sirolimus and Carcinoma--Squamous-Cell

While the majority of cutaneous squamous cell carcinomas (cSCCs) can be treated surgically, the additional work-up and treatments indicated for high-risk cSCC remain undefined. In recent years, improvements in tumor staging systems have allowed for the more accurate stratification of tumors into high- and low-risk categories. This insight, along with the publication of cSCC guidelines, brings us closer to the development of a consensus approach. The second article in this continuing medical education series addresses in question and answer format the most common questions related to advanced and high-stage cSCCs, with a simplified flowchart. The questions include the following: 1) Does my patient have high-risk cSCC?; 2) What is the next step for patients with cSCC and palpable lymphadenopathy?; 3) In patients with no clinically evident lymphadenopathy, who are candidates for lymph node staging?; 4) What forms of radiologic imaging can help detect subclinical lymph node metastases?; 5) What is the role of sentinel lymph node biopsy in cSCC?; 6) Which patients with cSCC need adjuvant radiation therapy?; 7) Is adjuvant chemotherapy an option for patients with high-stage cSCC after surgery?; 8) Are targeted and immunologic therapies an option for advanced cSCC?; 9) How often should I follow up with my patient after he/she has been diagnosed with a high-risk cSCC?; 10) What are the options for chemoprophylaxis in a patient with an increased risk of cSCC?; and 11) What chemopreventive measures can be started in coordination with medical oncology or transplant physicians?

Topics: Antimetabolites, Antineoplastic; Antineoplastic Agents, Immunological; Capecitabine; Carcinoma, Squamous Cell; Chemoprevention; Chemotherapy, Adjuvant; ErbB Receptors; Humans; Immunosuppression Therapy; Immunosuppressive Agents; Lymph Nodes; Lymphatic Metastasis; Molecular Targeted Therapy; Neoplasm Staging; Neoplasms, Multiple Primary; Organ Transplantation; Programmed Cell Death 1 Receptor; Radiotherapy, Adjuvant; Sirolimus; Skin Neoplasms

Patients with squamous cell carcinoma of the head and neck (SCCHN) typically present with locally advanced (LA) stage III or IV disease and are treated with combined-modality therapy with chemotherapy, radiotherapy, and surgery (if resectable). These aggressive, upfront treatment measures are often associated with substantial morbidity, and about half the patients develop locoregional or distant recurrences. Thus, new therapeutic strategies are needed that offer similar efficacy benefits with less toxicity. Current research is focused on selectively targeting signaling pathways involved in the proliferation and malignant transformation of SCCHN cells and the tumor microenvironment. For example, the ErbB receptor pathway has been implicated in the development and progression of SCCHN, and several agents targeting this pathway and downstream effectors are in various phases of clinical investigation. Cetuximab, a monoclonal antibody against epidermal growth factor receptor (EGFR), is the only currently approved targeted therapy for the treatment of LA SCCHN. Additional agents targeting EGFR and other ErbB family members, including monoclonal antibodies (eg, panitumumab, nimotuzumab) and small-molecule tyrosine kinase inhibitors (eg, erlotinib, afatinib, lapatinib) are being studied in LA SCCHN with varying results. Other treatment strategies for LA SCCHN include targeting downstream effectors of signaling and resistance mechanisms to EGFR inhibitors (eg, mammalian target of rapamycin, Src family, and Aurora kinase family). Data from ongoing and future clinical trials will continue to refine current treatment paradigms for LA SCCHN and provide new therapeutic options and potential predictive biomarkers to improve patient efficacy and safety and abrogate resistance.

Topics: Afatinib; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Azepines; Carcinoma, Squamous Cell; Cetuximab; Dasatinib; ErbB Receptors; Erlotinib Hydrochloride; Head and Neck Neoplasms; Humans; Lapatinib; Molecular Targeted Therapy; Panitumumab; Protein Kinase Inhibitors; Pyrimidines; Quinazolines; Quinazolinones; Signal Transduction; Sirolimus

The incidence of cutaneous squamous cell carcinomas (SCCs) in immunosuppressed solid organ transplant recipients (SOTRs) is 65- to 250-fold greater than in the general population. In addition, SCC in SOTRs is more aggressive than in the general population. SOTRs must undergo skin cancer screenings at intervals based on their risk stratification. The incidence of SCC in SOTRs varies with the type, intensity, and duration of the immunosuppressive regimen. Notably, patients on sirolimus have lower incidence of SCC compared to patients on calcineurin inhibitors. Revision of immunosuppressive regimen to include sirolimus may be a viable preventative measure against SCC in SOTRs who are high at risk for developing SCCs. Retinoids are also emerging as a means of chemoprophylaxis against development of new SCCs in high-risk patients. Treatments of SCC include electrodesiccation and curettage, surgical resection, cryosurgery, radiation, and systemic chemotherapy such as 5-fluorouracil and cetuximab.

Topics: Carcinoma, Squamous Cell; Cryosurgery; Fluorouracil; Humans; Immunosuppressive Agents; Microsurgery; Organ Transplantation; Photochemotherapy; Radiotherapy; Retinoids; Sentinel Lymph Node Biopsy; Sirolimus; Skin Neoplasms; Transplant Recipients

Topics: Carcinoma, Squamous Cell; Chemoradiotherapy; Humans; Immunosuppressive Agents; Kidney Transplantation; Laryngeal Neoplasms; Male; Middle Aged; Radiation-Sensitizing Agents; Sirolimus; TOR Serine-Threonine Kinases

Over the past two decades, advances in the fields of cancer genetics and molecular biology have elucidated molecular pathways that cause numerous cutaneous malignancies. This in turn has spurred the rational design of molecularly targeted therapies. In this review, we discuss the molecular pathways critical to the development of nonmelanoma skin cancers and the novel pharmacologic agents that target them. Included is a review of vismodegib for basal cell carcinoma, cetuximab for squamous cell carcinomas, imatinib for dermatofibrosarcoma protuberans, and sirolimus for Kaposi's sarcoma.

Topics: Anilides; Antibiotics, Antineoplastic; Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Benzamides; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cetuximab; Dermatofibrosarcoma; Humans; Imatinib Mesylate; Piperazines; Pyridines; Pyrimidines; Sarcoma, Kaposi; Sirolimus; Skin Neoplasms

Head and neck squamous cell carcinomas (HNSCC) represent 6% of all cancers diagnosed each year in the United States, affecting approximately 43,000 new patients and resulting in approximately 12,000 deaths. Currently, three main rapalogs exist for the treatment of cancer: CCI-779 (temsirolimus), RAD001 (everolimus), and AP235373 (deforolimus). Clinicians managing HNSCC need to be aware of the three rapalogs. Extensive evidence has shown rapamycin-analogs to be effective agents in the treatment of a number of solid tumors. While extensive preclinical data suggests that HNSCC would be an appropriate tumor type to benefit from inhibition of the mTOR pathway, limited clinical data is yet available to support this. Numerous phase II trials evaluating mTOR inhibitors for use in HNSCC are currently recruiting patients.

Topics: Antineoplastic Agents; Carcinoma, Squamous Cell; Clinical Trials as Topic; Everolimus; Female; Head and Neck Neoplasms; Humans; Male; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; United States

Patients with organ transplants have a significantly increased risk of skin cancer, especially squamous cell carcinoma, as a result of long-term treatment with immunosuppressive drugs. This paper provides a brief overview of the assumed underlying mechanisms.. The paper builds on relevant articles and studies identified in the course of many years of interest in immunopharmacology and skin cancer after organ transplantation.. Reduced immunological tumour surveillance as a result of chronic immunosuppression has long been assumed to underlie the increased risk of skin cancer after organ transplants. Recent studies indicate that immunosuppressive drugs may also have specific carcinogenic effects. Aziatropine, which inhibits proliferation of lymphocytes, increases oxidative DNA damage caused by UV radiation. Ciclosporin and tacrolimus, which have an immunosuppressive effect by inhibiting calcineurin, promote malignant phenotypes in cell culture and tumour growth in mouse models. Calcineurin has proved to be necessary in order for p53 protein to have a protective effect against skin cancer. A relatively new class of immunosuppressive drugs, mTOR inhibitors, have antineoplastic properties and are associated with less risk of skin cancer. A number of randomised studies are currently in progress to see whether mTOR inhibitors can reduce the risk of skin cancer after organ transplantation.. Immunosuppressive drugs contribute to skin cancer after organ transplantation, either as a result of immunosuppression or through specific carcinogenic mechanisms. Immunosuppressive drugs with antineoplastic properties are now starting to be used.

Topics: Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Humans; Immunosuppressive Agents; Organ Transplantation; Risk Factors; Sirolimus; Skin Neoplasms; Time Factors; TOR Serine-Threonine Kinases; Ultraviolet Rays

Whether sirolimus is useful in the prevention of non-melanoma skin cancer (NMSC) remains unclear and we therefore performed this meta-analysis of randomized controlled trials to test the hypothesis that Sirolimus-based immunosuppression is associated with a decrease in NMSC.. The main outcomes were NMSC, squamous-cell carcinoma and basal-cell carcinoma. The pooled risk ratio (RR) with its 95% confidence interval (95%CI) were used to assess the effects.. 5 randomized trials involving a total of 1499 patients receiving kidney transplantation were included. Patients undergoing Sirolimus-based immunosuppression had much lower risk of NMSC (RR = 0.49, 95%CI 0.32-0.76, P = 0.001). Subgroup analyses by tumor type showed that Sirolimus-based immunosuppression significantly decreased risk of both squamous-cell carcinoma (RR = 0.58, 95%CI 0.43-0.78, P < 0.001) and basal-cell carcinoma (RR = 0.56, 95%CI 0.37-0.85, P = 0.006). The quality of evidence was high for NMSC, and moderate for squamous-cell carcinoma and basal-cell carcinoma. No evidence of publication bias was observed.. High quality evidence suggests that Sirolimus-based immunosuppression decreases risk of non-melanoma skin cancer, and Sirolimus has an antitumoral effect among kidney-transplant recipients.

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Humans; Immunosuppression Therapy; Immunosuppressive Agents; Kidney Transplantation; Sirolimus; Skin Neoplasms

Topics: Abatacept; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Desensitization, Immunologic; Graft Rejection; Hand Disinfection; Heart Failure; HLA Antigens; Humans; Immunoconjugates; Immunomodulation; Immunosuppressive Agents; Kidney Transplantation; Living Donors; Mesenchymal Stem Cell Transplantation; Opportunistic Infections; Peritoneal Dialysis; Plasmapheresis; Postoperative Complications; Prognosis; Renal Dialysis; Renal Insufficiency; Sirolimus; Skin Neoplasms

Skin cancers are common in organ transplant recipients (OTRs). In this review, we discuss the epidemiology of and risk factors for cutaneous neoplasms, particularly squamous cell carcinoma (SCC) in OTRs. The pathogenesis of SCC is reviewed, as well as the potential mechanisms for tumor progression and metastasis associated with two commonly used immunosuppressive medications: tacrolimus and cyclosporine. Finally, we discuss the mechanism of action and potential preventative use of sirolimus, a member of a newer class of immunosuppressants, the mammalian target of rapamycin inhibitors. The authors have indicated no significant interest with commercial supporters.

Topics: Carcinoma, Squamous Cell; Humans; Immunosuppressive Agents; Organ Transplantation; Sirolimus; Skin Neoplasms

Skin cancer is the most frequent malignancy in organ transplant recipients, 95% of which are nonmelanoma skin cancer, especially squamous cell and basal cell carcinomas. This paper also discusses the incidence of other tumors (eg, melanoma, Merkel cell carcinoma, and Kaposi sarcoma) that are also increased in organ transplant patients compared to the general population. Part I of this two-part series describes the latest data concerning the epidemiologic and pathogenic aspects of nonmelanoma skin cancer development in solid organ transplant recipients. This review also highlights the concept of "field cancerization," represented by extensive areas of actinic damage and epidermal dysplasia, which accounts for increased risk of aggressive skin cancer development in susceptible patients.

Topics: Carcinoma, Basal Cell; Carcinoma, Merkel Cell; Carcinoma, Squamous Cell; Education, Medical, Continuing; Female; Humans; Immunocompromised Host; Incidence; Male; Melanoma; Organ Transplantation; Prognosis; Pyrimidines; Risk Assessment; Sarcoma, Kaposi; Sirolimus; Skin Neoplasms; Survival Analysis; Treatment Outcome; Triazoles; Voriconazole

Topics: Aurora Kinases; Carcinoma, Squamous Cell; Dasatinib; Enzyme Inhibitors; Focal Adhesion Protein-Tyrosine Kinases; Head and Neck Neoplasms; Humans; Protein Serine-Threonine Kinases; Pyrimidines; Sirolimus; src-Family Kinases; Thiazoles

Recently, new targets have been identified in head and neck squamous cell carcinomas (HNSCC) as playing key roles in tumour proliferation and metastases. The first target that has led to the approval of a molecularly based therapy in HNSCC has been the epidermal growth factor receptor (EGFR). Indeed, cetuximab, a monoclonal antibody directed against EGFR, has recently been approved in combination with radiation therapy in patients with locally advanced HNSCC, and in patients with platinum-refractory recurrent or metastatic (R/M) HNSCC. This review discusses novel targeted anticancer agents that do not exclusively target EGFR. The initial assessments of novel agents have typically been in patients with heavily pre-treated R/M HNSCC, with response rates and times to progression that are often disappointing. Evaluation of novel agents in the pre-operative 'window' setting, or as first-line therapy for R/M disease, may offer a more optimal understanding of their molecular and clinical effects.

Topics: Angiogenesis Inhibitors; Antibodies, Monoclonal; Carcinoma, Squamous Cell; ErbB Receptors; Farnesyltranstransferase; Head and Neck Neoplasms; Humans; Nuclear Proteins; Protein Kinase C; Protein Kinase Inhibitors; Receptor Protein-Tyrosine Kinases; Sirolimus

Topics: Carcinoma, Squamous Cell; Chemoprevention; Double-Blind Method; Humans; Organ Transplantation; Pilot Projects; Sirolimus; Skin Neoplasms; Transplant Recipients

Purpose Transplant recipients who develop cutaneous squamous cell carcinomas are at high risk for multiple subsequent skin cancers. Sirolimus has been shown to reduce the occurrence of secondary skin cancers, but no study included a follow-up exceeding 2 years. We extended at 5 years the TUMORAPA randomized trial of sirolimus-based immunosuppressive regimen versus calcineurin inhibitor-based immunosuppression. Methods Kidney transplant recipients receiving calcineurin inhibitors who had at least one cutaneous squamous cell carcinoma were randomly assigned to receive sirolimus as a substitute for calcineurin inhibitors (n = 64) or to maintain their initial treatment (n = 56). The primary end point was survival free of squamous cell carcinoma at 5 years. Secondary end points included the occurrence of other skin cancers, renal function, patient and graft survival, and treatment tolerance. Results Survival free of cutaneous squamous cell carcinoma was significantly longer in the sirolimus group than in the calcineurin inhibitor group ( P = .007). In the sirolimus group, the number of patients with new skin cancers was significantly lower compared with the calcineurin inhibitor group: 22% versus 59% for squamous cell carcinomas ( P < .001), 34% versus 66% for other skin cancers ( P < .001), and 20% versus 37.5% for basal cell carcinomas ( P < .05). Kidney graft function, patients, and graft survival were similar in both groups. In the sirolimus group, the mean number of serious adverse effects per patient decreased from 1.16 during the first 2 years, to 0.83 between years 2 and 5. Conclusion In kidney transplant recipients with previous cutaneous squamous cell carcinomas, the antitumoral effect of conversion from calcineurin inhibitors to sirolimus was maintained at 5 years, and sirolimus tolerance was satisfactory.

Topics: Calcineurin Inhibitors; Carcinoma, Squamous Cell; Humans; Immunocompromised Host; Immunosuppressive Agents; Kidney Transplantation; Secondary Prevention; Sirolimus; Skin Neoplasms; Transplant Recipients

Activating events along the PI3K/mTOR pathway are common in head and neck squamous cell carcinomas (HNSCC), and preclinical studies suggest additive or synergistic effects when combining mTORC1 inhibitors with carboplatin and paclitaxel chemotherapy.. In this single-institution phase II study, the combination of temsirolimus 25 mg, carboplatin AUC 1.5, and paclitaxel 80 mg/m2 administered on days 1 and 8 of a 21-day cycle was evaluated in 36 patients with recurrent and/or metastatic (R/M) HNSCC. The primary end point was objective response rate after two cycles of treatment. Secondary end points include the safety and tolerability profile and overall survival. Correlative studies with exome mutational analysis were performed in pre-treatment biopsy samples from 21 patients.. Fifteen (41.7%) patients had an objective response, which were all partial responses, and 19 (52.3%) patients had stable disease as best response. The two patients who were designated as 'non-responders' were removed from study prior to two cycles of treatment, but are included in the efficacy and safety analyses. The median duration on study was 5.3 months and the median progression-free survival and overall survival were 5.9 months (95% confidence interval, 4.8-7.1) and 12.8 months (95% confidence interval, 9.8-15.8), respectively. The most common grade 3 and 4 adverse events were hematologic toxicities. Three (3.8%) patients developed neutropenic fever on study. Three of four patients with PIK3CA mutations experienced tumor regressions, and responses were also seen in patients with other genetic alterations in the PI3K/mTOR pathway.. The combination of temsirolimus with low-dose weekly carboplatin and paclitaxel appears to have meaningful clinical efficacy in the treatment of R/M HNSCC. This regimen has a relatively high response rate compared to other treatments evaluated in R/M HNSCC, and potential associations with genetic alterations in the PI3K/mTOR pathway should be further explored.

Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Carboplatin; Carcinoma, Squamous Cell; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Head and Neck Neoplasms; Humans; Male; Middle Aged; Neoplasm Metastasis; Neoplasm Recurrence, Local; Paclitaxel; Sirolimus; Squamous Cell Carcinoma of Head and Neck

Squamous cell carcinoma of the head and neck (SCCHN) is a common disease, which has a poor prognosis after failure of therapy. Activation of the PI3K-AKT-mTOR axis is commonly detected in recurrent or metastatic SCCHN, and provided the rationale for the clinical phase II trial in pretreated SCCHN.. The primary end point was the progression-free survival rate (PFR) at 12 weeks. Forty eligible patients have been recruited after failure of platinum chemotherapy and cetuximab. A preplanned futility analysis was successfully passed after ≥1 success was detected in 20 patients. Secondary objectives consisted of progression-free survival (PFS), disease control rate (DCR), overall survival (OS), safety and tolerability, and predictive biomarkers for KRAS, BRAF, PIK3CA mutations, and HPV status. Archived tumor tissue was analyzed for DNA sequence.. A total of 40 patients were eligible. The PFR at 12 weeks was 40% (95% CI 25.0-54.6). The median PFS and OS were 56 days (95% CI 36-113 days) and 152 days (76-256 days), respectively. In 33 assessable patients, disease stabilization occurred in 57.6%, with tumor shrinkage in 13 patients (39.4%). Overall, the treatment was well tolerated. Fatigue (47.5%), anemia (25.0%), nausea (20.0%), and pneumonia (20.0%) were the most common adverse events. Neither PIK3CA mutations, nor HPV status were predictive for success with temsirolimus treatment. No mutations were found for KRAS or BRAF.. Tumor shrinkage and efficacy parameter indicate that inhibition of the PI3K-AKT-mTOR axis was a putative novel treatment paradigm for SCCHN. We could not identify parameters predictive for treatment success of temsirolimus, which underscores the need for refinement of the molecular analysis in future studies.. NCT01172769.

Topics: Adult; Aged; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cetuximab; Cisplatin; Disease-Free Survival; Female; Germany; Head and Neck Neoplasms; Humans; Male; Middle Aged; Neoplasm Recurrence, Local; Sirolimus; Squamous Cell Carcinoma of Head and Neck; Treatment Outcome

Squamous cell cancer of the head and neck (SCCHN) is a frequent aggressive malignancy with limited therapeutic options. Increasing evidence suggests that mammalian target of rapamycin (mTOR)-inhibitors might be effective in advanced SCCHN. However, non-invasive biomarkers for early prediction of treatment efficacy are not established in SCCHN. Highly proliferating tumours are characterised by enhanced cell turnover which is associated with enhanced apoptosis. During apoptosis of epithelial cells caspases cleave cytokeratin (CK)-18 can be detected in the blood. In this study we analysed sera from patients with relapsed or metastatic SCCHN patients who have been treated with temsirolimus for caspase-cleaved and total (caspase-cleaved and uncleaved) CK-18 by enzyme-linked immunosorbent assays (ELISAs). In addition, caspase-3 activity was detected by luminometric substrate assay. SCCHN patients revealed higher serum levels of those biomarkers compared to healthy controls. Importantly, patients with short progression-free survival (PFS) showed higher serum levels of caspase-3 activity compared to patients with longer PFS (⩾ 2months). Caspase-3 activity is inversely correlated with PFS. A cut-off value for caspase-3 activity was determined that correctly predicted PFS <2months with a sensitivity of 86% and a specificity of 67%. These data demonstrate that detection of serum caspase-3 activity might be a useful non-invasive biomarker for early identification of SCCHN patients not responding to treatment with novel targeted therapies such as mTOR-inhibitors.

Topics: Aged; Antineoplastic Agents; Biomarkers, Tumor; Carcinoma, Squamous Cell; Caspases; Disease-Free Survival; Enzyme-Linked Immunosorbent Assay; Female; Head and Neck Neoplasms; Humans; Male; Middle Aged; Predictive Value of Tests; Protein Kinase Inhibitors; Sirolimus; Squamous Cell Carcinoma of Head and Neck

The phosphatidylinositol 3-kinase/protein kinase-B/mammalian target of rapamycin (PI3K-AKT-mTOR) signalling pathway is aberrantly activated in several cancers. Notch signalling maintains cell proliferation, growth and metabolism in part by driving the PI3K pathway. Combining the mTOR inhibitor ridaforolimus with the Notch inhibitor MK-0752 may increase blockade of the PI3K pathway.. This phase I dose-escalation study (NCT01295632) aimed to define the dose-limiting toxicities (DLTs) and maximum tolerated dose (MTD) of combination oral ridaforolimus (rising doses starting at 20 mg, 5 days/week) and oral MK-0752 (1800 mg once weekly) in patients with solid tumours. No intrapatient dose escalation was permitted.. Twenty eight patients were treated on study. Ridaforolimus doses were escalated from 20 to 30 mg/day. Among 14 evaluable patients receiving ridaforolimus 20 mg, one DLT (grade 2 stomatitis, second episode) was reported. Among eight evaluable patients receiving ridaforolimus 30 mg, three DLTs were reported (one each grade 3 stomatitis, grade 3 diarrhoea, and grade 3 asthenia). The MTD was 20 mg daily ridaforolimus 5 days/week+1800 mg weekly MK-0752. The most common drug-related adverse events included stomatitis, diarrhoea, decreased appetite, hyperglycaemia, thrombocytopenia, asthenia and rash. Two of 15 (13%) patients with head and neck squamous cell carcinoma (HNSCC) had responses: one with complete response and one with partial response. In addition, one patient experienced stable disease ⩾6 months.. Combined ridaforolimus and MK-0752 showed activity in HNSCC. However, a high number of adverse events were reported at the MTD, which would require careful management during future clinical development.

Topics: Administration, Oral; Adult; Aged; Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Benzene Derivatives; Carcinoma, Squamous Cell; Drug Administration Schedule; Europe; Female; Head and Neck Neoplasms; Humans; Male; Maximum Tolerated Dose; Middle Aged; Molecular Targeted Therapy; Multimodal Imaging; Positron-Emission Tomography; Propionates; Protein Kinase Inhibitors; Signal Transduction; Sirolimus; Squamous Cell Carcinoma of Head and Neck; Sulfones; Time Factors; Tomography, X-Ray Computed; Treatment Outcome; United States

The addition of targeted agents to thoracic radiation has not improved outcomes in patients with locally advanced non-small-cell lung cancer (NSCLC). To improve cure rates in locally advanced NSCLC, effective targeted therapies need to be identified that can be given safely with radiation therapy. Temsirolimus is an inhibitor of the mammalian target of rapamycin (mTOR) pathway and has single-agent activity in lung cancer. Inhibition of the mTOR pathway has been found to augment the cytotoxic effect of radiation in preclinical studies. There is scant clinical experience with mTOR inhibitors and radiation.. This was a phase I study evaluating the combination of temsirolimus with thoracic radiation in patients with NSCLC.. Ten patients were enrolled in the study. The dose-limiting toxicities included sudden death, pneumonitis, and pulmonary hemorrhage. The maximum tolerated dose of temsirolimus that could be administered safely with concurrent radiotherapy (35 Gy in 14 daily fractions) was 15 mg intravenously weekly. Of the 8 evaluable patients, 3 had a partial response and 2 had stable disease.. The combination of temsirolimus 15 mg weekly and thoracic radiation is well tolerated and warrants further investigation, perhaps in a molecularly defined subset of patients.

Topics: Adenocarcinoma; Aged; Aged, 80 and over; Antineoplastic Agents; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Female; Follow-Up Studies; Humans; Lung Neoplasms; Male; Maximum Tolerated Dose; Middle Aged; Neoplasm Staging; Prognosis; Radiotherapy Dosage; Sirolimus; Survival Rate

Platinum-based therapy combined with cetuximab is standard first-line therapy for recurrent or metastatic squamous cell carcinoma of the head and neck (RMSCCHN). Preclinical studies have suggested that mammalian target of rapamycin inhibitors may overcome resistance to epidermal growth factor receptor blockers and may augment cetuximab antitumor activity. We conducted a phase 1b trial of carboplatin, cetuximab, and everolimus for untreated RMSCCHN.. Patients received carboplatin (area under the curve = 2 mg/ml/min; 3 weeks on, 1 week off), cetuximab (with a loading dose of 400 mg/m(2) and then 250 mg/m(2) weekly), and dose-escalating everolimus (2.5, 5.0, 7.5, and 10 mg/day) with a 3+3 design. After 4 cycles, patients without progression continued cetuximab/everolimus until progression or intolerable toxicity. Patients (age ≥ 18 years) had previously untreated, unresectable RMSCCHN not amenable to radiotherapy and an Eastern Cooperative Oncology Group performance status of 0 to 2.. The study enrolled 20 patients (male/female = 18/2) with RMSCCHN; the median age was 65 years (44-75 years). Thirteen patients received everolimus (male/female = 92%). Two of 6 patients receiving 2.5 mg/day experienced dose-limiting toxicity (DLT) with grade 3 hyponatremia and nausea. In 7 patients receiving de-escalated everolimus (2.5 mg every other day), grade 3 hyperglycemia produced DLT in 1 of 6 patients. The objective response rate (RR) was 61.5% (all partial responses). Progression-free survival (PFS) was 8.15 months. The pharmacokinetics of everolimus was described with a 2-compartment mixed-effects model. There was a significant correlation between tumor p-p44/42 staining and response (P = .044) and a marginally significant correlation between phosphorylated mammalian target of rapamycin and overall survival.. The maximum tolerated dose of everolimus with cetuximab and carboplatin was 2.5 mg every other day. The regimen was associated with an encouraging RR and PFS, and this suggested possible clinical efficacy in a select group of patients with squamous cell carcinoma of the head and neck.

Topics: Adult; Aged; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Carboplatin; Carcinoma, Squamous Cell; Cetuximab; Disease-Free Survival; Drug Administration Schedule; Everolimus; Female; Head and Neck Neoplasms; Humans; Male; Maximum Tolerated Dose; Middle Aged; Neoplasm Metastasis; Recurrence; Sirolimus; Squamous Cell Carcinoma of Head and Neck

We conducted a phase II study of docetaxel in combination with everolimus, a mammalian target of rapamycin (mTOR) inhibitor, for salvage therapy of advanced non-small-cell lung cancer (NSCLC) based on promising preclinical and early-phase clinical data. Patients with advanced-stage NSCLC treated with one or two previous systemic therapy regimens were given docetaxel (60 mg/m) and everolimus (5 mg orally once daily on days 1-19) every 3 weeks. Archived tumor specimens were evaluated for markers of mTOR pathway activation (total and phosphorylated mTOR, Akt, S6, eIF4e, and 4EBP1). Twenty-eight patients were enrolled (median age: 62 years; male: 13; Caucasians: 19; adenocarcinoma: 20; performance status 0, 3; performance status 1, 23; 1 previous regimen, 16). A median of 3.5 cycles of therapy was administered. Two patients experienced partial response and 15 had stable disease (clinical benefit rate, 70%). The 6-month progression-free survival rate was 5%, and the median overall survival was 9.6 months. Low pAkt expression correlated with clinical benefit rate (p = 0.01) but not with progression-free survival or overall survival. The combination of everolimus and docetaxel was tolerated well, but the efficacy was relatively modest in an unselected population of patients with NSCLC.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Biomarkers, Tumor; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Docetaxel; Everolimus; Female; Follow-Up Studies; Humans; Lung Neoplasms; Male; Middle Aged; Neoplasm Staging; Prognosis; Sirolimus; Survival Rate; Taxoids; TOR Serine-Threonine Kinases

Elevated expression of eukaryotic protein synthesis initiation factor 4E (eIF4E) in histologically cancer-free margins of resected head and neck squamous cell carcinomas (HNSCCs) is mediated by mammalian target of rapamycin complex 1 (mTORC1) and has been associated with increased risk of disease recurrence. Preclinically, inhibition of mTORC1 with everolimus sensitizes cancer cells to cisplatin and radiation.. This was single-institution phase 1 study to establish the maximum tolerated dose of daily everolimus given with fixed dose cisplatin (30 mg/m(2) weekly × 6) and concurrent intensity modulated radiation therapy for patients with locally and/or regionally advanced head-and-neck cancer. The study had a standard 3 + 3 dose-escalation design.. Tumor primary sites were oral cavity (4), salivary gland (4), oropharynx (2), nasopharynx (1), scalp (1), and neck node with occult primary (1). In 4 of 4 cases in which resected HNSCC surgical pathology specimens were available for immunohistochemistry, elevated expression of eIF4E was observed in the cancer-free margins. The most common grade ≥3 treatment-related adverse event was lymphopenia (92%), and dose-limiting toxicities (DLTs) were mucositis (n=2) and failure to thrive (n=1). With a median follow up of 19.4 months, 2 patients have experienced recurrent disease. The maximum tolerated dose was everolimus 5 mg/day.. Head-and-neck cancer patients tolerated everolimus at therapeutic doses (5 mg/day) given with weekly cisplatin and intensity modulated radiation therapy. The regimen merits further evaluation, especially among patients who are status post resection of HNSCCs that harbor mTORC1-mediated activation of eIF4E in histologically negative surgical margins.

Topics: Administration, Oral; Adult; Aged; Antineoplastic Agents; Carcinoma, Squamous Cell; Chemoradiotherapy; Cisplatin; Drug Administration Schedule; Eukaryotic Initiation Factor-4E; Everolimus; Female; Head and Neck Neoplasms; Humans; Injections, Intravenous; Male; Maximum Tolerated Dose; Middle Aged; Neoplasm Proteins; Prospective Studies; Radiation-Sensitizing Agents; Radiotherapy Dosage; Radiotherapy, Intensity-Modulated; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases

Conversion to sirolimus from calcineurin inhibitor- (CNI), azathioprine- (AZA) and mycophenolate-based regimens reduces the risk of development of squamous cell carcinoma of the skin (SCC) in kidney transplant recipients (KTRs). Sirolimus conversion may also be protective by permitting beneficial changes in immune phenotype. It is not known how sirolimus will affect immune phenotype in KTRs with SCC.. Thirty-two KTRs with SCC were enrolled into this single-blinded randomized study and 13 KTRs randomized to sirolimus (4-10 ng/mL) and prednisolone 5 mg/day.. Six-month post conversion to sirolimus FOXP3(+) CD127(low)CD25(high)CD69(-), the number of T cells (putative Treg) increased significantly (P = 0.008). Natural killer (NK) and CD56(bright) NK cells also increased significantly (P = 0.039 and 0.02). T-cell number only significantly increased in those KTRs where CNI was ceased as part of the conversion to mammalian target of rapamycin inhibitors (mTORi's) (P = 0.031) implying CNI cessation rather than mTORi initiation induced an increase in T-cell number. Increases in the NK cell number was only significant in those KTRs where AZA was ceased (P = 0.040), implying AZA cessation rather than mTORi initiation caused the NK cell number to increase. At 6 months, sirolimus conversion reduces new SCC/year, rate ratio 0.49 (95%CI: 0.15-1.63), P = 0.276. On therapy analysis and intention-to-treat analysis over 24 months, the rate ratios were 0.84 and 0.87, respectively, and did not reach significance.. Conversion to mTORi from CNI may reveal a pre-existing high Treg phenotype by unmasking CNI inhibition of FOXP3 expression. Cessation of AZA leads to increased NK cell number. High FOXP3(+) T-cell number on conversion to mTORi may predict those KTRs who continue to accrue SCC.

Topics: Aged; Aged, 80 and over; Azathioprine; Calcineurin Inhibitors; Carcinoma, Squamous Cell; CD56 Antigen; Female; Forkhead Transcription Factors; Humans; Immune System; Immunosuppressive Agents; Kidney Transplantation; Killer Cells, Natural; Male; Middle Aged; Phenotype; Risk Factors; Single-Blind Method; Sirolimus; Skin Neoplasms; T-Lymphocytes; TOR Serine-Threonine Kinases; Transplantation

In light of the significant morbidity and mortality of cutaneous invasive squamous cell carcinomas (SCCs) in renal transplant recipients, we investigated whether conversion to sirolimus-based immunosuppression from standard immunosuppression could diminish the recurrence rate of these skin cancers.. In a 2-year randomized controlled trial, 155 renal transplant recipients with at least one biopsy-confirmed SCC were stratified according to age (< 55 v ≥ 55 years) and number of previous SCCs (one to nine v ≥ 10) and randomly assigned to conversion to sirolimus (n = 74) or continuation of their original immunosuppression (n = 81). Development of a new SCC within 2 years after random assignment was the primary end point.. After 2 years of follow-up, the risk reduction of new SCCs in the multivariable analysis was not significant, with a hazard ratio (HR) of 0.76 (95% CI, 0.48 to 1.2; P = .255), compared with a non-sirolimus-based regimen. After the first year, there was a significant 50% risk reduction, with an HR of 0.50 (95% CI, 0.28 to 0.90; P = .021) for all patients together and an HR of 0.11 (95% CI, 0.01 to 0.94; P = .044) for patients with only one previous SCC. The tumor burden of SCC was reduced during the 2-year follow-up period in those receiving sirolimus (0.82 v 1.38 per year; HR, 0.51; 95% CI, 0.32 to 0.82; P = .006) if adjusted for the number of previous SCCs and age. Twenty-nine patients stopped taking sirolimus because of various adverse events.. Conversion to sirolimus-based immunosuppression failed to show a benefit in terms of SCC-free survival at 2 years.

Topics: Carcinoma, Squamous Cell; Diarrhea; Female; Follow-Up Studies; Humans; Immunosuppressive Agents; Kaplan-Meier Estimate; Kidney Transplantation; Male; Middle Aged; Multivariate Analysis; Neoplasm Invasiveness; Prospective Studies; Respiratory Tract Infections; Sirolimus; Skin Neoplasms; Time Factors; Treatment Outcome; Tumor Burden

The epidermal growth factor receptor (EGFR) is a validated target in head and neck squamous cell carcinoma (HNSCC). In recurrent and/or metastatic (R/M) HNSCC, resistance to anti-EGFR therapy inevitably occurs. Downstream activation of the PI3K/Akt/mTOR pathway is an established resistance mechanism. Concurrent mTOR blockade may improve efficacy of anti-EGFR therapy.. Erlotinib 150 mg daily and temsirolimus 15 mg weekly were administered to patients with platinum-refractory R/M HNSCC and ECOG performance status 0-2. The primary endpoint was progression-free survival (PFS). Correlative studies determined PIK3CA and HRAS mutation status; p16, EGFR, pS6K, pAkt and PTEN expression; and pre- and post-treatment plasma levels of 20 immunomodulatory cytokines.. Twelve patients enrolled; six withdrew within 6 weeks due to toxicity or death, prompting early closure of the trial. Grade ≥ 3 toxicities included fatigue, diarrhea, gastrostomy tube infection, peritonitis, pneumonia, dyspnea, and HN edema. Median PFS was 1.9 months. Median overall survival was 4.0 months. Six/12 tumors were p16(+), 9/11 lacked measurable PTEN expression, and 1/12 harbored a PIK3CA mutation. On exploratory analysis, high baseline plasma VEGF and interferon-gamma levels marginally associated with tumor progression.. The combination of erlotinib and temsirolimus was poorly tolerated. Low prevalence of PTEN expression and 8% incidence of PIK3CA mutations indicate biological relevance of this pathway in R/M disease. Investigation of more tolerable combinations of EGFR and PI3K/Akt/mTOR pathway inhibitors in selected HNSCC patients is warranted.

Topics: Aged; Aged, 80 and over; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Class I Phosphatidylinositol 3-Kinases; Cyclin-Dependent Kinase Inhibitor p16; Cytokines; Drug Resistance, Neoplasm; ErbB Receptors; Erlotinib Hydrochloride; Female; Head and Neck Neoplasms; Humans; Male; Middle Aged; Mutation; Neoplasm Recurrence, Local; Oncogene Protein v-akt; Phosphatidylinositol 3-Kinases; Platinum; Protein Kinase Inhibitors; Proto-Oncogene Proteins p21(ras); PTEN Phosphohydrolase; Quinazolines; Ribosomal Protein S6 Kinases; Sirolimus; Survival Rate; TOR Serine-Threonine Kinases; Tumor Suppressor Proteins

Sirolimus has antineoplastic effects and may reduce skin cancer rates in kidney transplant patients. This prospective, multicenter, randomized, open-label, controlled trial randomized 86 kidney transplant recipients (≥1 year posttransplant) with history of nonmelanoma skin cancer (NMSC) to continue calcineurin inhibitor (CNI) or convert to sirolimus. Patients were stratified by number of NMSC lesions (0-5, 6-20) in previous year. Primary end point was number of biopsy-confirmed new NMSC lesions per patient-year. Yearly NMSC rate was significantly lower with sirolimus (1.31 vs. 2.48 lesions/patient-year; p = 0.022). Squamous cell carcinoma occurred at a lower rate in the sirolimus versus CNI group (p = 0.038); basal cell carcinoma rate was similar in both. A lower proportion of patients receiving sirolimus developed new or recurrent NMSC (56.4% vs. 80.9%; p = 0.015) or new squamous cell carcinoma (41.0% vs. 70.2%; p = 0.006). No sirolimus patients and one CNI continuation patient experienced acute rejection. Incidence of treatment-emergent adverse events was similar between groups; however, discontinuation rates related to adverse events were significantly higher with sirolimus (46.2% vs. 0%; p < 0.001). In kidney transplant recipients with history of NMSC, conversion from CNI to sirolimus reduced rates of NMSC, without increasing acute rejection risk.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Female; Follow-Up Studies; Graft Rejection; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Prognosis; Prospective Studies; Risk Factors; Sirolimus; Skin Neoplasms; Survival Rate

The mammalian target of rapamycin complex 1 (mTORC1) is aberrantly activated in many head and neck squamous cell carcinomas (HNSCCs). This phase I study combines the mTORC1 inhibitor temsirolimus with carboplatin and paclitaxel.. This was a single institution phase I study for patients with R/M HNSCC with a standard 3 + 3 design. Three doses of temsirolimus were planned: 15, 20, and 25 mg. Due to excessive toxicity with the original study regimen, the protocol was amended to carboplatin AUC 1.5, paclitaxel 80 mg/m(2), and temsirolimus (according to dose escalation plan), all on days 1 and 8 of a 21-day cycle.. 18 patients (14 male, 4 female) enrolled, with median age 56 years (range 33-78). The most common toxicities were anemia, leukopenia, thrombocytopenia, and hyperglycemia. Among all patients treated, the confirmed objective partial response (cPR) rate was 22 %. DLT was not exceeded among 6 patients treated at dose level 3 of the revised protocol, and 4 of 6 subjects treated at this dose level had cPRs.. The phase II recommended regimen is temsirolimus 25 mg, carboplatin AUC 1.5, and paclitaxel 80 mg/m(2), all on days 1 and 8 of a 21-day cycle. A phase II study of this regimen in R/M HNSCC is ongoing.

Topics: Adult; Aged; Anemia; Antineoplastic Combined Chemotherapy Protocols; Carboplatin; Carcinoma, Squamous Cell; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Head and Neck Neoplasms; Humans; Hyperglycemia; Leukopenia; Male; Middle Aged; Neoplasm Metastasis; Neoplasm Recurrence, Local; Paclitaxel; Sirolimus; Thrombocytopenia; Treatment Outcome

Transplant recipients in whom cutaneous squamous-cell carcinomas develop are at high risk for multiple subsequent skin cancers. Whether sirolimus is useful in the prevention of secondary skin cancer has not been assessed.. In this multicenter trial, we randomly assigned transplant recipients who were taking calcineurin inhibitors and had at least one cutaneous squamous-cell carcinoma either to receive sirolimus as a substitute for calcineurin inhibitors (in 64 patients) or to maintain their initial treatment (in 56). The primary end point was survival free of squamous-cell carcinoma at 2 years. Secondary end points included the time until the onset of new squamous-cell carcinomas, occurrence of other skin tumors, graft function, and problems with sirolimus.. Survival free of cutaneous squamous-cell carcinoma was significantly longer in the sirolimus group than in the calcineurin-inhibitor group. Overall, new squamous-cell carcinomas developed in 14 patients (22%) in the sirolimus group (6 after withdrawal of sirolimus) and in 22 (39%) in the calcineurin-inhibitor group (median time until onset, 15 vs. 7 months; P=0.02), with a relative risk in the sirolimus group of 0.56 (95% confidence interval, 0.32 to 0.98). There were 60 serious adverse events in the sirolimus group, as compared with 14 such events in the calcineurin-inhibitor group (average, 0.938 vs. 0.250). There were twice as many serious adverse events in patients who had been converted to sirolimus with rapid protocols as in those with progressive protocols. In the sirolimus group, 23% of patients discontinued the drug because of adverse events. Graft function remained stable in the two study groups.. Switching from calcineurin inhibitors to sirolimus had an antitumoral effect among kidney-transplant recipients with previous squamous-cell carcinoma. These observations may have implications concerning immunosuppressive treatment of patients with cutaneous squamous-cell carcinomas. (Funded by Hospices Civils de Lyon and others; TUMORAPA ClinicalTrials.gov number, NCT00133887.).

Topics: Adult; Aged; Aged, 80 and over; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Cyclosporine; Enzyme Inhibitors; Female; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Sirolimus; Skin Neoplasms; Tacrolimus

The epidermal growth factor receptor inhibitor erlotinib is an approved treatment for chemotherapy-refractory advanced non-small-cell lung cancer (NSCLC). Because activated epidermal growth factor receptor signals through the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (mTOR) pathway, adding the oral mTOR inhibitor everolimus to erlotinib may improve efficacy by blocking multiple components of the same pathway. We conducted a phase I study to determine feasible dosages of combination therapy with erlotinib and everolimus for previously treated metastatic or unresectable NSCLC.. Participants had advanced NSCLC progressing after two or less previous chemotherapy regimens. Feasibility of daily/weekly everolimus plus daily erlotinib was determined using a 6 + 6 dose-escalation design based on the rate of dose-limiting toxicities. Antitumor activity was assessed by the Response Evaluation Criteria In Solid Tumors study.. Of the 94 patients enrolled, 90% had stage IV NSCLC, 19% never smoked, and 15% were current smokers. Eighty-nine patients experienced one or more adverse events possibly related to any study medication. The most common dose-limiting toxicities were stomatitis (n = 5), rash (n = 4), and diarrhea (n = 3). Maximum tolerated doses were everolimus 5 mg per day or 50 mg per week plus erlotinib 150 mg per day. In daily everolimus cohorts (n = 74), nine patients achieved a complete/partial response and 28 had stable disease (median duration disease control, 9.3 months). In weekly everolimus cohorts (n = 20), no tumor response was observed; seven patients had stable disease (median duration, 9.6 months).. Combination therapy with everolimus 5 mg per day or 50 mg per week and erlotinib 150 mg per day provided acceptable tolerability and disease control. A randomized phase II study evaluating this combination in comparison with erlotinib alone is complete and is being analyzed.

Topics: Adenocarcinoma; Adenocarcinoma, Bronchiolo-Alveolar; Adult; Aged; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Large Cell; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Cohort Studies; Erlotinib Hydrochloride; Everolimus; Feasibility Studies; Female; Follow-Up Studies; Humans; Lung Neoplasms; Male; Maximum Tolerated Dose; Middle Aged; Neoplasm Staging; Prognosis; Quinazolines; Sirolimus; Tissue Distribution

Activation of the mammalian target of rapamycin (mTOR) pathway in surgical margins of head and neck squamous cell carcinoma (HNSCC) is a predictor of recurrence and patients with minimal residual disease may benefit from adjuvant therapy with temsirolimus, an mTOR inhibitor.. The effects of 3 weekly doses of 25 mg of temsirolimus on Akt/mTOR pathway biomarkers were evaluated in tumor and peripheral blood mononuclear cells (PBMCs) of patients with HNSCC. Adverse events were assessed.. Temsirolimus significantly decreased pS6 and p4E-BP1 in tumors, and pS6 and pAkt in PBMCs (p < .05). There was no significant upregulation of pAkt(Ser(473)) in tumor tissue. Side effects were minimal and reversible.. Significant inhibition of the mTOR pathway was noted in both tumors and PBMCs of HNSCC with minimal side effects. The mTOR inhibitors can potentially be used as adjuvant therapy for patients with minimal residual disease and PBMCs are potential surrogate markers in this setting.

Topics: Adult; Aged; Antineoplastic Agents; Biomarkers, Tumor; Carcinoma, Squamous Cell; Chemotherapy, Adjuvant; Female; Head and Neck Neoplasms; Humans; Male; Middle Aged; Protein Kinase Inhibitors; Sirolimus; TOR Serine-Threonine Kinases

Concurrent signal transduction inhibition with the epidermal growth factor receptor (EGFR) inhibitor gefitinib and the mammalian target-of-rapamycin inhibitor everolimus has been hypothesized to result in enhanced antitumor activity in patients with non-small cell lung cancer (NSCLC). This phase II trial assessed the efficacy of the combination of gefitinib and everolimus in patients with advanced NSCLC.. Two cohorts of 31 patients with measurable stage IIIB/IV NSCLC were enrolled: (1) no prior chemotherapy and (2) previously treated with cisplatin or carboplatin and docetaxel or pemetrexed. All patients received daily everolimus 5 mg and gefitinib 250 mg. Response was assessed after 1 month and then every 2 months. Pretreatment tumor specimens were collected for mutation testing.. Sixty-two patients were enrolled (median age: 66 years, 50% women, 98% stage IV, all current/former smokers, and 85% adenocarcinoma). Partial responses were seen in 8 of 62 patients (response rate: 13%; 95% confidence interval: 5-21%); five responders had received no prior chemotherapy. Three partial responders had an EGFR mutation. Both patients with a KRAS (G12F) mutation responded. The median time to progression was 4 months. Median overall survival was 12 months, 27 months for no prior chemotherapy patients, and 11 months for patients previously treated with chemotherapy.. The 13% partial response rate observed did not meet the prespecified response threshold to pursue further study of the combination of gefitinib and everolimus. The response rate in patients with non-EGFR mutant tumors was 8%, likely reflecting activity of everolimus. Further investigation of mammalian target-of-rapamycin inhibitors in patients with NSCLC with KRAS G12F-mutated tumors is warranted.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Clinical Trials, Phase I as Topic; Cohort Studies; ErbB Receptors; Everolimus; Female; Gefitinib; Humans; Lung Neoplasms; Male; Middle Aged; Mutation; Neoplasm Staging; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); Quinazolines; ras Proteins; Salvage Therapy; Sirolimus; Survival Rate; Treatment Outcome

Preclinical studies have demonstrated that the inhibition of the PI3K/Akt/mTOR pathway restores gefitinib sensitivity in resistant cancer cell lines. A phase 1 study was conducted of the combination of everolimus, an mTOR inhibitor, and gefitinib to determine a daily dose of everolimus with gefitinib in patients with advanced nonsmall-cell lung cancer (NSCLC).. Oral everolimus and gefitinib were both administered daily to patients with progressive NSCLC. Patients were enrolled in 3-patient cohorts at everolimus dose levels of 5 and 10 mg daily. All patients received gefitinib 250 mg daily.. Ten patients were enrolled. The maximum tolerated dose of everolimus was 5 mg when administered daily with gefitinib 250 mg. Two patients who were treated at the 10 mg dose level of everolimus experienced dose-limiting toxicity, including grade 5 hypotension and grade 3 stomatitis. Pharmacokinetic studies demonstrated no consistent, significant interaction on the t(max), C(max), and AUC(0-8h) of either agent. Two partial radiographic responses were identified among the 8 response-evaluable patients.. For further study, everolimus at a dose of 5 mg daily in combination with daily gefitinib 250 mg is recommended. The 2 radiographic responses identified are encouraging. A phase 2 trial in patients with NSCLC is under way.

Topics: Adenocarcinoma; Aged; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Dose-Response Relationship, Drug; Everolimus; Female; Gefitinib; Humans; Lung Neoplasms; Male; Middle Aged; Quinazolines; Sirolimus; Survival Rate; United States

Recent studies have reported a significant increase of proteinuria in kidney transplant recipients who were switched from a calcineurin inhibitor (CI) to sirolimus. This has (partly) been ascribed to the hemodynamic renal effects of CI withdrawal. We have evaluated the evolution of proteinuria in renal transplant recipients who underwent conversion from azathioprine to sirolimus. In a randomized, prospective, multicenter study called RESCUE (Recurrent cutanEous Squamous cell Carcinoma Under RapamunE) the efficacy and safety is investigated of conversion to sirolimus in stable renal transplant recipients with a cutaneus squamous cell carcinoma (SCC). In our center 25 patients have been included in this study of which 13 patients were randomized to continue their current immunosuppressive treatment and 12 to conversion to sirolimus. After a mean follow-up of 360 days mean proteinuria increased from 0.37+/-0.34 to 1.81+/-1.73 g/24 h after conversion to sirolimus (P<0.005). In the control group there was no change in proteinuria. A significant increase of proteinuria was observed in all seven patients with proteinuria before conversion, whereas proteinuria remained absent in all patients without previous proteinuria. Two of the patients with proteinuria were converted from cyclosporine and five were converted from azathioprine to sirolimus. Sirolimus was discontinued in five patients with proteinuria, and in all of them proteinuria declined to baseline values. Our study demonstrates that conversion from azathioprine to sirolimus after kidney transplantation may cause a reversible increase of proteinuria. Sirolimus-induced proteinuria therefore cannot be ascribed to the hemodynamic renal effects of withdrawal of CI.

Topics: Aged; Azathioprine; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Creatinine; Female; Follow-Up Studies; Humans; Immunosuppression Therapy; Immunosuppressive Agents; Kidney; Kidney Transplantation; Male; Middle Aged; Prospective Studies; Proteinuria; Sirolimus; Skin Neoplasms; Time Factors

Transglutaminase 2 (TG2) is an important cancer stem-like cell survival protein that is highly expressed in epidermal squamous cell carcinoma and drives an aggressive cancer phenotype. In the present study, we show that TG2 knockdown or inactivation results in a reduction in mammalian target of rapamycin (mTOR) level and activity in epidermal cancer stem-like cells which are associated with reduced spheroid formation, invasion, and migration, and reduced cancer stem cell and epithelial-mesenchymal transition (EMT) marker expression. Similar changes were observed in both cultured cells and tumors. mTOR knockdown or treatment with rapamycin phenocopies the reduction in spheroid formation, invasion, and migration, and cancer stem cell and EMT marker expression. Moreover, mTOR appears to be a necessary mediator of TG2 action, as a forced expression of constitutively active mTOR in TG2 knockdown cells partially restores the aggressive cancer phenotype and cancer stem cell and EMT marker expression. Tumor studies show that rapamycin reduces tumor growth and cancer stem cell marker expression and EMT. These studies suggest that TG2 stimulates mTOR activity to stimulate cancer cell stemness and EMT and drive aggressive tumor growth.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Cell Survival; Epithelial-Mesenchymal Transition; GTP-Binding Proteins; Humans; Phenotype; Protein Glutamine gamma Glutamyltransferase 2; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

Immunosuppressive agents are essential for graft survival in solid-organ transplant recipients (SOTRs), but they have substantial durable side effects, including a higher incidence of aggressive nonmelanoma skin cancers (NMSCs). Hitherto, only one class of immunosuppressants, mammalian target of rapamycin inhibitors (mTORi), may inhibit skin tumor formation, however their durable effectiveness is controversial. To evaluate the sustained effectiveness of mTORi in reducing NMSCs' incidence in SOTRs, a retrospective study was conducted in a specialized dermatology clinic for SOTRs of a tertiary university-affiliated medical center. SOTRs with a history of at least one histologically proven NMSC were followed for 6 years: 3 years after transplantation, before initiation of mTORi, and 3 years under mTORi treatment. The cohort consisted of 44 SOTRs. Treatment with mTORi was initiated on average 6.27 (3.34-6.34) years following transplantation. In the 3 years before mTORi treatment initiation, the mean number of new NMSCs per patient was 2.11 (1-14). This value decreased to 1.2 (0-19) in the 3 years under mTORi treatment (p = 0.0007). Analysis by NMSC type yielded a significant decrease in both SCCs and BCCs. This study found that mTORi are effective for prolonged secondary prevention of NMSCs in SOTRs.

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Humans; Immunosuppressive Agents; MTOR Inhibitors; Organ Transplantation; Retrospective Studies; Secondary Prevention; Sirolimus; Skin Neoplasms; TOR Serine-Threonine Kinases

Organ transplant recipients show a high incidence for the formation of cutaneous squamous cell carcinoma (cSCC), while sirolimus appears to reduce the risk. GRO-α is a chemokine, which is overexpressed in many tumor entities and associated with malignant transformation. However, little is known about the expression and function of GRO-α in human cSCC.. Our aim was to investigate the relevance of the GRO-α (CXCL-1)/ CXCR2 axis in human cSCC and the potential impact of sirolimus.. We analyzed the GRO-α expression in human keratinocytes, different cSCC cell lines as well as cSCC tissue and investigated its effect on cell proliferation and migration. Additionally, we incubated cells with sirolimus and measured the expression of GRO-α and its receptor CXCR2.. We showed that both constitutive as well as induced GRO-α expression is higher in in cSCC cell lines compared to keratinocytes and that GRO-α protein is detectable in human cSCC tissue. By GRO-α exposure and shRNA knock down, we identified GRO-α as a driving factor in proliferation and migration. Moreover, in a dermis equivalent GRO-α knocked down cSCC cell lines displayed a reduced capacity in tumor nest formation. Incubation with sirolimus significantly inhibited GRO-α expression in keratinocytes as well as tumor cell lines. Moreover, sirolimus decreased the expression of the corresponding receptor CXCR2.. Taken together, our results suggest that the GRO-α/CXCR2 axis plays a role in human keratinocyte carcinogenesis and might represent a molecular mechanism for the preventive effect of mTOR inhibitors in cSCC development.

Topics: Carcinogenesis; Carcinoma, Squamous Cell; Cell Line, Tumor; Chemokine CXCL1; Gene Knockdown Techniques; Humans; Keratinocytes; MTOR Inhibitors; Receptors, Interleukin-8B; Signal Transduction; Sirolimus; Skin Neoplasms

There is not much progress in the treatment for lung squamous cell carcinoma LSCC in the past few years. Rapamycin Rapa, an inhibitor of mammalian target of rapamycin mTOR, has exhibited antitumor efficacy in a variety of malignant tumors. It has recently been reported that Rapamycin can induce autophagy signaling pathway in lung cancer and Glypican-3GPC3 can promote the growth of hepatocellular carcinoma by stimulating canonical Wnt signaling pathway. The aim of this study is to investigate the mechanisms of rapamycin's antitumor efficacy in relation to GPC3/Wnt/β-catenin pathway and autophagy in LSCC.. SK-MES-1 cells, a LSCC cell line, were treated with various concentrations of rapamycin with or without Glypican-3 GPC3-targeting siRNA. SK-MES-1 cell proliferation was determined by MTT assay. Protein expression levels of GPC3, β-catenin, Beclin-1 were checked via western blotting. We established the xenograft mice model to investigate the suppression effect of rapamycin on LSCC. In addition, we further testified the metabolism protein of autophagy process using the xenograft tumor tissue.. Rapamycin could inhibit the SK-MES-1 cell proliferation in a concentration-dependent manner both in vitro and in vivo by decreasing the GPC3 expression and downregulating the glypican-3/Wnt/β-catenin signaling pathway. In addition, we found that GPC3 silencing can activate the glypican-3/Wnt/β-catenin pathway and autophagy, which contribute to the suppression of tumor growth both in vitro and in vivo.. Rapamycin suppresses the growth of lung cancer through down-regulating glypican-3/Wnt/β-catenin signaling, which mediates with activation of autophagy. This study suggests GPC3 is a new promising target for rapamycin in the treatment of lung cancer.

Topics: Animals; Apoptosis; Autophagy; beta Catenin; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Down-Regulation; Female; Glypicans; Humans; Lung Neoplasms; Mice; Mice, Inbred BALB C; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Wnt Signaling Pathway

Tumor-associated macrophages (TAMs) promote tumor progression. The number of infiltrating TAMs is associated with poor prognosis in esophageal squamous cell carcinoma (ESCC) patients; however, the mechanism underlying this phenomenon is unclear. cDNA microarray analysis indicates that the expression of chemokine (C-C motif) ligand 1 (CCL1) is up-regulated in peripheral blood monocyte-derived macrophages stimulated using conditioned media from ESCC cells (TAM-like macrophages). Here, we evaluated the role of CCL1 in ESCC progression. CCL1 was overexpressed in TAM-like macrophages, and CCR8, a CCL1 receptor, was expressed on ESCC cell surface. TAM-like macrophages significantly enhanced the motility of ESCC cells, and neutralizing antibodies against CCL1 or CCR8 suppressed this increased motility. Recombinant human CCL1 promoted ESCC cell motility via the Akt/proline-rich Akt substrate of 40 kDa/mammalian target of rapamycin pathway. Phosphatidylinositol 3-kinase or Akt inhibitors, CCR8 silencing, and neutralizing antibody against CCR8 could significantly suppress these effects. The overexpression of CCL1 in stromal cells or CCR8 in ESCC cells was significantly associated with poor overall survival (P = 0.002 or P = 0.009, respectively) and disease-free survival (P = 0.009 or P = 0.047, respectively) in patients with ESCC. These results indicate that the interaction between stromal CCL1 and CCR8 on cancer cells promotes ESCC progression via the Akt/proline-rich Akt substrate of 40 kDa/mammalian target of rapamycin pathway, thereby providing novel therapeutic targets.

Topics: Carcinoma, Squamous Cell; Cell Movement; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Humans; Ligands; Macrophages; Proto-Oncogene Proteins c-akt; Receptors, CCR8; Sirolimus; TOR Serine-Threonine Kinases; Tumor-Associated Macrophages

The immunosuppressive efficiency obtained in the last decades in kidney transplantation significantly improved graft survival. However, there is still a high risk and incidence of cancer in transplant patients strongly and directly related to the type of immunosuppression. An increasing body of evidence suggests that the PI3K/Akt/mTOR pathway may play a pivotal role in the development and progression of several neoplastic diseases.. We describe a 47-year-old male patient who received a cadaveric primary renal transplant in November 2008 developing a poorly differentiated infiltrating and ulcerated squamous cell carcinoma (SCC) at the eye level. In this patient, the modification of an immunosuppressive regimen with introduction of rapamycin (mTOR) inhibitors and withdrawal of calcineurin inhibitors (CNIs) led to the resolution of this severe condition.. The introduction of mTOR inhibitors and withdrawal of CNIs in kidney-transplanted patients with de novo eye SCC should be considered in this clinical setting.

Topics: Antineoplastic Agents; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Everolimus; Eye Neoplasms; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Sirolimus; TOR Serine-Threonine Kinases

The therapeutic effect of dihydroartemisinin (DHA) against cutaneous squamous cell carcinoma (cSCC) has been previously demonstrated; however, the underlying mechanism remains unclear. This study sought to verify the therapeutic effect of DHA against cSCC and explore its underlying mechanism in A431 cSCC cells. This study reported that DHA inhibited A431 cells proliferation in a time- and concentration-dependent manner and promoted A431 cells apoptosis. Moreover, DHA inhibited the invasion and migration of A431 cells. Mechanistically, DHA promoted autophagy and inhibited activation of the absent in melanoma 2 (AIM2) inflammasome pathway and NF-κB/HIF-1α/VEGF pathway. Treatment of A431 cells with the mTOR inhibitor, and autophagy promoter, rapamycin also inhibited these two pathways. In conclusion, DHA inhibited activation of the AIM2 inflammasome pathway and NF-κB/HIF-1α/VEGF pathway by promoting autophagy in A431 cells, thus accounting for its therapeutic effect. Induction of autophagy by DHA may be mediated by inhibiting the mTOR pathway and promoting reactive oxygen species production.

Topics: Apoptosis; Artemisinins; Autophagy; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Drug Screening Assays, Antitumor; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Inflammasomes; NF-kappa B; Signal Transduction; Sirolimus; Skin Neoplasms; TOR Serine-Threonine Kinases; Vascular Endothelial Growth Factor A

Cutaneous squamous cell carcinoma (CSCC) represents the most common malignant tumour of the feline skin. Emerging evidence suggests that the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway may represent a potential target for pharmacological intervention in human and canine CSCC.. The present study aimed to explore the expression pattern and status of activation of relevant signalling proteins of the PI3K/Akt/mTOR signalling pathway in feline CSCC.. The expression of pEGFR. The immunodetection using phosphospecific antibodies to detect the activated forms of signalling proteins showed that the PI3K/Akt/mTOR signalling pathway is frequently activated in feline CSCCs, and may be independent of the activation of EGFR. The results also showed that PTEN expression is not significantly altered in feline CSCCs.. Our study shows that the persistent activation of the PI3K/Akt/mTOR signalling pathway represents a key event in feline CSCC, pointing to this signalling pathway being a potential therapeutic target in feline patients with CSCC.. Le carcinome épidermoïde cutané (CSCC) représente la tumeur maligne la plus fréquente de la peau chez le chat. Des preuves émergentes suggèrent que la voie de signal PI3K/Akt/mTOR (phosphatidylinositol 3-kinase/Akt/mammalian cible de rapamycine) pourrait représenter une cible potentielle pour le traitement pharmacologique de CSCC du chien et de l’homme. HYPOTHÈSES/OBJECTIFS: L’étude a pour objectif d’explorer le patron d’expression et le statut d’activation des protéines de la voie de signal PI3K/Akt/mTOR dans les CSCC du chat. MÉTHODES: L’expression de pEGFRTyr1068, pAktSer473, pS6Ser235/236 combinée avec Ki-67, et PTEN protéine suppresseur de tumeur a été évaluée par analyse immunohistochimique dans 45 échantillons de CSCC félins à l’aide de microéchantillons tissulaires. RÉSULTATS: L’immunodétection utilisant des anticorps phosphospécifiques pour détecter les formes activées de protéines de signal a montré que la voie de signal PI3K/Akt/mTOR est fréquemment activée dans les CSCC félins et pourrait être indépendant de l’activation de EGFR. Les résultats ont aussi montré que l’expression de PTEN n’est pas significativement altérée dans les CSCC du chat.. Notre étude montre que l’activation persistante de la voie de signal PI3K/Akt/mTOR représente un événement clé dans les CSCC félins, pointant cette voie de signal comme potentielle cible thérapeutique chez les chats atteints de CSCC.. Das kutane Plattenepithelkarzinom (CSCC) stellt den häufigsten bösartigen Tumor der Katzenhaut dar. Entstehende Evidenz deutet darauf, dass der Signalweg von Phosphatidylinositol 3-Kinase/Akt/Säugetier Target von Rapamycin (PI3K/Akt/mTOR) als möglicher Angriffspunkt einer pharmakologischen Intervention beim CSCC des Menschen und des Hundes dienen könnte.. Die vorliegende Studie zielte darauf ab, das Exprimierungsmuster und den Status der Aktivierung relevanter Signalproteine des PI3K/Akt/mTOR Signalwegs des CSCCs der Katze zu untersuchen.. Die Exprimierung von pEGFR. Die Immundetektion mittels Phospho-spezifischer Antikörper, um die aktivierten Formen des Signalproteins zu finden, zeigte, dass der PI3K/Akt/mTOR Signalweg häufig bei felinen CSCCs aktiviert wird und von der Aktivierung von EGFR unabhängig sein könnte. Die Ergebnisse zeigten, dass die Exprimierung von PTEN bei felinen CSCCs nicht signifikant verändert waren.. Unsere Studie zeigt, dass die persistierende Aktivierung des PI3K/Akt/mTOR Signalweges ein Schlüsselevent beim felinen CSCC darstellt, was zeigt, dass dieser Signalweg ein potentieller therapeutischer Ansatzpunkt für feline Patienten mit CSCC sein könnte.. 背景: 膚扁平上皮癌 (CSCC) は、猫の皮膚の最も一般的な悪性腫瘍である。新たなエビデンスは、ホスファチジルイノシトール3-キナーゼ/ Akt /哺乳類のラパマイシン標的 (PI3K / Akt / mTOR) シグナル伝達経路が、ヒトおよびイヌのCSCCにおける薬理学的介入の潜在的な標的となる可能性があることを示唆している。 仮説/目的: 本研究は、ネコのCSCCにおけるPI3K / Akt / mTORシグナル伝達経路の関連するシグナル伝達タンパク質の発現パターンおよび活性化の状態を調査することを目的としている。 方法: Ki-67と組み合わせたpEGFRTyr1068、pAktSer473、pS6Ser235 / 236、および腫瘍抑制タンパク質PTENの発現を、組織マイクロアレイ法を使用して、ネコCSCCの45サンプルで免疫組織化学的分析によって評価した。 結果: リン酸化特異的抗体を使用してシグナル伝達タンパク質の活性化型を検出する免疫検出は、PI3K / Akt / mTORシグナル伝達経路がネコのCSCCで頻繁に活性化され、EGFRの活性化とは無関係である可能性があることを示した。結果は、PTEN発現がネコのCSCCで有意に変化しないことも示した。 結論と臨床的重要性: 我々の研究は、PI3K / Akt / mTORシグナル伝達経路の持続的な活性化がネコCSCCの重要なイベントであり、このシグナル伝達経路がCSCCに罹患したネコの潜在的な治療標的であることを示している。.. O carcinoma de células escamosas (CCE) representa o tumor maligno mais comum da pele de felinos. Evidências recentes sugerem que a via de sinalização da fosfatidilinositol 2-quinase/Akt/alvo mamífero da rapamicina (PI3K/Akt/mTOR) pode ser um alvo potencial de intervenções farmacológicas no CCE humano e canino. HIPÓTESE/OBJETIVOS: O presente estudo objetivou explorar o padrão de expressão e o status de ativação das proteínas sinalizadoras relevantes da via de sinalização PI3K/Akt/mTOR no CCE felino. MÉTODOS: A expressão de pEGFR. A imunodetecção utilizando anticorpos fosfoespecíficos para detectar as formas ativas de proteínas de sinalização demonstraram que a via de sinalização PI3K/Akt/mTOR é frequentemente ativada no CCE felino, e pode ser independente da ativação de EGFR. Os resultados também demonstram que a expressão de PTEN não está significativamente alterada no CCE felino. CONCLUSÕES E IMPORTÂNCIA CLÍNICA: Nosso estudo revelou que a ativação persistente da via de sinalização PI3K/Akt/mTOR representa um evento chave na CCE felina, indicando que esta via de sinalização representa um alvo terapêutico potencial em pacientes felinos com CCE.

Topics: Animals; Carcinoma, Squamous Cell; Cat Diseases; Cats; Phosphatidylinositol 3-Kinases; Sirolimus; Skin Neoplasms; TOR Serine-Threonine Kinases

The mechanistic target of rapamycin (MTOR) plays a key role in regulating cell growth and metabolism and is commonly overexpressed in head and neck cancer (HNSCC). This study investigated the association of MTOR with clinical outcome in human papilloma virus (HPV) positive and negative HNSCC patients treated by chemoradiation.. A tissue microarray (TMA) consisting of cores from 109 HNSCC patients treated by definitive chemoradiation was constructed and stained with antibodies against p16 and MTOR and expression correlated with clinicopathological features and clinical outcome.. MTOR varied widely between tumor cores and was not associated with HPV status or clinicopathological features. There was a positive correlation with pre-treatment FDG uptake. (P = .01). In HPV negative patients, MTOR predicted for shorter locoregional control (P = .02), diseases free survival (P = .02), and overall survival (P = .04). MTOR expression was not associated with outcome in HPV positive patients.. Prognostic significance of MTOR expression depends on HPV status.

Topics: Carcinoma, Squamous Cell; Head and Neck Neoplasms; Humans; Papillomaviridae; Papillomavirus Infections; Prognosis; Sirolimus; TOR Serine-Threonine Kinases

Studies have shown the mammalian target of rapamycin (mTOR) and 70-kDa ribosomal protein S6 kinase (p70S6K) to be tumor suppressors in many cancers. These factors may have synergistic functions in tongue squamous cell carcinoma (TSCC), which is the most common malignant cancer in the oral region. We aimed to investigate the expression of the mTOR-p70S6K axis in TSCC patients and its biological function in TSCC cell lines.. Sixty-eight TSCC patients were included in this study, and their features, including age, gender, tumor differentiation, lymphatic metastasis, and clinical stage, were recorded. The expression of mTOR and p70S6K was detected by immunohistochemistry. Small interfering RNA constructs were delivered into TSCC cells to downregulate mTOR and p70S6K expression in vitro. After transfection, cell proliferation, migration or invasion, apoptosis, and chemoresistance assays were performed to examine cellular variations of biological function.. High expression of the mTOR-p70S6K axis was associated with higher tumor stage, lymph node metastasis, and poor tumor differentiation. Suppression of mTOR and p70S6K in TSCC cells resulted in the inhibition of cell proliferation, metastases, and chemoresistance. Inhibiting mTOR expression could inhibit p70S6K expression but not vice versa.. The high expression of mTOR and p70S6K is closely associated with malignant characterization of TSCC patients, and it could inhibit biological functions of TSCC cell lines. Taken together, the mTOR-p70S6K axis may serve as a potential therapeutic strategy for TSCC.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Humans; Ribosomal Protein S6 Kinases, 70-kDa; Sirolimus; Tongue Neoplasms; TOR Serine-Threonine Kinases

Topics: Acitretin; Aged; Carcinoma, Squamous Cell; Drug Therapy, Combination; Humans; Immunosuppressive Agents; Keratolytic Agents; Male; Organ Transplantation; Sirolimus; Skin Neoplasms; Transplant Recipients; Wound Healing

Hypopharyngeal carcinoma (HPC) is an aggressive malignancy with the worst prognosis among all head and neck cancers. MicroRNAs (miRNAs) are involved in the development of many human cancers, and may function as oncogenes or tumor suppressors. The present study aimed to evaluate the effects of miRNA (miR)‑194‑5p on the proliferation and invasion of HPC cells and to identify the potential regulatory mechanism. First, miR‑194‑5p and Smad ubiquitin regulatory factor 1 (SMURF1) expression levels were examined in HPC tissues. Subsequently, to explore the effects of miR‑194‑5p on SMURF1, a dual‑luciferase reporter gene assay was performed to verify the target relationship. To define the role of miR‑194‑5p in HPC progression, miR‑194‑5p upregulation and depletion were used to evaluate its effects on cell viability, invasion and migration. SMURF1 silencing and rapamycin [an inhibitor of the mammalian target of rapamycin (mTOR) signaling pathway] treatment were also used to analyze the regulatory mechanism in HPC. Finally, tumor growth was assessed in xenografted tumors in nude mice. SMURF1 was demonstrated to be highly expressed, whereas miR‑194‑5p was poorly expressed in HPC tissues; SMURF1 was identified as a target gene of miR‑194‑5p. FaDu hypopharyngeal squamous cell carcinoma cells treated with miR‑194‑5p mimics exhibited decreased viability, invasion and migration. The results indicated that miR‑194‑5p may inactivate the mTOR signaling pathway by targeting SMURF1. In addition, the in vivo experiments further verified these regulatory effects. These data suggested that miR‑194‑5p‑targeted SMURF1 inhibition may be involved in the disruption of HPC progression through the repression of the mTOR signaling pathway.

Topics: 3' Untranslated Regions; Aged; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; Hypopharyngeal Neoplasms; Male; Mice; MicroRNAs; Middle Aged; Neoplasm Invasiveness; Neoplasm Transplantation; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Ubiquitin-Protein Ligases; Up-Regulation

Head and neck squamous cell carcinoma (HNSCC) clearly involves activation of the Akt mammalian target of rapamycin (mTOR) signalling pathway. However, the effectiveness of treatment with the mTOR inhibitor rapamycin is often limited by chemoresistance. Melatonin suppresses neoplastic growth via different mechanisms in a variety of tumours. In this study, we aimed to elucidate the effects of melatonin on rapamycin-induced HNSCC cell death and to identify potential cross-talk pathways. We analysed the dose-dependent effects of melatonin in rapamycin-treated HNSCC cell lines (Cal-27 and SCC-9). These cells were treated with 0.1, 0.5 or 1 mmol/L melatonin combined with 20 nM rapamycin. We further examined the potential synergistic effects of melatonin with rapamycin in Cal-27 xenograft mice. Relationships between inhibition of the mTOR pathway, reactive oxygen species (ROS), and apoptosis and mitophagy reportedly increased the cytotoxic effects of rapamycin in HNSCC. Our results demonstrated that combined treatment with rapamycin and melatonin blocked the negative feedback loop from the specific downstream effector of mTOR activation S6K1 to Akt signalling, which decreased cell viability, proliferation and clonogenic capacity. Interestingly, combined treatment with rapamycin and melatonin-induced changes in mitochondrial function, which were associated with increased ROS production, increasing apoptosis and mitophagy. This led to increase cell death and cellular differentiation. Our data further indicated that melatonin administration reduced rapamycin-associated toxicity to healthy cells. Overall, our findings suggested that melatonin could be used as an adjuvant agent with rapamycin, improving effectiveness while minimizing its side effects.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Head and Neck Neoplasms; Humans; Melatonin; Mice; Mice, Nude; Mitophagy; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Honokiol, an active natural product derived from Magnolia officinalis, exerted anticancer effects through a variety of mechanisms on multiple types of cancers. In this study, the molecular mechanisms of honokiol in suppressing the human oral squamous cell carcinoma (OSCC) cells were evaluated. Treatment of two OSCC cell lines with honokiol resulted in reducing the cell proliferation and arresting the cell cycle at G1 stage which was correlated with the down-regulation of Cdk2 and Cdk4 and the up-regulation of cell cycle suppressors, p21 and p27. In addition, the caspase-dependent programmed cell death was substantially detected, and the autophagy was induced as the autophagosome formation and autophagic flux proceeded. Modulation of autophagy by autophagic inducer, rapamycin or inhibitors, 3-MA or bafilomycin, potentiated the honokiol-mediated anti-OSCC effects where honokiol exerted multiple actions in suppression of MAPK pathway and regulation of Akt/mTOR or AMPK pathways. As compared to clinical therapeutic agent, 5-FU, honokiol exhibited more potent activity against OSCC cells and synergistically enhanced the cytotoxic effect of 5-FU. Furthermore, orally administrated honokiol exerted effective antitumour activity in vivo in OSCC-xenografted mice. Thus, this study revealed that honokiol could be a promising candidate in preventing human OSCCs.

Topics: Adenine; Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Autophagy; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Fluorouracil; G1 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Humans; Lignans; Macrolides; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Mouth Neoplasms; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Tumor Burden; Xenograft Model Antitumor Assays

Inhibitors of the mammalian target of rapamycin (mTORis) exert immunosuppressive and antitumor effects and are used in organ-transplant recipients (OTR) as immunosuppressants able to reduce skin tumor burden. This study investigated the effects of mTORis on the expression of mTOR pathway proteins in cutaneous squamous-cell carcinomas (SCC) developing in OTR, before and after switching to mTORis.. An immunohistochemical study was performed on 23 SCC sections excised from OTR with post-transplant SCC, before or after switch to mTORis, with antibodies against pAkt, pmTOR and PI3K.. pmTOR expression was found in 8/12 SCC pre-switch, and in 8/11 SCC post-switch, to mTORis. All (but 2) SCC expressed PI3K, and all SCCs expressed pAkt.. mTORis do not significantly change the immunohistochemical expression of molecules upstream of the mTOR inhibition (pmTOR, PI3K, pAkt), in cutaneous SCC.

Topics: Carcinoma, Squamous Cell; Humans; Immunohistochemistry; Immunosuppressive Agents; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; Skin Neoplasms; Time Factors; TOR Serine-Threonine Kinases; Transplantation

Vulvar squamous cell carcinoma (VSCC) constitutes over 90% of vulvar cancer. Its pathogenesis can follow two different pathways; high risk human papillomavirus (hrHPV)-dependent and HPV-independent. Due to the rarity of VSCC, molecular mechanisms underlying VSCC development remain largely unknown. The study aimed to identify pathogenic mutations implicated in the two pathways of VSCC development.. Using next generation sequencing, 81 VSCC tumors, 52 hrHPV(+) and 29 hrHPV(-), were screened for hotspot mutations in 50 genes covered by the Ion AmpliSeq Cancer Hotspot Panel v2 Kit (Thermo Fisher Scientific).. Mutations of TP53 (46% and 41%, of hrHPV(+) and hrHPV(-) cases respectively) and CDKN2A (p16) (25% and 21%, of hrHPV(+) and hrHPV(-) cases respectively) were the most common genetic alterations identified in VSCC tumors. Further mutations were identified in PIK3CA, FBXW7, HRAS, FGFR3, STK11, AKT1, SMAD4, FLT3, JAK3, GNAQ, and PTEN, albeit at low frequencies. Some of the identified mutations may activate the PI3K/AKT/mTOR pathway. The activation of mTOR was confirmed in the vast majority of VSCC samples by immunohistochemical staining.. Detecting pathogenic mutations in 13/50 genes examined at comparable frequencies in hrHPV(+) and hrHPV(-) tumors suggest that genetic mechanisms of the two routes of VSCC pathogenesis may be similar, despite being initiated from different premalignant lesions. Importantly, our data provide a rationale for new anti-VSCC therapies targeting the PI3K/AKT/mTOR pathway.

Topics: Adult; Aged; Aged, 80 and over; AMP-Activated Protein Kinase Kinases; Antibiotics, Antineoplastic; Antineoplastic Agents; Benzamides; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Survival; Class I Phosphatidylinositol 3-Kinases; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p18; Disease-Free Survival; DNA Mutational Analysis; DNA, Neoplasm; Everolimus; F-Box-WD Repeat-Containing Protein 7; Female; fms-Like Tyrosine Kinase 3; GTP-Binding Protein alpha Subunits, Gq-G11; High-Throughput Nucleotide Sequencing; Humans; Janus Kinase 3; Middle Aged; Morpholines; Mutation; Papillomaviridae; Papillomavirus Infections; Phosphatidylinositol 3-Kinase; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins p21(ras); PTEN Phosphohydrolase; Pyrimidines; Receptor, Fibroblast Growth Factor, Type 3; Signal Transduction; Sirolimus; Smad4 Protein; TOR Serine-Threonine Kinases; Tumor Suppressor Protein p53; Vulvar Neoplasms

Due to their immunosuppressive therapy, organtransplant recipients (OTRs) exhibit a high incidence for the development of cutaneous squamous cell carcinoma (cSCC). Randomized studies of kidney-transplanted patients indicate a significant lower susceptibility for cSCC among patients receiving the mTOR-inhibitor Sirolimus, compared to patients without mTOR-regimen. The exact mechanism, how mTOR inhibition affects keratinocyte carcinogenesis remains unclear.. Our aim was to investigate the impact of Sirolimus on the expression level of the oncogene ATF3, which is involved in the development and progression of cSCC.. We incubated human keratinocytes, cSSC cell lines and 3D skin equivalents with Sirolimus, exposed the cells to calcineurin inhibitors (CNI) and UVA-radiation and measured the expression level of ATF3 by real-time PCR and western blot.. We show that Sirolimus downregulates the expression of ATF3 induced by cyclosporine or cyclosporine plus UV-radiation in keratinocytes. In line with this we demonstrate a decrease in ATF3 expression, by incubating 3D skin equivalents with Sirolimus prior to cyclosporine and UV-light. However, Sirolimus has no significant impact on the ATF3 expression levels of cyclosporine stimulated cSCC cell lines.. Taken together, our study demonstrates that Sirolimus downregulates the CNI or UV-induced ATF3 expression in human keratinocytes, which could be a potential molecular mechanism how Sirolimus reduces cSCC in OTRs. The lack of ATF3 suppression by Sirolimus in cSCC cell lines fits to observations from clinical studies which demonstrated a clinical benefit from the switch to a mTOR-regimen in patients with low tumor burden in early stage of disease.

Topics: Activating Transcription Factor 3; Calcineurin Inhibitors; Carcinogenesis; Carcinoma, Squamous Cell; Cell Culture Techniques; Cell Line, Tumor; Cyclosporine; Down-Regulation; Humans; Immunosuppressive Agents; Keratinocytes; Oncogenes; Organ Transplantation; Sirolimus; Skin; Skin Neoplasms; TOR Serine-Threonine Kinases; Ultraviolet Rays

Hyperactivation of mTOR signaling pathway has been viewed as a significant molecular pathogenesis of cancer. However, inhibition of mTOR by rapamycin and its analogs could induce numerous negative feedback loops to attenuate their therapeutic efficacy. As a traditional Chinese herbal medicine, Rabdosia rubescens has been used to treat esophageal squamous cell carcinoma (ESCC) for hundreds of years, and its major effective component is oridonin. Here we reported that OP16, a novel analog of oridonin, showed potent inhibition of cell proliferation and Akt phosphorylation in ESCC cells. The combination of OP16 and rapamycin possesses synergistic anti-proliferative and pro-apoptotic effects both in ESCC cells and ESCC xenografts, and no obvious adverse effect was observed in vivo. Mechanistic analysis revealed that OP16 could inhibit rapamycin-induced Akt activation through the p70S6K-mediated negative feedback loops, and the combination of OP16 and rapamycin was more effective in activating caspase-dependent apoptotic signaling cascade. This study supports the combined use of OP16 with rapamycin as a feasible and effective therapeutic approach for future treatment of ESCC.

Topics: Animals; Antibiotics, Antineoplastic; Antineoplastic Agents, Phytogenic; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Diterpenes, Kaurane; Drug Synergism; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Feedback, Physiological; Female; Humans; Mice, Nude; Phosphorylation; Protein Processing, Post-Translational; Proto-Oncogene Proteins c-akt; Random Allocation; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; Specific Pathogen-Free Organisms; Tumor Burden; Xenograft Model Antitumor Assays

YM155, a small molecule inhibitor of survivin, has been studied in many tumors. It has been shown that YM155 inhibited oral squamous cell carcinoma through promoting apoptosis and autophagy and inhibiting proliferation. It was found that YM155 also inhibited the oral squamous cell carcinoma-mediated angiogenesis through the inactivation of the mammalian target of rapamycin pathway. Rapamycin, a mammalian target of rapamycin inhibitor, played an important role in the proliferation and angiogenesis of oral squamous cell carcinoma cell lines. In our study, cell proliferation assay, transwell assay, tube formation assay, and western blot assay were used to investigate the synergistic effect of rapamycin on YM155 in oral squamous cell carcinoma. Either in vitro or in vivo, rapamycin and YM155 exerted a synergistic effect on the inhibition of survivin and vascular endothelial growth factor through mammalian target of rapamycin pathway. Overall, our results revealed that low-dose rapamycin strongly promoted the sensitivity of oral squamous cell carcinoma cell lines to YM155.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Autophagy; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Imidazoles; Inhibitor of Apoptosis Proteins; Mice; Mouth Neoplasms; Naphthoquinones; Neovascularization, Pathologic; Sirolimus; Survivin; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

Our aim was to evaluate whether repetition of C-ion (carbon ion beam) irradiation induces radioresistance as well as repeated X-ray irradiation in cancer cell lines, and to find the key molecular pathway for radioresistance by comparing radioresistant cancer cells with their parental cells. A mouse squamous cell carcinoma cell line, NR-S1, and radioresistant cancer cells, NR-S1-C30 (C30) and NR-S1-X60 (X60), established by repetition of C-ion and X-ray irradiation, respectively, were used. X-ray and C-ion sensitivity, changes in lysosome, mitochondria, intracellular ATP and reactive oxygen species (ROS) level, and mechanistic target of rapamycin (mTOR) signaling were evaluated. Moreover, the effect of rapamycin on radioresistance was also assessed. X-ray and C-ion resistance of C30 cells was moderate, and the resistance of X60 cells was the highest in this study. In X60 cells, the amount of lysosome, mitochondria, intracellular ATP and ROS level were significantly increased, and mTOR and p70S6K (ribosomal protein S6 kinase p70) phosphorylation were enhanced compared with C30 and NR-S1 cells. The inhibition of mTOR signaling was effective for X-ray and C-ion radiosensitization in both cell lines, especially in X60 cells in which X-ray and C-ion resistance was decreased to the same level as that in NR-S1 cells. Our results indicated that the contribution to generate X-ray and C-ion resistance was less for repeated C-ion irradiations compared with repeated X-ray irradiation. Moreover, we found that activated mTOR signaling contributes to X-ray and C-ion resistance in the X60 cancer cells.

Topics: Adenosine Triphosphate; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Heavy Ion Radiotherapy; Mice; Mitochondria; Phosphorylation; Radiation Tolerance; Reactive Oxygen Species; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; X-Ray Therapy; Xenograft Model Antitumor Assays

Esophageal squamous cell carcinoma (ESCC) is one of the most frequently diagnosed cancers in China, but the etiology and mode of carcinogenesis of this disease remain poorly understood. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN), as a negative regulator of Akt/mTOR pathway, frequently mutates or is inactive in many cancers. Although mTOR has been thought a promising cancer therapeutic target, the sensitivity of tumor cells to rapamycin was still to be revaluated. In this study, we measured the effects of rapamycin on cell proliferation and phosphorylation of Akt in ESCC cells with varying degrees of differentiation. And then, the relationship between PTEN status and the sensitivity of cells to rapamycin was investigated in EC9706 cells with or without wild-type PTEN in vitro and in vivo. The results demonstrated ESCC cells with poor differentiation were insensitive to rapamycin of high concentration and rapamycin obviously promoted the phosphorylation of Akt in these cells, but it had no obvious effects on p-Akt in cells with well differentiation. Also, we showed that wild-type PTEN improved the sensitivity of poor differentiation cells to rapamycin through inhibiting phosphorylation of Akt in vitro and in vivo. This study explored the possible molecular mechanism of some ESCC cells insensitive to rapamycin and provided a measure for treating ESCC patients with PTEN inactivation using mTOR inhibitors.

Topics: Animals; Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Pharmacogenetics; Phosphorylation; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Random Allocation; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

Recent preclinical and clinical studies on head and neck squamous cell carcinoma (HNSCC) revealed synergistic effects when combining anti-EGFR agents with conventional chemotherapeutic drugs. Activation of the PI3-kinase/AKT/mTOR signaling pathway has been identified as an important mechanism implicated in tumor progression and resistance to EGFR inhibitors. The aim of this study was to investigate the effects of combining the mTOR inhibitor temsirolimus (Tem) with the anti-EGFR agent cetuximab (Cet) and conventional chemotherapeutic drugs (cisplatin and fluorouracil (C/F)) on an orthotopic model of HNSCC.. Preclinical in vivo study.. We evaluated the anti-tumor efficacy (measured tumor volume) of Tem, Cet, and C/F, administered alone or in combination. Investigations were performed using a human HNSCC cell line, CAL33, injected into the mouth floor of nude mice.. As compared with the control, the combination of Tem and Cet led to the highest tumor inhibition and induced almost complete tumor growth arrest (P = 0.001). Tem significantly enhanced the impact of the Cet-C/F combination on tumor growth (P < 0.001). The highest inhibitory effects of treatments on cell proliferation (Ki67 labeling), MAPK (pP42/44 labeling), and PI3K/AKT/mTOR (pS6R labeling) signaling pathways were found with the Tem-Cet association.. In this orthotopic HNSCC model, the combination of Tem with Cet produced synergistic effects on tumor growth. These results were corroborated by a strong inhibition of both MAPK and PI3K-mTOR signaling pathways.. N/A.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cell Line, Tumor; Cetuximab; Cisplatin; ErbB Receptors; Fluorouracil; Head and Neck Neoplasms; Humans; Mice; Mice, Nude; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) frequently occurs in many human cancers and hampers their therapeutic use. A large body of evidence has demonstrated the pro-survival role of autophagy in many human cancers. However, whether autophagy is involved in the induction of erlotinib resistance in tongue squamous cell carcinoma (TSCC) remains unknown. In this report, we found that autophagy prior to or induced by erlotinib treatment plays an important role in erlotinib resistance in tongue cancer cells. Using LC3 transfection, we observed that autophagy is upregulated and further induced when treated with erlotinib. Moreover, we found that autophagy plays a cytoprotective role by MTT analysis of the cell viability in TSCCs when treated with rapamycin or hydroxychloroquine (HCQ) in combination with erlotinib. However, 3-methyladenine (3-MA) did not influence the autophagy. Then, through siRNA technology and WB, we found that erlotinib-induced autophagy is mediated by ATG5 but not Beclin1. Also, knockdown of ATG5 significantly decreased the erlotinib resistance and knockdown of Beclin1 did not affect the sensitivity to erlotinib in TSCCs. Taken together, this indicates the critical role of non-canonical autophagy in erlotinib resistance in TSCCs.

Topics: Adenine; Antineoplastic Combined Chemotherapy Protocols; Autophagy; Autophagy-Related Protein 5; Beclin-1; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Survival; Drug Resistance, Neoplasm; Erlotinib Hydrochloride; Humans; Hydroxychloroquine; RNA, Small Interfering; Sirolimus; Tongue Neoplasms; Up-Regulation

Aberrant epidermal growth factor receptor (EGFR) signaling is associated with tumor growth in head and neck squamous cell carcinoma (HNSCC) and is a major focus of targeted therapy. The phosphatidylinositol-3-kinase/AKT/mammalian target of the rapamycin (PI3K/AKT/mTOR) signaling pathway is frequently mutated in HNSCC and is involved in disease progression and resistance to EGFR inhibitors. The aim of this study was to assess the antiproliferative effects of mTOR inhibition (temsirolimus) combined with the anti-EGFR monoclonal antibody cetuximab, administered according to different combination schedules. Antiproliferative effects of the combination of temsirolimus and cetuximab were determined on the representative HNSCC CAL33 cell line (PI3KCA H1047R mutated and K-RAS wild-type). In addition, key proteins related to the EGFR pathway (pEGFR/EGFR, pAKT/AKT) and the mTOR pathway (p-p70S6K1, p4E-BP1) were determined to explain the cytotoxic effects. Temsirolimus and cetuximab showed a synergistic effect when administered in combination. Supra-additive effect was lost when the two drugs were administered sequentially, irrespective of which drug was administered first. Synergistic effect of the combination was corroborated by a marked downregulation of pEGFR, significant downregulation of pAKT expression, and a marked diminution of p70S6K1 and p4E-BP1 expression. Our study demonstrated a synergistic effect of temsirolimus and cetuximab administered in combination, well illustrated by a simultaneous blockade of intracellular signaling pathways regulating cell proliferation and survival. These results establish the notion of a schedule dependency for the combined treatment, which can be of importance at the clinical level.

Topics: Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cell Line, Tumor; Cetuximab; Drug Administration Schedule; Drug Interactions; Drug Synergism; ErbB Receptors; Head and Neck Neoplasms; Humans; Mechanistic Target of Rapamycin Complex 1; Proto-Oncogene Proteins c-akt; ras Proteins; Sirolimus; Squamous Cell Carcinoma of Head and Neck

Effective treatments for recurrent/metastatic human papillomavirus-positive (HPV+) head and neck squamous cell cancer (HNSCC) are limited. To aid treatment development, we characterized a novel murine model of recurrent/metastatic HPV+ HNSCC. Further analysis of the parental tumor cell line and its four recurrent/metastatic derivatives led to preclinical testing of an effective treatment option for this otherwise fatal disease. Reverse phase protein arrays identified key signaling cascades in the parental and recurrent/metastatic cell lines. While protein expression profiles differed among the recurrent/metastatic cell lines, activated proteins associated with the mTOR signaling cascade were a commonality. Based on these data, mTOR inhibition was evaluated as an adjuvant treatment for recurrent/metastatic disease. mTOR activity and treatment response were assessed in vitro by western blot, Seahorse, proliferation, clonogenic, and migration assays. Standard-of-care cisplatin/radiation therapy (CRT) versus CRT/rapamycin were compared in vivo. Low-dose rapamycin inhibited mTOR signaling, decreasing proliferation (43%) and migration (62%) while it enhanced CRT-induced cytotoxicity (3.3 fold) in clonogenic assays. Furthermore, rapamycin re-sensitized CRT-resistant, metastatic tumors to treatment in vivo, improving long-term cures (0-30% improved to 78-100%, depending on the recurrent/metastatic cell line) and limiting lymph node metastasis (32%) and lung metastatic burden (30 fold). Studies using immune compromised mice suggested rapamycin's effect on metastasis is independent of the adaptive immune response. These data suggest a role of mTOR activation in HPV+ HNSCC recurrent/metastatic disease and that adjuvant mTOR inhibition may enhance treatment of resistant, metastatic cell populations at the primary site and limit distant metastasis.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Proliferation; Chemotherapy, Adjuvant; Cisplatin; Head and Neck Neoplasms; Humans; Lymphatic Metastasis; Male; Mice; Mice, Inbred C57BL; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Papillomaviridae; Papillomavirus Infections; Sirolimus; TOR Serine-Threonine Kinases; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Topics: Carcinoma, Squamous Cell; Female; Humans; Immunosuppressive Agents; Male; Organ Transplantation; Sirolimus; Skin Neoplasms; Transplant Recipients

Dysfunctional clock signaling is observed in a variety of pathological conditions. Many members of the clock gene family are upregulated in tumor cells. Here, we explored the consequences of a commonly disrupted signaling pathway in head and neck cancer on the regulation of circadian clock genes. PTEN is a key molecular controller of the PI3K signaling, and loss of PTEN function is often observed in a variety of cancers. Our main goal was to determine whether PTEN regulates circadian clock signaling. We found that oxidation-driven loss of PTEN function resulted in the activation of mTOR signaling and activation of the core clock protein BMAL1 (also known as ARNTL). The PTEN-induced BMAL1 upregulation was further confirmed using small interference RNA targeting PTEN, and in vivo conditional depletion of PTEN from the epidermis. We observed that PTEN-driven accumulation of BMAL1 was mTOR-mediated and that administration of Rapamycin, a specific mTOR inhibitor, resulted in in vivo rescue of normal levels of BMAL1. Accumulation of BMAL1 by deletion of PER2, a Period family gene, was also rescued upon in vivo administration of mTOR inhibitor. Notably, BMAL1 regulation requires mTOR regulatory protein Raptor and Rictor. These findings indicate that mTORC1 and mTORC2 complex plays a critical role in controlling BMAL1, establishing a connection between PI3K signaling and the regulation of circadian rhythm, ultimately resulting in deregulated BMAL1 in tumor cells with disrupted PI3K signaling.

Topics: Animals; ARNTL Transcription Factors; Carcinoma, Squamous Cell; Cell Line, Tumor; Circadian Rhythm; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; HeLa Cells; Humans; Lymph Nodes; Mice; Mice, Knockout; Neoplasm Metastasis; Neoplasms, Glandular and Epithelial; Oxygen; Phosphatidylinositol 3-Kinases; PTEN Phosphohydrolase; Reactive Oxygen Species; Risk Factors; RNA, Small Interfering; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

The rising incidence of human papillomavirus (HPV)-associated malignancies, especially for oropharyngeal cancers, has highlighted the urgent need to understand how the interplay between high-risk HPV oncogenes and carcinogenic exposure results in squamous cell carcinoma (SCC) development. Here, we describe an inducible mouse model expressing high risk HPV-16 E6/E7 oncoproteins in adults, bypassing the impact of these viral genes during development. HPV-16 E6/E7 genes were targeted to the basal squamous epithelia in transgenic mice using a doxycycline inducible cytokeratin 5 promoter (cK5-rtTA) system. After doxycycline induction, both E6 and E7 were highly expressed, resulting in rapid epidermal hyperplasia with a remarkable expansion of the proliferative cell compartment to the suprabasal layers. Surprisingly, in spite of the massive growth of epithelial cells and their stem cell progenitors, HPV-E6/E7 expression was not sufficient to trigger mTOR activation, a key oncogenic driver in HPV-associated malignancies, and malignant progression to SCC. However, these mice develop SCC rapidly after a single exposure to a skin carcinogen, DMBA, which was increased by the prolonged exposure to a tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA). Thus, only few oncogenic hits may be sufficient to induce cancer in E6/E7 expressing cells. All HPV-E6/E7 expressing SCC lesions exhibited increased mTOR activation. Remarkably, rapamycin, an mTOR inhibitor, abolished tumor development when administered to HPV-E6/E7 mice prior to DMBA exposure. Our findings revealed that mTOR inhibition protects HPV-E6/E7 expressing tissues form SCC development upon carcinogen exposure, thus supporting the potential clinical use of mTOR inhibitors as a molecular targeted approach for prevention of HPV-associated malignancies.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Proliferation; Gene Expression Regulation, Neoplastic; Human papillomavirus 16; Humans; Mice; Oncogene Proteins, Viral; Oropharyngeal Neoplasms; Papillomavirus E7 Proteins; Papillomavirus Infections; Phorbol Esters; Repressor Proteins; Sirolimus; TOR Serine-Threonine Kinases

It has been demonstrated that mTOR/p70S6K pathway was abnormally activated in many cancers and rapamycin and its analogs can restrain tumor growth through inhibiting this pathway, but some tumors including esophageal squamous cell carcinoma (ESCC) appear to be insensitive to rapamycin in recent studies. In the present study, we explored the measures to improve the sensitivity of ESCC cells to rapamycin and identified the clinical significance of the expression of phosphorylated p70S6K (p-p70S6K). The results showed that, after downregulating the expression of p70S6K and p-p70S6K by p70S6K siRNA, the inhibitory effects of rapamycin on cell proliferation, cell cycle, and tumor growth were significantly enhanced in vitro and in vivo. Furthermore, p-p70S6K had strong positive expression in ESCC tissues and its expression was closely related to lymph node metastasis and the TNM staging. These results indicated that p-p70S6K may participate in the invasion and metastasis in the development of ESCC and downregulation of the expression of p-p70S6K could improve the sensitivity of cells to rapamycin in ESCC.

Topics: Animals; Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Down-Regulation; Drug Resistance, Neoplasm; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Gene Expression; Humans; Lymphatic Metastasis; Male; Mice; Neoplasm Staging; Phosphorylation; Ribosomal Protein S6 Kinases, 70-kDa; RNA Interference; RNA, Small Interfering; Sirolimus; Xenograft Model Antitumor Assays

Topics: Antibodies, Monoclonal, Murine-Derived; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cyclophosphamide; Doxorubicin; Female; Humans; Immunosuppressive Agents; Kidney Transplantation; Lymphoma, Large B-Cell, Diffuse; Middle Aged; Neoplasms, Multiple Primary; Postoperative Complications; Prednisone; Remission Induction; Rituximab; Sirolimus; Skin Diseases; Skin Neoplasms; Vincristine; Warts

Patient-derived xenograft (PDX) models have shown to reflect original patient tumors better than any other preclinical model. We embarked in a study establishing a large panel of head and neck squamous cell carcinomas PDX for biomarker analysis and evaluation of established and novel compounds. Out of 115 transplanted specimens 52 models were established of which 29 were characterized for response to docetaxel, cetuximab, methotrexate, carboplatin, 5-fluorouracil and everolimus. Further, tumors were subjected to sequencing analysis and gene expression profiling of selected mTOR pathway members. Most frequent response was observed for docetaxel and cetuximab. Responses to carboplatin, 5-fluorouracil and methotrexate were moderate. Everolimus revealed activity in the majority of PDX. Mutational profiling and gene expression analysis did not reveal a predictive biomarker for everolimus even though by trend RPS6KB1 mRNA expression was associated with response. In conclusion we demonstrate a comprehensively characterized panel of head and neck cancer PDX models, which represent a valuable and renewable tissue resource for evaluation of novel compounds and associated biomarkers.

Topics: Adult; Aged; Aged, 80 and over; Animals; Antineoplastic Agents; Carcinoma, Squamous Cell; DNA Mutational Analysis; Everolimus; Female; Gene Expression Profiling; Head and Neck Neoplasms; Human papillomavirus 16; Humans; Interleukin Receptor Common gamma Subunit; Male; Mice, Inbred NOD; Mice, Knockout; Mice, SCID; Middle Aged; Papillomavirus Infections; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Treatment Outcome; Xenograft Model Antitumor Assays

Rapamycin inhibits products of molecular pathways in esophageal squamous cell carcinoma and limits tumor cell growth by targeting 4E-BP1- and eIF4E-dependent gene translation. In this study, we investigate the influence of 4E-BP1-to-eIF4E ratio on rapamycin response in esophageal squamous cell carcinoma cells, and the underlying mechanism is discussed.. The response to rapamycin treatment was examined in 6 esophageal cancer cell lines. Adjustment of the 4E-BP1/eIF4E ratio was carried out by knockdown or overexpression of 4E-BP1 and eIF4E. The relationship between Egr-1 and 4E-BP1 expression in esophageal cancer cells was also studied.. The 4E-BP1/eIF4E ratio was adjusted to evaluate the response to rapamycin treatment in TE1 and TE2 esophageal cancer cells. TE2 cells are sensitized to rapamycin treatment after overexpression of 4E-BP1 or knockdown of eIF4E; TE1 cells become resistant to rapamycin after knockdown of 4E-BP1 or overexpression of eIF4E. These data suggest that the 4E-BP1/eIF4E ratio is a determinant for the response of TE1 and TE2 cells to rapamycin treatment. Egr-1 expression was higher in TE2 cells compared with other esophageal cancer cell lines, and its knockdown increased 4E-BP1 expression in TE2 cells, which became sensitive to rapamycin treatment.. The 4E-BP1/eIF4E ratio is a determinant of the response of rapamycin treatment in esophageal cancer cells. Egr-1 can reduce 4E-BP1 gene expression and render esophageal squamous cell carcinoma cells resistant to rapamycin with a relatively low 4E-BP1/eIF4E ratio. Thus, the 4E-BP1/eIF4E ratio may represent a therapeutic index for the prediction of clinical outcome of rapamycin treatment in patients with esophageal squamous cell carcinoma.

Topics: Adaptor Proteins, Signal Transducing; Antineoplastic Agents; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Cycle Proteins; Cell Line, Tumor; Dose-Response Relationship, Drug; Early Growth Response Protein 1; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Eukaryotic Initiation Factor-4F; Gene Expression Regulation, Neoplastic; Humans; Phosphoproteins; RNA Interference; Signal Transduction; Sirolimus; Transfection

Topics: Adaptor Proteins, Signal Transducing; Antineoplastic Agents; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Cycle Proteins; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Eukaryotic Initiation Factor-4F; Humans; Phosphoproteins; Sirolimus

Although marked improvements have been made in surgical technique and chemoradiotherapy, the prognosis for patients with esophageal squamous cell carcinoma (ESCC) is still unsatisfactory. The mammalian target of rapamycin (mTOR) and its downstream signaling, p70 ribosomal S6 protein kinase (p70S6K) and eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1), seem to play central roles in the regulation of cancer cell proliferation and survival. The significance of mTOR and its downstream targets, p70S6K and 4E-BP1, on the prognosis of ESCC remains uncertain, but this pathway is of particular concern because effective inhibitors are already available.. Immunohistochemistry performed to evaluate the expression of phosphorylated mTOR (p-mTOR), phosphorylated p70S6K (p-p70S6K), phosphorylated 4E-binding protein 1 (p-4E-BP1), and Ki-67 using 105 surgically resected ESCC correlated with treatment outcome. The effect of the mTOR signaling pathway inhibitor everolimus on ESCC cell lines were investigated in vitro by the 3-(4.5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and in vivo by a nude mouse xenograft model.. Univariate analysis showed that p-mTOR overexpression (P = .022), p-p70S6K overexpression (P = .002), and Ki-67 labeling index >50% (P = .045) were associated with inferior overall survival (OS). In a multivariate comparison, p-p70S6K overexpression (P = .001; hazard ratio, 2.247) remained independently associated with worse OS. In cell lines and the xenograft model, everolimus significantly inhibited ESCC growth.. Overexpression of p-p70S6K is associated independently with a poor prognosis among patients with ESCC. The mTOR signaling pathway inhibitor everolimus can inhibit ESCC growth in vitro and in vivo. Our findings suggest that inhibition of mTOR signaling pathway may be a promising novel target for ESCC.

Topics: Adult; Aged; Aged, 80 and over; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Everolimus; Female; Humans; Male; Mice; Middle Aged; Phosphorylation; Prognosis; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

This study aims to investigate how immunosuppression influences the protein expression of antiapoptotic members of the Bcl-2 family-namely, Bcl-xL and Mcl-1-in nonmelanoma skin cancer (NMSC) and the peritumoral epidermis of renal transplant recipients.. NMSC and peritumoral epidermis protein expression of Bcl-xL and Mcl-1 were assessed by immunohistochemistry in renal transplant recipients receiving tacrolimus or sirolimus and the general population not receiving immunosuppression.. NMSC from renal transplant recipients compared with patients not receiving immunosuppressant medications had a reduced Bcl-xL expression intensity (P = .042). Mcl-1 expression intensity in NMSC was decreased in tacrolimus-treated patients compared with sirolimus-treated patients and the nonimmunosuppressed population (P = .024). Bcl-xL expression intensity was increased in peritumoral epidermis compared with NMSC (P = .002).. It was shown for the first time that Bcl-xL and Mcl-1 expression are widespread in the peritumoral epidermis and NMSC of renal transplant recipients. Importantly in NMSC, Bcl-xL expression was reduced with immunosuppression exposure, and Mcl-1 expression was reduced in tacrolimus-treated compared with sirolimus-treated patients.

Topics: Adult; Aged; Aged, 80 and over; Apoptosis; bcl-X Protein; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Female; Humans; Immunohistochemistry; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Mitosis; Myeloid Cell Leukemia Sequence 1 Protein; Sirolimus; Skin Neoplasms; Tacrolimus; Transplant Recipients

Head and neck squamous cell carcinoma (HNSCC) is an aggressive epithelial malignancy. It is the most common neoplasm appearing in the upper aerodigestive tract and the sixth most common cancer worldwide. The five-year survival rate remains poor despite advances in surgery, radiation and chemotherapy. Furthermore, the incidence of human papillomavirus (HPV)-associated oropharyngeal cancer is rising. Thus, innovative therapy approaches are imperative in order to improve the situation. Everolimus, an inhibitor of the mammalian target of rapamycin (mTOR) and sorafenib and sunitinib, multityrosine kinase inhibitors, have been notably effective in the therapy of different tumor entities. The modest side-effects and oral application of the drugs might improve patient compliance. Expression levels of mTOR and Amphiregulin (AREG) in p16-positive and -negative SCC (squamous cell carcinoma) and the effect of everolimus, sorafenib or sunitinib on the expression levels of these target proteins were assessed. As far as we are aware of, this is one of the first in vitro studies to evaluate the effect of these small-molecule drugs with regard to the p16 status of SCC cells.. p16-negative HNSCC 11A and 14C cells and p16-positive CERV196 cells were exposed to different concentrations of everolimus, sorafenib and sunitinib for 2-8 days. Expression levels of mTOR and AREG were determined by enzyme-linked immunosorbent assay (ELISA) and compared against a chemonaïve control.. AREG and mTOR were expressed in all tested cell lines. CERV196 displayed a remarkable increase of mTOR expression compared to p16-negative HNSCC. On the contrary, AREG levels were reduced by 50% in CERV196. Everolimus, sorafenib and sunitinib significantly reduced mTOR expression. Everolimus significantly decreased AREG expression independently of the HPV status. Sunitinib and sorafenib increased AREG expression in HNSCC 11A and 14C but not in CERV196.. The applied drugs showed remarkable suppression of mTOR expression, which might delay tumor progression. Interestingly, sorafenib and sunitinib increased AREG in HNSCC 11A and 14C, which could be a possible evasive mechanism following incubation with these drugs. On the contrary, p16-positive CERV196 showed increased susceptibility to sorafenib and sunitinib concerning suppression of AREG expression. Further studies are required to evaluate the HPV-dependent differences of therapy response and the possible consequences for treatment options.

Topics: Amphiregulin; Carcinoma, Squamous Cell; EGF Family of Proteins; Everolimus; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; Human papillomavirus 16; Humans; Indoles; Molecular Targeted Therapy; Niacinamide; Papillomaviridae; Phenylurea Compounds; Pyrroles; Sirolimus; Sorafenib; Squamous Cell Carcinoma of Head and Neck; Sunitinib; TOR Serine-Threonine Kinases

The mammalian target of rapamycin (mTOR) signaling pathway plays a pivotal role in numerous cellular processes involving growth, proliferation and survival. The purpose of this study was to investigate the anti-tumoral effect of the mTOR inhibitor (mTORi) CCI-779 in HNSCC cell lines and its potency in cisplatin- and cetuximab-resistant cells.. A panel of 10 HNSCC cell lines with differences in TP53 mutational status and basal cisplatin sensitivity and two isogenic models of acquired resistance to cisplatin and cetuximab, respectively were studied. Cell survival after treatment with CCI-779, cisplatin and cetuximab alone or in combination was determined by MTT assays. Potential predictive biomarkers for tumor cell sensitivity to CCI-779 were evaluated.. We observed considerable heterogeneity in sensitivity of HNSCC cell lines to CCI-779 monotherapy. Sensitivity was observed in TP53 mutated as well as wild-type cell lines. Total and p-EGFR expression levels but not the basal activity of the mTOR and MAPK signaling pathways were correlated with sensitivity to CCI-779. Resistant cells with increased EGFR activation could be sensitized by the combination of CCI-779 with cetuximab. Interestingly, cell lines with acquired resistance to cisplatin displayed a higher sensitivity to CCI-779 whereas cetuximab-resistant cells were less sensitive to the drug, but could be sensitized to CCI-779 by EGFR blockade.. Activity of CCI-779 in HNSCC cells harboring TP53 mutations and displaying a phenotype of cisplatin resistance suggests its clinical potential even in patients with dismal outcome after current standard treatment. Cetuximab/mTORi combinations might be useful for treatment of tumors with high expression of EGFR/p-EGFR and/or acquired cetuximab resistance. This combinatorial treatment modality needs further evaluation in future translational and clinical studies.

Topics: Antineoplastic Agents; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cetuximab; Cisplatin; Drug Resistance, Neoplasm; ErbB Receptors; Head and Neck Neoplasms; Humans; Mutation; RNA, Messenger; Signal Transduction; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases

Here, we report the chemotherapeutic effect of honokiol, a phytochemical from Magnolia plant, on human head and neck squamous cell carcinoma (HNSCC). Treatment of HNSCC cell lines from different sub-sites, SCC-1 (oral cavity), SCC-5 (larynx), OSC-19 (tongue) and FaDu (pharynx) with honokiol inhibited their cell viability, which was associated with the: (i) induction of apoptosis, (ii) correction of dysregulatory cell cycle proteins of G0/G1 phase. Honokiol decreased the expression levels of epidermal growth factor receptor (EGFR), mTOR and their downstream signaling molecules. Treatment of FaDu and SCC-1 cell lines with rapamycin, an inhibitor of mTOR pathway, also reduced cell viability of HNSCC cells. Administration of honokiol by oral gavage (100 mg/kg body weight) significantly (P < 0.01-0.001) inhibited the growth of SCC-1 and FaDu xenografts in athymic nude mice, which was associated with: (i) inhibition of tumor cell proliferation, (ii) induction of apoptosis, (iii) reduced expressions of cyclins and Cdks, and (iv) inhibition of EGFR signaling pathway. Molecular docking analysis of honokiol in EGFR binding site indicated that the chemotherapeutic effect of honokiol against HNSCC is mediated through its firm binding with EGFR, which is better than that of gefitinib, a commonly used drug for HNSCC treatment.

Topics: Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Binding Sites; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Separation; Cell Survival; Dose-Response Relationship, Drug; ErbB Receptors; Female; Flow Cytometry; Head and Neck Neoplasms; Humans; Lignans; Magnolia; Mice; Mice, Nude; Molecular Docking Simulation; Phytotherapy; Plant Extracts; Signal Transduction; Sirolimus

Little is known about the use of sirolimus for primary prevention of cutaneous squamous cell carcinoma (SCC) among solid organ transplant recipients (SOTRs).. We examined the association between sirolimus exposure and incident SCC risk among SOTRs within Kaiser Permanente Northern California.. Using a retrospective cohort of all Kaiser Permanente Northern California members given a diagnosis of SOTR from 2000 through 2010, we evaluated incident posttransplantation SCC risk in relation to sirolimus exposure. Sirolimus use was determined from electronic pharmacy records, and incident posttransplantation SCCs were identified from health plan electronic pathology records. We used extended Cox regression to examine the independent association between receipt of sirolimus and risk of SCC.. Among 3539 SOTRs, 488 were exposed to sirolimus and 47 developed an incident SCC. SCC risk was not associated with ever use of sirolimus (adjusted hazard ratio 1.18, 95% confidence interval 0.84-1.16) or cumulative duration of sirolimus exposure (adjusted hazard ratio 2.75, 95% confidence interval 0.84-9.04, comparing long-term users with nonusers).. No information was available for some known SCC risk factors, such as skin type and sun exposure.. Among a large cohort of SOTRs, sirolimus exposure was not associated with a reduction in incident posttransplantation SCC risk.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Cohort Studies; Confidence Intervals; Female; Follow-Up Studies; Humans; Immunosuppressive Agents; Incidence; Male; Middle Aged; Organ Transplantation; Proportional Hazards Models; Reference Values; Retrospective Studies; Risk Assessment; Sensitivity and Specificity; Sirolimus; Skin Neoplasms; Transplant Recipients; Treatment Outcome

Long-term immunosuppression in the organ transplant recipient (OTR) population places these individuals at higher risk of developing skin malignancies. Oral retinoids have become a useful tool for pharmacologic prophylaxis in the OTR population. Immunosuppressants that inhibit mTOR, such as sirolimus, may be used in combination with a systemic retinoid for chemoprophylaxis of cutaneous malignancies. We present the case of a male patient status post second renal transplant who developed an abrupt and unexpected rise in sirolimus levels to supra-therapeutic levels after initiation of prophylactic acitretin for innumerable squamous cell carcinomas (SCC). The sirolimus levels returned to baseline after cessation of acitretin. Systemic drug-drug interactions are an important phenomenon, especially in the solid OTR population. It is postulated that this interaction was mediated by acitretin inhibition of CYP3A4, the primary enzyme responsible for sirolimus metabolism. The Drug Interaction Probability Scale (DIPS) indicates this was a "probable" drug-drug interaction. To date, this interaction has not been reported in the literature. This case accentuates the importance of close monitoring of solid OTRs for adverse medication interactions when multiple medications are taken.

Topics: Acitretin; Carcinoma, Squamous Cell; Drug Interactions; Humans; Immunosuppressive Agents; Keratolytic Agents; Kidney Transplantation; Male; Middle Aged; Sirolimus; Skin Neoplasms

We present a case of a 71-year-old man with a history of liver transplantation who was treated with adjuvant radiotherapy with concurrent cisplatin for recurrent cutaneous squamous cell carcinoma of the head and neck. The patient was transitioned from tacrolimus to sirolimus for immunosuppression immediately prior to the start of radiation therapy, with the goal of reducing the risk for further skin cancer recurrence. The patient developed severe normal tissue toxicity, disproportionate to the dose delivered. He was diagnosed with Grade 4 esophagitis and mucositis after just 2,400 cGy in 12 fractions (planned 6,400 cGy in 32 fractions), requiring cessation of therapy. Six months later, the patient was diagnosed with local recurrence and distant metastases in the lung, and unfortunately passed away one month later. Randomized data have demonstrated the anti-neoplastic benefit of sirolimus. Pre-clinical studies and animal models have suggested that sirolimus may be a radiation sensitizer; however, the literature is limited regarding the clinical translation of these biologic findings. The case we presented reflects that concurrent radiation therapy with sirolimus may enhance the cytotoxic effects of radiation therapy and contribute to dose-limiting toxicity. Certainly, further study is necessary to explore this observation.

Topics: Aged; Carcinoma, Squamous Cell; Chemoradiotherapy; Head and Neck Neoplasms; Humans; Immunosuppressive Agents; Liver Transplantation; Male; Neoplasm Staging; Prognosis; Radiation Injuries; Radiotherapy, Adjuvant; Sirolimus; Skin Neoplasms

To explore the differences in sensitivity to rapamycin of five esophageal squamous cell carcinoma cell lines with different differentiation and the changes of sensitivity of the cells after siRNA-interfered expression of p70S6K.. Effects of rapamycin on proliferation of ESCC cell lines with different differentiation, EC9706, TE-1, Eca109, KYSE790 and KYSE450 cells, were investigated using cell counting kit 8 (CCK-8) assay, and according to the above results, the EC9706 cells non-sensitive to rapamycin were chosen to be transfected with p70S6K-siRNA. The changes in sensitivity of cells to rapamycin were measured in vitro and in vivo using CCK-8 kit, flow cytometry and tumor formation in nude mice.. CCK-8 results showed that all the five cell line cells were sensitive to low concentration of rapamycin (<100 nmol/L), but TE-1 and EC9706 cells, which were with poor differentiation, showed resistance to high concentration of rapamycin. After EC9706 cells were treated with 50, 100, 200, 500 and 1 000 nmol/L rapamycin and p70S6K-siRNA, the proliferation rates of EC9706 cells were (48.67 ± 1.68)%, (15.45 ± 1.54)%, (14.00 ± 0.91)%, (10.97 ± 0.72)% and (2.70 ± 0.32)%, respectively, and were significantly lower than that of cells treated with 50, 100, 200, 500 and 1 000 nmol/L rapamycin and control siRNA [(74.53 ± 1.71)%, (68.27 ± 1.35)%, (71.74 ± 2.44)%, (76.23 ± 1.02)% and (80.21 ± 2.77)%] (P<0.05 for all). The results of flow cytometry showed that the ratios of cells in G1 phase of the p70S6K-siRNA, rapamycin and p70S6K-siRNA+ rapamycin groups were (53.82 ± 1.78)%, (57.87 ± 4.01)% and (73.73 ± 3.68)%, respectively, significantly higher than that in the control group (46.09 ± 2.31)% (P<0.05 for all). The results of tumor formation test in vivo showed that the inhibitory effect of rapamycin on tumor growth was stronger after the cells were transfected with p70S6K-siRNA, and the inhibition rate was 96.5%.. ESCC cells with different differentiation have different sensitivity to rapamycin, and p70S6K-siRNA can improve the sensitivity of cells to rapamycin in vitro and in vivo.

Topics: Animals; Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Humans; Mice; Mice, Nude; Ribosomal Protein S6 Kinases, 70-kDa; RNA, Small Interfering; Signal Transduction; Sirolimus; Transfection

In the present study, the ability of metformin to inhibit skin tumor promotion by 12-O-tetradecanoylphorbol-13-acetate (TPA) was analyzed in mice maintained on either an overweight control diet or an obesity-inducing diet. Rapamycin was included for comparison, and a combination of metformin and rapamycin was also evaluated. Metformin (given in the drinking water) and rapamycin (given topically) inhibited development of both papillomas and squamous cell carcinomas in overweight and obese mice in a dose-dependent manner. A low-dose combination of these two compounds displayed an additive inhibitory effect on tumor development. Metformin treatment also reduced the size of papillomas. Interestingly, all treatments seemed to be at least as effective for inhibiting tumor formation in obese mice, and both metformin and rapamycin were more effective at reducing tumor size in obese mice compared with overweight control mice. The effect of metformin on skin tumor development was associated with a significant reduction in TPA-induced epidermal hyperproliferation. Furthermore, treatment with metformin led to activation of epidermal AMP-activated protein kinase (AMPK) and attenuated signaling through mTOR complex (mTORC)-1 and p70S6K. Combinations of metformin and rapamycin were more effective at blocking epidermal mTORC1 signaling induced by TPA consistent with the greater inhibitory effect on skin tumor promotion. Collectively, the current data demonstrate that metformin given in the drinking water effectively inhibited skin tumor promotion in both overweight and obese mice and that the mechanism involves activation of epidermal AMPK and attenuated signaling downstream of mTORC1.

Topics: Adenylate Kinase; Adiponectin; Animals; Body Weight; Carcinogenesis; Carcinoma, Squamous Cell; Diet; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Insulin; Insulin-Like Growth Factor I; Leptin; Mechanistic Target of Rapamycin Complex 1; Metformin; Mice; Mice, Obese; Multiprotein Complexes; Neoplasms, Experimental; Obesity; Overweight; Papilloma; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; Skin Neoplasms; Tetradecanoylphorbol Acetate; TOR Serine-Threonine Kinases

Radiotherapy is widely used to treat cancer because it has the advantage of physically and functionally conserving the affected organ. To improve radiotherapy and investigate the molecular mechanisms of cellular radioresistance, we established a clinically relevant radioresistant (CRR) cell line, SAS-R, from SAS cells. SAS-R cells continue to proliferate when exposed to fractionated radiation (FR) of 2 Gy/day for more than 30 days in vitro. A xenograft tumor model of SAS-R was also resistant to 2 Gy/day of X-rays for 30 days. The density of blood vessels in SAS-R tumors was higher than in SAS tumors. Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, sensitized microvascular endothelial cells to radiation, but failed to radiosensitize SAS and SAS-R cells in vitro. Everolimus with FR markedly reduced SAS and SAS-R tumor volumes. Additionally, the apoptosis of endothelial cells (ECs) increased in SAS-R tumor tissues when both Everolimus and radiation were administered. Both CD34-positive and tomato lectin-positive blood vessel densities in SAS-R tumor tissues decreased remarkably after the Everolimus and radiation treatment. Everolimus-induced apoptosis of vascular ECs in response to radiation was also followed by thrombus formation that leads to tumor necrosis. We conclude that FR combined with Everolimus may be an effective modality to overcome radioresistant tumors via targeting tumor ECs.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Endothelial Cells; Everolimus; Female; Head and Neck Neoplasms; HeLa Cells; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Radiation-Sensitizing Agents; Sirolimus; Squamous Cell Carcinoma of Head and Neck; Uterine Cervical Neoplasms; Xenograft Model Antitumor Assays

Toll-like receptor 4 (TLR4) is expressed in head and neck squamous cell carcinoma (HNSCC) cells and is associated with HNSCC cancer progression. Rapamycin has been proven to be efficient for the treatment of HNSCC in vivo, yet the mechanism is not understood and rapamycin demonstrates little effect in vitro. In the present study, the HNSCC cell lines CAL27 and SCC4 were pre-treated with rapamycin then stimulated with a TLR4 ligand lipopolysaccharide (LPS). Cell proliferation, migration, invasion, resistance to TRAIL-induced apoptosis, cytokine production, NF-κB and p65 activation were determined. The results indicated that LPS significantly stimulated HNSCC cell proliferation, cytokine production, migration, invasion and resistance to apoptosis induced by tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL). Pretreatment with rapamycin significantly attenuated LPS-induced pro-oncogenic effects by inhibiting the activation of NF-κB by LPS. siRNA knockdown of TLR4 in HNSCC cells demonstrated that rapamycin attenuated LPS-induced pro-oncogenic effects via TLR4. Hence, this study suggests rapamycin may be efficient for the treatment of HNSCC by attenuating TLR4-induced pro-oncogenic effects.

Topics: Apoptosis; Carcinoma, Squamous Cell; Cell Proliferation; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; Humans; Lipopolysaccharides; NF-kappa B; Signal Transduction; Sirolimus; TNF-Related Apoptosis-Inducing Ligand; Toll-Like Receptor 4

Inhibition of mammalian target of rapamycin (mTOR) kinase enhances the radiosensitivity of some cancer cells. We investigated the effect of RAD001, an mTOR inhibitor, on irradiated oral cancer cell lines.. Clonogenic assays were performed to determine the radiosensitivity of SCC4 and SCC25 cells after treatment with RAD001. Target protein phosphorylation, apoptosis, and cell-cycle progression were assessed in SCC4 cells treated with RAD001 with and without ionizing radiation.. RAD001 increased the radiosensitivity of SCC4 cells without affecting cell death; it also inhibited phosphorylation of mTOR, S6, and factor 4E binding protein 1 and reduced the clonogenic survival of irradiated cancer cells. RAD001 combined with radiation increased G2 arrest by activating CHK1, which phosphorylates CDC25C at Ser216, thereby inhibiting CDC2-cyclin B 1 complex formation.. RAD001 enhances the radiosensitivity of SCC4 cells by inhibiting mTOR signaling and inducing G2 cell-cycle arrest through disruption of the G2 checkpoint.

Topics: Apoptosis; Blotting, Western; Carcinoma, Squamous Cell; Cell Cycle; Cell Cycle Checkpoints; Cell Proliferation; Everolimus; Humans; Immunosuppressive Agents; Phosphorylation; Radiation Tolerance; Radiation, Ionizing; Signal Transduction; Sirolimus; Tongue Neoplasms; TOR Serine-Threonine Kinases; Tumor Cells, Cultured

Cetuximab, a monoclonal blocking antibody against the epidermal growth factor receptor EGFR, has been approved for the treatment of squamous cell carcinomas of the head and neck (HNSCC). However, only few patients display long-term responses, prompting the search for cetuximab resistance mechanisms and new therapeutic options enhancing cetuximab effectiveness.. Cetuximab-sensitive HNSCC cells were retro-engineered to express PIK3CA and RAS oncogenes. These cells and HNSCC cells harboring endogenous PIK3CA and RAS oncogenes were xenografted into mice (n = 10 per group) and studied for their biochemical, antitumor, antiangiogenic, and antilymphangiogenic responses to cetuximab and mTOR targeting agents. All P values are two-sided.. Cetuximab treatment of PIK3CA- and RAS-expressing HNSCC xenografts promoted an initial antitumor response, but all tumors relapsed within few weeks. In these tumors, cetuximab did not decrease the activity of mTOR, a downstream signaling target of EGFR, PIK3CA, and RAS. The combined administration of cetuximab and mTOR inhibitors exerted a remarkably increased antitumor activity, particularly in HNSCC cells that are resistant to cetuximab as a single agent. Indeed, cotargeting mTOR together with cetuximab caused a rapid tumor collapse of both PIK3CA- and RAS-expressing HNSCC xenografts (P < .001), concomitant with reduced proliferation (P < .001) and lymphangiogenesis (P < .001).. The presence of PIK3CA and RAS mutations and other alterations affecting the mTOR pathway activity in HNSCC could be exploited to predict the potential resistance to cetuximab, and to select the patients that may benefit the most from the concomitant administration of cetuximab and PI3K and/or mTOR inhibitors as a precision molecular therapeutic option for HNSCC patients.

Topics: Animals; Antibiotics, Antineoplastic; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cetuximab; Class I Phosphatidylinositol 3-Kinases; Drug Resistance, Neoplasm; ErbB Receptors; Head and Neck Neoplasms; Humans; Lymphangiogenesis; Mice; Mutation; Phosphatidylinositol 3-Kinases; ras Proteins; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Autophagy is a catabolic process involving the degradation of cells' own unnecessary, injured, or aged proteins and recycling of degraded products to maintain hemostasis. Recently, studies indicated that autophagy plays a crucial role in cancer development. However, the role of autophagy in tongue squamous cell carcinoma (TSCC) has not been well documented. This study aims to assess the expression of autophagy-related protein and investigate its effect on TSCC.. Archival 50 TSCC samples were enrolled. Immunohistochemistry were performed to examine the expression of Beclin1 and LC3. Statistical analyses were carried out to assess the associations among clinicopathologic parameters. In vitro, cells were treated with rapamycin or 3-MA. Then, qPCR, western blot and immunofluorescence were performed to detect the expression of Beclin1 and LC3. Transmission electron microscopy was utilized to identify autophagsomes. For functional analysis, cell proliferation and cell cycle were evaluated with MTT assay and flow cytometer, respectively. At last, cell migration and invasion potentials were assessed by wound healing assay and transwell assay.. We confirmed that down-regulation of Beclin1 and LC3 is a frequent event in TSCC. Then, we demonstrated that decreased expression of Beclin1 was associated with T stage, clinical stage and differentiation. Furthermore, we showed that activation of autophagy by rapamycin suppressed proliferation, migration and invasion while inhibition of autophagy by 3-MA promoted proliferation, migration and invasion in TSCC cells.. Taken together, these data suggest that autophagy plays a pivotal role in the progression of TSCC.

Topics: Adenine; Antibiotics, Antineoplastic; Apoptosis Regulatory Proteins; Autophagy; Beclin-1; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Disease Progression; Female; Humans; Male; Membrane Proteins; Microtubule-Associated Proteins; Middle Aged; Neoplasm Invasiveness; Neoplasm Staging; Phagosomes; Sirolimus; Tongue Neoplasms

The transcription factor SOX2 is essential for the maintenance of embryonic stem cells and normal development of the esophagus. Our previous study revealed that the SOX2 gene is an amplification target of 3q26.3 in esophageal squamous cell carcinoma (ESCC), and that SOX2 promotes ESCC cell proliferation in vitro. In the present study, we aimed to identify the mechanisms by which SOX2 promotes proliferation of ESCC cells. Using a phosphoprotein array, we assayed multiple signaling pathways activated by SOX2 and determined that SOX2 activated the AKT/mammalian target of rapamycin complex 1 (mTORC1) signaling pathway. LY294002, an inhibitor of phosphatidylinositol 3-kinase, and rapamycin, an inhibitor of mTORC1, suppressed the ability of SOX2 to enhance proliferation of ESCC cells in vitro. Effects of SOX2 knockdown, including reduced levels of phosphorylated AKT and decreased ESCC cell proliferation, were reversed with constitutive activation of AKT with knockdown of phosphatase and tensin homolog. In mouse xenografts, SOX2 promoted in vivo tumor growth of ESCC, which was dependent on AKT/mTORC1 activation. LY294002 suppressed the ability of SOX2 to enhance tumor growth of ESCC by reducing cell proliferation, but not by enhancing apoptosis. Furthermore, tissue microarray analysis of 61 primary ESCC tumors showed a positive correlation between expression levels of SOX2 and phosphorylated AKT. Our findings suggest that SOX2 promotes in vivo tumor growth of ESCC through activation of the AKT/mTORC1 signaling pathway, which enhances cell proliferation.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Chromones; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Heterografts; Humans; Male; MAP Kinase Signaling System; Mechanistic Target of Rapamycin Complex 1; Mice; Mice, Inbred BALB C; Mice, Nude; Morpholines; Multiprotein Complexes; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; SOXB1 Transcription Factors; TOR Serine-Threonine Kinases

Ultraviolet B (UVB) in the sun light is a major cause of skin damage, which accompanies complex alterations in irradiated skin cells, including DNA lesions, oxidative stress, inflammation and caspase activation. The protection against UVB damage requires multiple interruptions such as repair of the DNA lesions, scavenging of the reactive oxygen species (ROS), repression of the inflammation and others. Silibinin is suggested as an anti-UVB reagent, but the underlying mechanisms have not been fully elucidated. In this study, we found a role of autophagy in the anti-UVB effect of silibinin in A431 cells. Autophagy was reduced after UVB-irradiation while restored by silibinin through the suppression of the IGF-1R signalling pathway. The protective effect of silibinin in UVB-irradiated A431 cells was further enhanced by pre-treatment with an autophagy inducer, rapamycin, while it was reversed by an autophagy inhibitor, wortmannin, indicating that elevated autophagy contributed to the cell survival. Consistently, cell apoptosis was augmented by siRNAs targeting Beclin 1 and Atg5, supporting the hypothesis that autophagy induced by silibinin plays a protective role against UVB-induced epidermal apoptosis.

Topics: Androstadienes; Apoptosis; Apoptosis Regulatory Proteins; Autophagy; Autophagy-Related Protein 5; Beclin-1; Carcinoma, Squamous Cell; Humans; Membrane Proteins; Microtubule-Associated Proteins; Receptor, IGF Type 1; RNA, Small Interfering; Silybin; Silymarin; Sirolimus; Tumor Cells, Cultured; Ultraviolet Rays; Wortmannin

Topics: Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; ErbB Receptors; Female; Head and Neck Neoplasms; Humans; Male; Neoplasm Recurrence, Local; Protein Kinase Inhibitors; Quinazolines; Sirolimus

Human papillomavirus (HPV)-related head and neck squamous cell carcinoma (HNSCC) incidence is increasing at a near epidemic rate. We investigated whether the mammalian (or mechanistic) target of rapamycin (mTOR) inhibitor, rapamycin, can be used as a concurrent agent to standard-of-care cisplatin/radiation therapy (CRT) to attenuate tumor lactate production, thus enhancing CRT-induced immune-mediated clearance of this antigenic tumor type. A C57Bl/6-derived mouse oropharyngeal epithelial cell line retrovirally transduced with HPV type 16 E6/E7 and human squamous cell carcinoma cell lines were evaluated for their response to rapamycin in vitro with proliferation assays, Western blots, and lactate assays. Clonogenic assays and a preclinical mouse model were used to assess rapamycin as a concurrent agent to CRT. The potential of rapamycin to enhance immune response through lactate attenuation was assessed using quantitative tumor lactate bioluminescence and assessment of cell-mediated immunity using E6/E7-vaccinated mouse splenocytes. Rapamycin alone inhibited mTOR signaling of all cancer cell lines tested in vitro and in vivo. Furthermore, rapamycin administered alone significantly prolonged survival in vivo but did not result in any long-term cures. Given concurrently, CRT/rapamycin significantly enhanced direct cell killing in clonogenic assays and prolonged survival in immunocompromised mice. However, in immunocompetent mice, concurrent CRT/rapamycin increased long-term cures by 21%. Preliminary findings suggest that improved survival involves increased cell killing and enhanced immune-mediated clearance in part due to decreased lactate production. The results may provide rationale for the clinical evaluation of mTOR inhibitors concurrent with standard-of-care CRT for treatment of HPV-positive HNSCC.

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cisplatin; Head and Neck Neoplasms; Human papillomavirus 16; Humans; Immunity, Cellular; Lactates; Male; Mice; Mice, Inbred C57BL; Papillomavirus Infections; Signal Transduction; Sirolimus; Squamous Cell Carcinoma of Head and Neck; Survival Rate; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Tumor dissemination to cervical lymph nodes via lymphatics represents the first step in the metastasis of head and neck squamous cell carcinoma (HNSCC) and is the most significant predictor of tumor recurrence decreasing survival by 50%. The lymphatic suppressing properties of mTOR inhibitors are not yet well understood.. Lymphatic inhibiting effects of rapamycin were evaluated in vitro using two lymphatic endothelial cell (LEC) lines. An orthotopic mouse model of HNSCC (OSC-19 cells) was used to evaluate anti-lymphangiogenic effects of rapamycin in vivo. The incidence of cervical lymph node metastases, numbers of tumor-free lymphatic vessels and those invaded by tumor cells in mouse lingual tissue, and expression of pro-lymphangiogenic markers were assessed.. Rapamycin significantly decreased lymphatic vascular density (p = 0.027), reduced the fraction of lymphatic vessels invaded by tumor cells in tongue tissue (p = 0.013) and decreased metastasis-positive lymph nodes (p = 0.04). Rapamycin also significantly attenuated the extent of metastatic tumor cell spread within lymph nodes (p < 0.0001). We found that rapamycin significantly reduced LEC proliferation and was correlated with decreased VEGFR-3 expression in both LEC, and in some HNSCC cell lines.. The results of this study demonstrate anti-lymphangiogenic properties of mTOR inhibitors in HNSCC. mTOR inhibitors suppress autocrine and paracrine growth stimulation of tumor and lymphatic endothelial cells by impairing VEGF-C/VEGFR-3 axis and release of soluble VEGFR-2. In a murine HNSCC orthotopic model rapamycin significantly suppressed lymphovascular invasion, decreased cervical lymph node metastasis and delayed the spread of metastatic tumor cells within the lymph nodes.

Topics: Animals; Antineoplastic Agents; Apoptosis; Blotting, Western; Carcinoma, Squamous Cell; Cell Proliferation; Disease Models, Animal; Head and Neck Neoplasms; Humans; Lymphangiogenesis; Lymphatic Metastasis; Male; Mice; Mice, SCID; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Topics: Carcinoma, Squamous Cell; Female; Humans; Male; Sirolimus; Skin Neoplasms

Topics: Carcinoma, Squamous Cell; Female; Humans; Male; Sirolimus; Skin Neoplasms

Topics: Antineoplastic Agents; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Graft Rejection; Humans; Immunosuppressive Agents; Lung Transplantation; Male; Middle Aged; Risk Factors; Sirolimus; Skin Neoplasms; Tacrolimus; Treatment Outcome

Mammalian Target of Rapamycin (mTOR) inhibitors, such as sirolimus and everolimus, have been shown to reduce cutaneous carcinogenesis in organ-transplant recipients requiring for immunosuppressive treatment to prevent from allograft rejection. Clinical observations suggest that cutaneous squamous cell carcinomas (SCC) are more sensitive than basal cell carcinomas (BCC) to the antitumoral effect of these inhibitors.. To investigate if the different response of SCC and BCC to mTOR inhibitors can be explained by differential expression of molecules involved in the mTOR signaling pathway.. The expression of phospho-mTOR was immunohistocemically studied in specimens of cutaneous SCC and BCC. Results. All 15 SCCs expressed significant cytoplasmic phospho-mTOR immunoreactivity; by contrast, 12/13 BCC were completely negative, only one BCC exhibited weak phospho-mTOR immunoreactivity.. The considerably higher expression of phospho-mTOR in SCC compared to BCC is a likely explanation for their higher sensitivity to mTOR inhibitors.

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Everolimus; Humans; Immunosuppressive Agents; Organ Transplantation; Sirolimus; Skin Neoplasms; TOR Serine-Threonine Kinases

Heat shock protein 90 (HSP90) and mammalian target of rapamycin (mTOR) are involved in the molecular pathogenesis of advanced oral squamous cell carcinoma. HSP90 inhibitors are capable of effectively interfering with multiple signaling pathways, including the mTOR signaling pathway. However, the combined effects of HSP90 and mTOR inhibitors on oral squamous cell carcinoma are still unknown. In this study, we investigated the dual treatment of the novel HSP90 inhibitor NVP-AUY922 and temsirolimus against oral squamous cell carcinoma.. The effect of the combination of NVP-AUY922 and temsirolimus on oral squamous cell carcinoma in vitro and in vivo was determined by MTS assay and mouse xenograft models. The effect of the combination on angiogenesis was determined by tube formation assay and angioreactor.. The combination treatment of NVP-AUY922 and temsirolimus significantly inhibited the proliferation of SAS oral squamous cell carcinoma cells in vitro and suppressed the growth of oral squamous cell carcinoma xenografts in vivo. We have clearly shown that the combination treatment of NVP-AUY922 and temsirolimus inhibited vascular formation both in vitro and in vivo. Moreover, the combination treatment of NVP-AUY922 and temsirolimus prolonged the survival rate in mice xenografted with oral squamous cell carcinoma.. Here, we showed the activity of a combination of mTOR and HSP90 inhibitors for the treatment of advanced oral squamous carcinoma.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Squamous Cell; Cell Line, Tumor; Cells, Cultured; Female; Head and Neck Neoplasms; HSP90 Heat-Shock Proteins; Human Umbilical Vein Endothelial Cells; Humans; Isoxazoles; Mice; Mice, Nude; Molecular Targeted Therapy; Mouth Neoplasms; Neoplasm Proteins; Neovascularization, Pathologic; Protein Kinase Inhibitors; Random Allocation; Resorcinols; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Esophageal squamous cell carcinoma (ESCC) remains one of the most aggressive cancers with poor prognosis regardless of a several reports that indicate a better therapeutic efficacy using some new chemotherapeutic agents. Recent drug development has contributed to an improved specificity to suppress mTOR activity by which many types of malignancies can be explosively progressed. Temsirolimus (CCI-779, TricelTM) is one of recently synthesized analogs of rapamycin and has provided better outcomes for patients with renal cell carcinoma. In this study, we experimentally evaluated an efficacy of targeting mTOR by temsirolimus for ESCC treatment, with an assessment of its survival advantage using an advanced ESCC animal model. First, we confirmed that the expression of phosphorylated mTOR was increased in 46 of 58 clinical ESCC tumor tissues (79.3%) and appeared to get strengthened with tumor progression. All of ESCC cell lines used in this study revealed an increase of mTOR phosphorylation, accompanied with the upregulation of hypoxia inducible factor-I α (HIF-1α), one of the critical effectors regulated by mTOR. Temsirolimus treatment apparently suppressed the activation of mTOR and its downstream effectors, resulting in the reduced ability of ESCC cell proliferation. Finally, the weekly administration of temsirolimus significantly diminished the size of subcutaneous tumors (vehicle, 3261.6 ± 722.0; temsirolimus, 599.2 ± 122.9; p = 0.007) in nude mice and effectively prolonged orthotopic esophageal cancer-bearing mice (median survival periods: control, 31 d; temsirolimus, 43 d; p = 0.0024). These data suggests that targeting mTOR by temsirolimus may become a therapeutic alternative for esophageal cancer, with a contribution to a better outcome.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Enzyme Activation; Esophageal Neoplasms; Humans; Mice; Protein Kinase Inhibitors; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Tumor Burden; Xenograft Model Antitumor Assays

The mammalian target of rapamycin (mTOR) protein is important for cellular growth and homeostasis. The presence and prognostic significance of inappropriate mTOR activation have been reported for several cancers. Mammalian target of rapamycin inhibitors, such as everolimus (RAD001), are in development and show promise as anti-cancer drugs; however, the therapeutic effect of everolimus on oesophageal squamous cell carcinoma (OSCC) remains unknown.. Phosphorylation of mTOR (p-mTOR) was evaluated in 167 resected OSCC tumours and 5 OSCC cell lines. The effects of everolimus on the OSCC cell lines TE4 and TE11 in vitro and alone or in combination with cisplatin on tumour growth in vivo were evaluated.. Mammalian target of rapamycin phosphorylation was detected in 116 tumours (69.5%) and all the 5 OSCC cell lines. Everolimus suppressed p-mTOR downstream pathways, inhibited proliferation and invasion, and induced apoptosis in both TE4 and TE11 cells. In a mouse xenograft model established with TE4 and TE11 cells, everolimus alone or in combination with cisplatin inhibited tumour growth.. The mTOR pathway was aberrantly activated in most OSCC tumours. Everolimus had a therapeutic effect both as a single agent and in combination with cisplatin. Everolimus could be a useful anti-cancer drug for patients with OSCC.

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cisplatin; Esophageal Neoplasms; Everolimus; Humans; Mice; Phosphorylation; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

The incidence of head and neck squamous cell carcinomas (HNSCC) associated with human papillomavirus (HPV) infection has increased over the past decades in the United States. We aimed at examining the global impact of HPV-associated HNSCC and whether the established key role of mTOR activation in HNSCC is also observed in HPV(+) HNSCC lesions, thereby providing novel treatment options for HPV-associated HNSCC patients.. An international HNSCC tissue microarray (TMA) was used to analyze the expression of p16(INK4A), a surrogate for HPV infection, and Akt-mTOR pathway activation. Results were confirmed in a large collection of HPV(-) and HPV(+) HNSCC cases and in a cervical cancer (CCSCC) TMA. Observations were validated in HNSCC and CCSCC-derived cell lines, which were xenografted into immunodeficient mice for tumorigenesis assays.. Approximately 20% of all HNSCC lesions could be classified as HPV(+), irrespective of their country of origin. mTOR pathway activation was observed in most HPV(+) HNSCC and CCSCC lesions and cell lines. The preclinical efficacy of mTOR inhibition by rapamycin and RAD001 was explored in HPV(+) HNSCC and CCSCC tumor xenografts. Both mTOR inhibitors effectively decreased mTOR activity in vivo and caused a remarkable decrease in tumor burden. These results emphasize the emerging global impact of HPV-related HNSCCs and indicate that the activation of the mTOR pathway is a widespread event in both HPV(-) and HPV-associated HNSCC and CCSCC lesions.. The emerging results may provide a rationale for the clinical evaluation of mTOR inhibitors as a molecular targeted approach for the treatment of HPV-associated malignancies.

Topics: Animals; Blotting, Western; Carcinoma, Squamous Cell; Cell Line, Tumor; Cervix Uteri; Cohort Studies; Cyclin-Dependent Kinase Inhibitor p16; DNA, Viral; Female; Humans; Immunoenzyme Techniques; Mice; Mice, Nude; Mouth Neoplasms; Neoplasm Proteins; Papillomaviridae; Papillomavirus Infections; Polymerase Chain Reaction; Proto-Oncogene Proteins c-akt; Sirolimus; Tissue Array Analysis; TOR Serine-Threonine Kinases; Uterine Cervical Neoplasms; Xenograft Model Antitumor Assays

Head and neck squamous cell carcinoma (HNSCC) is a group of tumors known to be sensitive to chemotherapy and radiotherapy in patients who are treatment naive. However, when recurrences do occur, these tumors generally become resistant and objective responses to therapy at that point tend to be less effective. There has been an increasing interest in developing novel molecular-targeted agents that specifically modulate growth factor and signaling pathways that are unregulated in HNSCC tumor cells. Combinations of vascular endothelial growth factor and mammalian target of rapamycin inhibitors have been used in some types of neoplasms, but no such efforts have been made in HNSCC. In this study, we investigated the in vitro, in vivo, and clinical effects of the temsirolimus (mammalian target of rapamycin inhibitor, Tem) and bevacizumab (antivascular endothelial growth factor antibody, Bev) combination. In-vitro studies were carried out on the A431 human squamous epidermoid carcinoma cell line and in-vivo studies were carried out on A431 tumor cells implanted on female Nu/Nu*nuBR (athymic nude) mice. Also, the effectiveness of the Tem and Bev combination was tested clinically in two separate clinical cases of chemoresistant HNSCC. The in-vitro, in-vivo, and clinical results showed that this combination can be significantly effective. In conclusion, we discuss the theoretical basis of the molecular pharmacological interactions between Bev and Tem that could explain these good results. If the therapeutic index is ultimately well determined, the antitumor effect of Bev and Tem is very likely to yield fruitful results.

Topics: Animals; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Bevacizumab; Carcinoma, Squamous Cell; Cell Line, Tumor; Female; Head and Neck Neoplasms; Humans; Male; Mice; Mice, Nude; Middle Aged; Molecular Targeted Therapy; Sirolimus; Treatment Outcome; Xenograft Model Antitumor Assays

Topics: Carcinoma, Squamous Cell; Female; Graft Rejection; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Sirolimus; Skin Neoplasms

The mammalian target of rapamycin signaling pathway has been implicated in therapeutic resistance in several types of cancer. However, the significance of mammalian target of rapamycin activation in chemoradiotherapy sensitivity and its effect on the prognosis of esophageal squamous cell carcinoma treated with chemoradiotherapy remain unknown. However, this pathway is of particular interest because an effective inhibitor is available.. By using immunohistochemistry, phosphorylated mammalian target of rapamycin expression was examined in 77 patients with esophageal squamous cell carcinoma treated with preoperative chemoradiotherapy followed by surgery between 1999 and 2009, and correlated with treatment outcome. With the use of CE81T/VGH and TE2 cell lines, cells were treated with chemotherapy, temsirolimus (mammalian target of rapamycin inhibitor), or a combination of chemotherapy and temsirolimus, and investigated by 3-(4.5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.. Pathologic complete response rates were 42% and 16% in patients with negative and positive phosphorylated mammalian target of rapamycin expression, respectively (P = .01). The 3-year overall survivals were 57% and 30% in patients with negative and positive phosphorylated mammalian target of rapamycin expression, respectively (P = .005). Positive phosphorylated mammalian target of rapamycin expression was independently associated with inferior overall and disease-free survival. In patients who did not achieve pathologic complete response, postchemoradiotherapy esophagectomy specimens showed significantly higher phosphorylated mammalian target of rapamycin expression than pretreatment biopsy specimens. In cell lines, concomitant administration of temsirolimus enhanced the effect of chemotherapy.. Phosphorylated mammalian target of rapamycin expression is independently associated with the response to chemoradiotherapy and prognosis of patients with esophageal squamous cell carcinoma treated with preoperative chemoradiotherapy. Mammalian target of rapamycin inhibition can sensitize esophageal cancer cells to chemotherapy. Our results suggest the potential for mammalian target of rapamycin as a therapeutic target for patients with esophageal squamous cell carcinoma who receive multimodality treatment.

Topics: Adult; Aged; Biomarkers, Tumor; Biopsy; Carcinoma, Squamous Cell; Cell Line, Tumor; Chemoradiotherapy, Adjuvant; Chi-Square Distribution; Disease-Free Survival; Esophageal Neoplasms; Esophagectomy; Female; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Logistic Models; Male; Middle Aged; Multivariate Analysis; Neoadjuvant Therapy; Odds Ratio; Phosphorylation; Proportional Hazards Models; Protein Kinase Inhibitors; Retrospective Studies; Risk Assessment; Risk Factors; Sirolimus; Time Factors; TOR Serine-Threonine Kinases; Treatment Outcome

To assess the efficacy of rapamycin treatment in chemoprevention and chemotherapy of tumorigenesis in a genetically defined mouse model of head and neck squamous cell carcinoma (HNSCC).. Knockdown of Tgfbr1 and/or Pten using siRNA-mediated RNA interference was carried out in human HNSCC cell lines to analyze molecular changes in the mTOR pathway. Tgfbr1(flox/flox); Pten(flox/flox); K14-CreER(tam) mice were treated with oral gavage of tamoxifen for the conditional deletion of Tgfbr1 and Pten in oral mucosa, resulting in HNSCC. Tgfbr1 and Pten conditonal deletion (2cKO) mice were treated with rapamycin before or after the onset of HNSCC, and the efficacy of this treatment was assessed by determining tumor burden, longevity, and molecular analysis of the mTOR pathway. Molecular changes observed in human HNSCC cell lines and 2cKO mice were compared to identify key alterations in the mTOR pathway.. Knockdown of Tgfbr1 and/or Pten in human HNSCC cell lines resulted in activation of mTOR activity complex 1 and increased levels of survivin. Furthermore, we observed similar changes in HNSCC of the 2cKO mouse. In the human HNSCC tissue array, a loss of Tgfbr1 expression correlated with increased survivin levels. Chemopreventive rapamycin treatment significantly delayed the onset of the HNSCC tumors and prolonged survival in 2cKO mice. In addition, we also found that rapamycin had a therapeutic effect on squamous cell carcinomas in these mice. In 2cKO HNSCC tongue tumors, rapamycin treatment induced apoptosis, inhibited cell proliferation and phosphorylation of Akt and S6, and decreased survivin expression.. These findings indicate that tumorigenesis in 2cKO HNSCC is associated with activation of the Akt/mTOR/survivin pathway, and inhibition of this pathway by rapamycin treatment successfully ameliorates the onset and progression of tumorigenesis.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Disease Models, Animal; Head and Neck Neoplasms; Humans; Inhibitor of Apoptosis Proteins; Mice; Protein Serine-Threonine Kinases; PTEN Phosphohydrolase; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Signal Transduction; Sirolimus; Survivin; TOR Serine-Threonine Kinases

The epidermal growth factor receptor (EGFR) signaling pathway is frequently dysregulated in a variety of human malignancies. As a result, agents have been developed to selectively inhibit the tyrosine kinase function of EGFR (EGFR-TKI) for cancer therapy. However, the clinical efficacy of these drugs to date has been limited by both acquired and intrinsic resistance. Macroautophagy, a process of intracellular proteolysis, has been shown to be activated in response to EGFR targeted therapy. However, the specific role of the induction of autophagy remains controversial. Here we show that autophagy is induced in a dose-dependent manner by in vitro treatment of multiple cancer cell lines with EGFR-TKI. Additionally, we find that in cells highly resistant to EGFR-TKI, autophagy is not robustly activated and that co-treatment of these cells with rapamycin, a known inducer of autophagy, can partially restore sensitivity to EGFR-TKI. Finally, we demonstrate that, in resistant cell lines, EGFR-TKI sensitivity can be further inhibited by siRNA-mediated depletion of the critical autophagy protein ATG7. Thus, our data suggests that defective autophagy may be an EGFR-TKI resistance mechanism and that activation of autophagy may be a viable strategy to augment the cytotoxic effect of EGFR-TKIs.

Topics: Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Autophagy; Autophagy-Related Protein 7; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Survival; Cetuximab; ErbB Receptors; Erlotinib Hydrochloride; HeLa Cells; Humans; Lung Neoplasms; Mouth Neoplasms; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Quinazolines; RNA Interference; RNA, Small Interfering; Sirolimus; Ubiquitin-Activating Enzymes

The integrity of the epidermis and mucosal epithelia is highly dependent on resident self-renewing stem cells, which makes them vulnerable to physical and chemical insults compromising the repopulating capacity of the epithelial stem cell compartment. This is frequently the case in cancer patients receiving radiation or chemotherapy, many of whom develop mucositis, a debilitating condition involving painful and deep mucosal ulcerations. Here, we show that inhibiting the mammalian target of rapamycin (mTOR) with rapamycin increases the clonogenic capacity of primary human oral keratinocytes and their resident self-renewing cells by preventing stem cell senescence. This protective effect of rapamycin is mediated by the increase in expression of mitochondrial superoxide dismutase (MnSOD), and the consequent inhibition of ROS formation and oxidative stress. mTOR inhibition also protects from the loss of proliferative basal epithelial stem cells upon ionizing radiation in vivo, thereby preserving the integrity of the oral mucosa and protecting from radiation-induced mucositis.

Topics: Animals; Carcinoma, Squamous Cell; Cell Compartmentation; Cell Death; Cell Proliferation; Cells, Cultured; Cellular Senescence; Clone Cells; Cytoprotection; Epithelial Cells; Head and Neck Neoplasms; Humans; Keratinocytes; Mice; Mouth Mucosa; Mucositis; Oxidative Stress; Radiation Injuries; Radiation, Ionizing; Sirolimus; Stem Cells; Superoxide Dismutase; TOR Serine-Threonine Kinases

Squamous cell carcinoma of the skin (SCC) is the most frequent cancer in renal transplant recipients. Conversion to mammalian target of rapamycin inhibitors after diagnosis of SCC may reduce the incidence of recurrence of skin cancer. This retrospective study evaluated the outcome of renal transplant recipients followed by the Renal Unit with posttransplant diagnosis of SCC treated with conversion from calcineurin inhibitors (CNIs) to Everolimus (EVR) associated with low-dose cyclosporine. Eleven patients developed SCC at a median time from renal transplantation of 107 months (range 36-264). Five patients with creatinine clearance (CCl) below 40 mL/min before conversion developed end stage renal disease (two cases) or further deterioration of renal function (two cases); only one patient in this group maintained a stable renal function. The remaining six patients with a CC1 greater than 40 mL/min and proteinuria below 0.8 g/24 hours maintained a stable renal function after conversion to EVR at a median follow-up of 22 months (range 15-75). Conversion from CNIs to EVR has been proven safe, effective, and associated with low recurrence of SCC in patients with a CCl >40 mL/min. In the case of preexisting deterioration of renal function or significant proteinuria, conversion to EVR should be carefully evaluated.

Topics: Calcineurin Inhibitors; Carcinoma, Squamous Cell; Cyclosporine; Dose-Response Relationship, Drug; Everolimus; Humans; Kidney Transplantation; Sirolimus; Skin Neoplasms

Topics: Carcinoma, Squamous Cell; Graft Rejection; Humans; Immunosuppressive Agents; Kidney Transplantation; Risk Factors; Sirolimus; Skin Neoplasms

Topics: Aged; Antineoplastic Agents; Carcinoma, Squamous Cell; Drug Therapy, Combination; ErbB Receptors; Gefitinib; Humans; Lymphatic Metastasis; Male; Quinazolines; Sirolimus; Skin Neoplasms; Skin Ulcer; TOR Serine-Threonine Kinases

Topics: Calcineurin Inhibitors; Carcinoma, Squamous Cell; Female; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Sirolimus; Skin Neoplasms

Rapamycin is an allosteric inhibitor of mammalian target of rapamycin, and inhibits tumor growth and angiogenesis. Recent studies suggested a possibility that rapamycin renormalizes aberrant tumor vasculature and improves tumor oxygenation. The longitudinal effects of rapamycin on angiogenesis and tumor oxygenation were evaluated in murine squamous cell carcinoma (SCCVII) by electron paramagnetic resonance imaging (EPRI) and magnetic resonance imaging (MRI) to identify an optimal time after rapamycin treatment for enhanced tumor radioresponse. Rapamycin treatment was initiated on SCCVII solid tumors 8 days after implantation (500-750 mm(3)) and measurements of tumor pO(2) and blood volume were conducted from day 8 to 14 by EPRI/MRI. Microvessel density was evaluated over the same time period by immunohistochemical analysis. Tumor blood volume as measured by MRI significantly decreased 2 days after rapamycin treatment. Tumor pO(2) levels modestly but significantly increased 2 days after rapamycin treatment; whereas, it decreased in non-treated control tumors. Furthermore, the fraction of hypoxic area (pixels with pO(2)<10 mm Hg) in the tumor region decreased 2 days after rapamycin treatments. Immunohistochemical analysis of tumor microvessel density and pericyte coverage revealed that microvessel density decreased 2 days after rapamycin treatment, but pericyte coverage did not change, similar to what was seen with anti-angiogenic agents such as sunitinib which cause vascular renormalization. Collectively, EPRI/MRI co-imaging can provide non-invasive evidence of rapamycin-induced vascular renormalization and resultant transient increase in tumor oxygenation. Improved oxygenation by rapamycin treatment provides a temporal window for anti-cancer therapies to realize enhanced response to radiotherapy.

Topics: Angiogenesis Inhibitors; Animals; Biomarkers; Carcinoma, Squamous Cell; Diagnostic Imaging; Electron Spin Resonance Spectroscopy; Immunohistochemistry; Magnetic Resonance Imaging; Mice; Mice, Inbred C3H; Microcirculation; Neovascularization, Pathologic; Oxygen; Sirolimus; TOR Serine-Threonine Kinases; Tumor Microenvironment

Cancer after heart transplant is recognized as a leading cause of morbidity and mortality. A man's clinical course after receiving a heart transplant is described, with emphasis on important clinical considerations in the care of heart transplant recipients.

Topics: Azathioprine; Carcinoma, Squamous Cell; Drug Therapy, Combination; Fatal Outcome; Glucocorticoids; Heart Transplantation; Humans; Immunosuppressive Agents; Kidney Transplantation; Lung Neoplasms; Male; Mediastinal Neoplasms; Middle Aged; Mycophenolic Acid; Neoplasm Recurrence, Local; Pleural Neoplasms; Prednisone; Sirolimus; Skin Neoplasms; Tacrolimus

FOXO3a, a well-known transcriptional regulator, controls a wide spectrum of biological processes. The phosphoinositide-3-kinase (PI3K)/Akt signaling pathway inactivates FOXO3a via phosphorylation-induced nuclear exclusion and degradation. A loss or gain of FOXO3a activity has been correlated with efficiency of chemotherapies in various cancers including oral squamous cell carcinoma (OSCC). Therefore, in the current study, we have investigated the FOXO3a activity modulating and antitumor effects of rapamycin and cisplatin in OSCC cells. Cisplatin inhibited proliferation and induced apoptosis in a dose-dependent way in OSCC Tca8113 cells. Rapamycin alone had no effect on cell proliferation and apoptosis. Rapamycin downregulated the expression of S-phase kinase associated protein-2 (Skp2) and increased the FOXO3a protein stability but induced the upregulation of feedback Akt activation-mediated FOXO3a phosphorylation. Cisplatin decreased the phosphorylation of FOXO3a via Akt inhibition. Rapamycin combined with cisplatin as its feedback Akt activation inhibitor revealed the most dramatic FOXO3a nuclear localization and reactivation with the prevention of its feedback loop and exposed significant synergistic effects of decreased cell proliferation and increased apoptosis in vitro and decreased tumor size in vivo. Furthermore, the downstream effects of FOXO3a reactivation were found to be accumulation of p27 and Bim. In conclusion, rapamycin/cisplatin combination therapy boosts synergistic antitumor effects through the significant FOXO3a reactivation in OSCC cells. These results may represent a novel mechanism by which rapamycin/cisplatin combination therapy proves to be a potent molecular-targeted strategy for OSCC.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cisplatin; Cyclin-Dependent Kinase Inhibitor p27; Dose-Response Relationship, Drug; Drug Synergism; Forkhead Box Protein O3; Forkhead Transcription Factors; Humans; Male; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Nude; Molecular Targeted Therapy; Phosphorylation; Protein Kinase Inhibitors; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; S-Phase Kinase-Associated Proteins; Sirolimus; Time Factors; Tongue Neoplasms; Tumor Burden; Xenograft Model Antitumor Assays

Bladder cancer and head and neck squamous cell carcinoma (HNSCC) are frequent but lack efficient therapies especially in advanced disease. Almost no studies on mTOR function and inhibition in these tumor entities have been reported. We examined the gene and protein expression levels of mTOR and its activated form (pmTOR) in three human bladder carcinoma cell lines (RT-4, T24, EJ28) and three HNSCC cell lines (PCI-1, PCI-13, BHY). Furthermore, the consequences of mTOR inhibition by mTOR-specific siRNAs and the mTOR inhibitor temsirolimus were analysed in vitro using immunohistochemical Ki-67 staining, mTOR and pmTOR western blot analysis, MTT assay, as well as cell cycle analysis with flow cytometry. Especially pmTOR protein expression levels showed marked differences between cell lines. siRNA transfection was associated with dose-dependent target protein reduction but not proliferation inhibition or apoptosis. On the contrary, temsirolimus significantly reduced cell viability and induced apoptosis and cell cycle arrest. According to these data, bladder cancer and HNSCC are promising tumor entities for mTOR inhibition with temsirolimus.

Topics: Antineoplastic Agents; Carcinoma, Squamous Cell; Cell Cycle; Cell Proliferation; Drug Evaluation, Preclinical; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; Humans; Protein Kinase Inhibitors; RNA, Small Interfering; Sirolimus; TOR Serine-Threonine Kinases; Tumor Cells, Cultured; Urinary Bladder Neoplasms

We previously reported on head and neck tumor xenografts that the tumor regression induced by a triple combination of irradiation (RT), anti-EGFR and anti-angiogenic therapies was followed, after treatment arrest, by tumor re-growth characterized by activation of the AKT signaling pathway. Since mTOR is the main AKT-related messenger, the aim of this study was to add the mTOR inhibitor temsirolimus to a tri-therapy with RT plus anti-EGFR and anti-angiogenic drugs in order to improve anti-tumor effects. The human head and neck cancer cell line CAL33 (over-expressing EGFR and secreting VEGF-A) was xenografted in nude mice. Treatment (20 mice per treatment group) was administered for 2 weeks and consisted of either vehicle (control), temsirolimus (5mg/kg i.p. five times a week), tri-therapy with RT (6 Gy three times a week) combined with cetuximab (0.5mg/kg i.p. five times a week) and bevacizumab (5mg/kg i.p. five times a week) or the temsirolimus-tri-therapy association. The time to reach a tumor volume of 2000 mm(3) was significantly different between the four treatment groups (Log Rank p<0.0001), with a median of 29.5, 44.5, 67.0 and 70.0 days for control, temsirolimus, tri-therapy and combination groups, respectively. The combination of temsirolimus plus tri-therapy produced the longest growth-inhibiting effects (tri-therapy versus combination, p=0.01). No significant interaction was observed between temsirolimus and the tri-therapy, suggesting that temsirolimus, on the one hand, and RT-cetuximab-bevacizumab, on the other, exert additive effects on tumor growth inhibition. These decreases observed on tumor growth were corroborated by the parallel decreases observed on tumor proliferation (Ki67) and on anti-apoptotic markers (Bcl2). These results suggest that temsirolimus exhibits synergistic antiproliferative effects when administered in combination with irradiation, anti-EGFR and anti-angiogenic therapies in head and neck cancer patients.

Topics: Angiogenesis Inhibitors; Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Bevacizumab; Carcinoma, Squamous Cell; Cell Line, Tumor; Cetuximab; Combined Modality Therapy; Drug Synergism; Head and Neck Neoplasms; Humans; Mice; Mice, Nude; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Rapamycin is an mTOR inhibitor with preclinical efficacy in squamous cell carcinoma of the head and neck (SCCHN). However, mTOR inhibitors also increase Akt activity in SCCHN cell lines, which would promote survival and oncogenesis. Enzastaurin is an AGC kinase inhibitor with nanomolar inhibitory concentrations for Akt and protein kinase C (PKC). Moreover, Akt and PKC inhibitors have demonstrated efficacy in SCCHN.. We hypothesized that the combination of rapamycin and enzastaurin would be more effective than either agent alone.. Rapamycin and enzastaurin generally inhibited putative targets in SCCHN cell lines in culture. In mice xenografted with CAL27 cells, rapamycin and enzastaurin produced growth delay. In contrast, the combination of rapamycin and enzastaurin caused regression of CAL27 tumors with evidence of inhibition of putative targets, survival, angiogenesis and proliferation.. These data demonstrate that the combination of rapamycin and enzastaurin disrupts critical oncogenic pathways in SCCHN and has efficacy in preclinical models.

Topics: Animals; Antibiotics, Antineoplastic; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Drug Synergism; Female; Head and Neck Neoplasms; In Situ Nick-End Labeling; Indoles; MAP Kinase Signaling System; Mice; Mice, Nude; Neoplasm Transplantation; Neovascularization, Pathologic; Protein Kinase Inhibitors; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Tumor Cells, Cultured

Recent reports have shown that cancer stem cells exist in many malignancies. Side population (SP) cells are used to enrich cancer stem-like cells in many cell lines and fresh tumor specimens. In this study, we cultured primary esophageal squamous cell carcinoma (ESCC) cells from ESCC tissue specimens. SP cells from primary ESCC cells were more resistant to chemotherapeutic reagents and formed more colonies in vitro than non-SP cells. In addition, xenograft experiments revealed that SP cells were more tumorigenic in vivo. Further results indicated that the PI3K/Akt pathway is essential to SP cells through the regulation of ABCG2 transporter function. Furthermore, PTEN, rather than mTOR, was found to be involved in SP cell regulation in primary ESCC cells. These findings reveal that SP cells are enriched for cancer stem-like cells in primary ESCC cells and that the PTEN/PI3K/Akt pathway regulates this stem-like population. This study indicates that SP cells in primary culture cells from tissue specimens could be a promising model for cancer stem cell research and may help researchers develop novel therapeutic strategies or efficient drugs that target ESCC stem-like cells.

Topics: Antineoplastic Agents; ATP-Binding Cassette Transporters; Carcinoma, Squamous Cell; Cell Proliferation; Chromones; Drug Resistance, Neoplasm; Enzyme Inhibitors; Esophageal Neoplasms; Humans; Morpholines; Neoplastic Stem Cells; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; RNA Interference; Side-Population Cells; Signal Transduction; Sirolimus; Tumor Cells, Cultured

To evaluate antitumor efficacy of the generic mammalian target of rapamycin (mTOR) inhibitor sirolimus in preclinical animal models of head and neck squamous cell carcinoma (HNSCC) and compare its effects with those of the patented analogue temsirolimus.. In vivo study.. To develop xenograft established tumor model (ETM) of HNSCC, FaDu cells were injected subcutaneously into nude mice. When tumors reached 50 to 60 mm(3), mice were randomized into five groups and treated daily intraperitoneally with sirolimus at various doses for 5 days per week for 3 weeks. Tumor volumes were measured. The results were compared with historical data on temsirolimus effects. In the minimal residual disease (MRD) model, surgical wounds were created and FaDu cells implanted. After 72 hours, animals were randomized into two groups and were injected intraperitoneally with 0 or 5 mg/kg sirolimus for 5 days per week for 30 days.. In the ETM, sirolimus significantly inhibited tumor growth (P < .01), although there was no overall significant difference in tumor growth inhibition between sirolimus and temsirolimus. In the MRD model, sirolimus significantly suppressed growth of tumors (P < .001) and improved survival compared with controls (P < .01). There was a significant decrease in pS6 expression, indicating mTOR inhibition.. In this study, we demonstrate that the generic mTOR inhibitor sirolimus shows potent antitumor activity in HNSCC and produces comparable effects to the patent drug temsirolimus. Sirolimus has the potential of serving as an economic and comparative targeted agent to temsirolimus in the treatment of HNSCC.

Topics: Animals; Antineoplastic Agents; Carcinoma, Squamous Cell; Disease Models, Animal; Head and Neck Neoplasms; Mice; Mice, Inbred BALB C; Mice, Nude; Sirolimus

To investigate the synergistic effects of rapamycin and cisplatin on head and neck squamous cancer cells regulated by chemokine (C-C motif) ligand 19 (CCL19).. The role of rapamycin and cisplatin was detected on cell-cycle and apoptosis in CCL19 induced PCI-4B and PCI-37B cells by methyl thiazolyl tetrazolium (MTT) and flow cytometry (FCM). Dose-effect relationship parameters and combination index (CI) were calculated on the median-effect equation and multiple drug effect equation using computer software CalcuSyn. Statistical analysis was performed by the unpaired student's t-test.. Rapamycin and cisplatin could respectively increase the growth arrest, the proportion of G(1) phase and apoptosis of CCL19 induced cancer cells (P < 0.05). Under inhibitory concentration 50% (IC(50)), CI was less than 1, and in IC(75), it was more than 1 in PCI-4B cells. In PCI-37B cells, under IC(75), CI was less than 1, and in IC(90), it was more than 1.. Rapamycin and cisplatin can inhibit CCL19-regulated PCI-4B and PCI-37B cells' survival. The two drugs have synergistic effects when used in combination.

Topics: Antibiotics, Antineoplastic; Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Chemokine CCL19; Cisplatin; Drug Synergism; Head and Neck Neoplasms; Humans; Sirolimus

Recently it has been shown that radiation induces migration of glioma cells and facilitates a further spread of tumor cells locally and systemically. The aim of this study was to evaluate whether radiotherapy induces migration in head and neck squamous cell carcinoma (HNSCC). A further aim was to investigate the effects of blocking the epidermal growth factor receptor (EGFR) and its downstream pathways (Raf/MEK/ERK, PI3K/Akt) on tumor cell migration in vitro.. Migration of tumor cells was assessed via a wound healing assay and proliferation by a MTT colorimeritric assay using 3 HNSCC cell lines (BHY, CAL-27, HN). The cells were treated with increasing doses of irradiation (2 Gy, 5 Gy, 8 Gy) in the presence or absence of EGF, EGFR-antagonist (AG1478) or inhibitors of the downstream pathways PI3K (LY294002), mTOR (rapamycin) and MEK1 (PD98059). Biochemical activation of EGFR and the downstream markers Akt and ERK were examined by Western blot analysis.. In absence of stimulation or inhibition, increasing doses of irradiation induced a dose-dependent enhancement of migrating cells (p < 0.05 for the 3 HNSCC cell lines) and a decrease of cell proliferation (p < 0.05 for the 3 HNSCC cell lines). The inhibition of EGFR or the downstream pathways reduced cell migration significantly (almost all p < 0.05 for the 3 HNSCC cell lines). Stimulation of HNSCC cells with EGF caused a significant increase in migration (p < 0.05 for the 3 HNSCC cell lines). After irradiation alone a pronounced activation of EGFR was observed by Western blot analysis.. Our results demonstrate that the EGFR is involved in radiation induced migration of HNSCC cells. Therefore EGFR or the downstream pathways might be a target for the treatment of HNSCC to improve the efficacy of radiotherapy.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Dose-Response Relationship, Radiation; Epidermal Growth Factor; ErbB Receptors; Flavonoids; Gamma Rays; Head and Neck Neoplasms; Humans; MAP Kinase Kinase 1; MAP Kinase Signaling System; Phosphatidylinositol 3-Kinases; Quinazolines; Signal Transduction; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Tyrphostins

Despite our improved understanding of cancer, the 5-year survival rate for head and neck squamous cell carcinomas (HNSCC) patients remains relatively unchanged at 50% for the past three decades. HNSCCs often metastasize to locoregional lymph nodes, and lymph node involvement represents one of the most important prognostic factors of poor clinical outcome. Among the multiple dysregulated molecular mechanism in HNSCCs, emerging basic, preclinical, and clinical findings support the importance of the mTOR signaling route in HNSCC progression. Indeed, we observed here that the activation of mTOR is a widespread event in clinical specimens of HNSCCs invading locoregional lymph nodes. We developed an orthotopic model of HNSCC consisting of the implantation of HNSCC cells into the tongues of immunocompromised mice. These orthotopic tumors spontaneously metastasize to the cervical lymph nodes, where the presence of HNSCC cells can be revealed by histologic and immunohistochemical evaluation. Both primary and metastatic experimental HNSCC lesions exhibited elevated mTOR activity. The ability to monitor and quantitate lymph node invasion in this model system enabled us to explore whether the blockade of mTOR could impact HNSCC metastasis. We found that inhibition of mTOR with rapamycin and the rapalog RAD001 diminished lymphangiogenesis in the primary tumors and prevented the dissemination of HNSCC cancer cells to the cervical lymph nodes, thereby prolonging animal survival. These findings may provide a rationale for the future clinical evaluation of mTOR inhibitors, including rapamycin and its analogues, as part of a molecular-targeted metastasis preventive strategy for the treatment of patients with HNSCC.

Topics: Animals; Carcinoma, Squamous Cell; Everolimus; Female; Head and Neck Neoplasms; Humans; Lymphangiogenesis; Lymphatic Metastasis; Mice; Mice, SCID; Sirolimus; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Transcription Factors

Metastatic squamous cell carcinoma (SCC) of the head and neck expresses chemokine receptor 7 (CCR7), which activates phosphoinositide-3 kinase (PI3K) to promote invasion and survival of SCC cells in the head and neck. We hypothesised that mammalian target of rapamycin (mTOR) may be the downstream molecule of the CCR7-PI3K pathway. Results have shown that interaction between CCR7 and its ligand CCL19 induces the phosphorylation of mTOR and its target p70s6k. This phosphorylation is abolished by inhibition of CCR7 and PI3K/Akt, indicating that mTOR is involved in the CCR7-PI3K cascade. The inhibitors of mTOR and CCR7-PI3K also lead to a significant increase in CCL19-induced death, apoptosis, and cell-cycle arrest of metastatic SCC cells in the head and neck. Taken together, our data indicate the important part played by mTOR in CCR7-induced survival of such SCC cells.

Topics: Antibodies, Monoclonal; Antineoplastic Agents; Apoptosis; Blotting, Western; Carcinoma, Squamous Cell; Cell Cycle; Cell Death; Cell Line, Tumor; Cell Survival; Chemokine CCL19; Chromones; Cisplatin; Coloring Agents; Enzyme Inhibitors; Flow Cytometry; Head and Neck Neoplasms; Humans; Intracellular Signaling Peptides and Proteins; Morpholines; Neoplasm Invasiveness; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Polymerase Chain Reaction; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Receptors, CCR7; Ribosomal Protein S6 Kinases, 70-kDa; Sirolimus; Tetrazolium Salts; Thiazoles; TOR Serine-Threonine Kinases

Accumulating evidences have demonstrated that mTOR pathway has a central role not only in cell growth but also in invasion and metastasis of cancers. Here we reported that rapamycin or cisplatin alone inhibited significantly the tumor growth and their combination had the strongest anticancer effect on transplantable tumor growth of human ESCC cell line EC9706 in nude mice. Furthermore, western blots, RT-PCR and TUNEL assay revealed that rapamycin specifically blocked mTOR pathway and induced apoptosis of ESCC cells in vivo. These findings indicate a rationale for using mTOR inhibitors as a mechanism-based therapeutic approach to patients with ESCC.

Topics: Aged; Animals; Antineoplastic Agents; Apoptosis; Blotting, Western; Carcinoma, Squamous Cell; Cell Proliferation; Cisplatin; Esophageal Neoplasms; Female; Humans; Immunohistochemistry; In Situ Nick-End Labeling; Intracellular Signaling Peptides and Proteins; Male; Mice; Mice, Nude; Middle Aged; Neoplasms, Experimental; Protein Serine-Threonine Kinases; Reverse Transcriptase Polymerase Chain Reaction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Topics: Adult; Aged; Calcineurin Inhibitors; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Everolimus; Graft Rejection; Heart Transplantation; Humans; Immunosuppressive Agents; Male; Middle Aged; Sirolimus; Skin Neoplasms

Renal transplant recipients (RTR) have a 50-200-fold higher risk for nonmelanoma-skin cancer (NMSC) causing high rates of morbidity and sometimes mortality. Cohort-studies gave evidence that a sirolimus-based immunosuppression may inhibit skin tumor growth. This single-center, prospective, assessor-blinded, randomized trial investigated if switching to sirolimus treatment inhibits the progression of premalignancies and moreover how many new NMSC occur compared to continuation of the original immunosuppressive therapy. Forty-four RTR (mean age 59.9 years, mean duration of immunosuppression 229.5 months) with skin lesions were randomized to sirolimus or continuation of their original immunosuppression. Blinded dermatological assessment at month 6 and 12 by the same dermatologist evaluated the clinical change compared to baseline. Biopsy was performed in suspected malignancy. Already the 6-month-assessment showed significant superiority of sirolimus-therapy: a stop of progression, even regression of preexisting premalignancies (p < 0.0005). This effect was increased at month 12 (p < 0.0001). Nine patients developed histologically confirmed NMSC: one in the sirolimus group, eight in the control group, p = 0.0176. Sirolimus-based immunosuppression in RTR, even when established many years after transplantation, can delay the development of premalignancies, induce regression of preexisting lesions and decelerate the incidence of new NMSC.

Topics: Aged; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Female; Humans; Immune Tolerance; Immunosuppression Therapy; Immunosuppressive Agents; Kidney Transplantation; Longitudinal Studies; Male; Middle Aged; Precancerous Conditions; Prospective Studies; Sirolimus; Skin Neoplasms

The mammalian target of rapamycin (mTOR) plays central roles in the regulation of cell growth and proliferation by monitoring nutrient availability, cellular energy level, oxygen level, and mitogenic signals. The aberrant activation of mTOR in relation to clinical outcome has been reported in several types of cancers. mTOR is increasingly important as a potential target for anticancer therapy. Nonetheless, a prognostic feature of mTOR activation in esophageal squamous cell carcinoma (ESCC) remains uncertain.. First, in order to validate phospho-specific mTOR antibody (Ser2448), phosphorylated mTOR (p-mTOR) expression levels in five ESCC cell lines under cultural conditions with or without everolimus (mTOR inhibitor, also known as RAD001) were evaluated by in vitro immunohistochemistry and immunoblotting. Second, we examined p-mTOR expression by immunohistochemistry using 143 resected ESCC specimens. Prognostic significance of p-mTOR expression was examined by Cox regression and Kaplan-Meier analyses.. Among 143 patients, 71 (49.7%) were classified into p-mTOR-positive and 72 (50.3%) were classified into p-mTOR-negative. Compared with p-mTOR-negative patients, p-mTOR-positive patients experienced high overall mortality [hazard ratio (HR) 2.44; 95% confidence interval (CI), 1.24-4.83; P = 0.008], which persisted in multivariate analysis (multivariate HR 2.92; 95% CI, 1.48-5.78; P = 0.002). A similar finding was observed for esophageal cancer-specific mortality. p-mTOR expression was not related with any clinical or pathologic variables including age, sex, tumor location, histological grading, operative procedure, T classification (tumor invasion), or lymph-node metastasis.. p-mTOR overexpression was independently associated with poor prognosis in ESCC, supporting the potential for mTOR as a therapeutic target for ESCC.

Topics: Aged; Carcinoma, Squamous Cell; Esophageal Neoplasms; Everolimus; Female; Humans; Immunoblotting; Immunoenzyme Techniques; Immunosuppressive Agents; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Staging; Phosphorylation; Prognosis; Sirolimus; Survival Rate; TOR Serine-Threonine Kinases; Tumor Cells, Cultured

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Humans; Longitudinal Studies; Sirolimus; Skin Neoplasms

Curcumin appears to be a safe, bioactive food compound that is a potential chemopreventive for patients at a high risk for head and neck squamous cell carcinoma (HNSCC). Identification and validation of intermediate endpoints is an important step in evaluating chemopreventive agents. AKT/MTOR pathway biomarkers are intrinsic to the carcinogenic process as well as the mechanism of intervention with curcumin. Antiproliferative effects of curcumin were assayed in 9 HNSCC and a keratinocyte cell line. Nicotine, a genotoxic alkaloid involved in tobacco addiction, forms DNA adducts and has been implicated in upper aerodigestive tract cancer promotion. The antiproliferative effects of curcumin were associated with inhibition of the AKT/MTOR pathway in presence and absence of nicotine, which also induced this pathway. Curcumin was highly effective at suppressing growth of SCC40 xenografts and its activity is associated with modulation of MTOR's downstream target pS6. Curcumin at 15 mg significantly increased survival (286 ± 37 vs. 350 days) in the 4NQO carcinogenic model survival study. A major cause of lethal progression of HNSCC is local regional migration and invasion of malignant cells, and curcumin significantly inhibited cancer cell migration and invasion in vitro and in vivo where downregulation of pS6 was associated with a significant decrease in MMP-9. This is the first study to demonstrate that curcumin inhibits the adverse effects of nicotine by blocking nicotine-induced activation of the AKT/MTOR pathway in HNSCC, which retards cell migration. These studies indicate that inhibiting the AKT/MTOR pathway with curcumin may be useful as an oral chemopreventive agent.

Topics: Animals; Antibiotics, Antineoplastic; Antineoplastic Agents; Apoptosis; Blotting, Western; Carcinoma, Squamous Cell; Cell Adhesion; Cell Line; Cell Movement; Cell Proliferation; Curcumin; Head and Neck Neoplasms; Humans; Immunoenzyme Techniques; Keratinocytes; Mice; NF-kappa B; Nicotine; Phosphatidylinositol 3-Kinases; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

The mammalian target of rapamycin (mTOR) is engaged in the molecular pathogenesis of oral squamous cell carcinoma, which frequently invades the maxilla or the mandible. However, the effects of a mTOR inhibitor on bone destruction associated with oral squamous cell carcinoma are still unclear. In this study, we investigated the antitumor effect of temsirolimus-mediated mTOR inhibition against advanced oral squamous cell carcinoma. Temsirolimus inhibited the proliferation and migration of HSC-2 oral squamous cell carcinoma cells in vitro and suppressed the growth of oral squamous cell carcinoma xenografts in vivo. Significantly, we clearly show that temsirolimus inhibited osteoclast formation both in vitro and in vivo. Reverse transcriptase-PCR analysis showed that temsirolimus decreased the mRNA expression of receptor activator for nuclear factor-κB ligand, known as an osteoclast differentiation factor in bone stromal ST2 cells. Moreover, temsirolimus normalized blood-free calcium concentration in mouse models for humoral hypercalcemia. These findings suggest that mTOR signaling is a potential target of oral squamous cell carcinoma associated with bone destruction, and hence we describe the efficacy of temsirolimus for the treatment of advanced oral squamous carcinoma.

Topics: Animals; Antineoplastic Agents; Bone Neoplasms; Bone Resorption; Carcinoma, Squamous Cell; Cell Movement; Cells, Cultured; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Mouth Neoplasms; Osteoclasts; Sirolimus; Xenograft Model Antitumor Assays

Non-melanoma skin cancers (NMSCs) are the most common malignancies after solid organ transplantation. Their incidence increases with time after transplantation. Calcineurin-inhibitors (CNIs) and azathioprine are known as skin neoplasia-initiating and -enhancing immunosuppressants. In contrast, increasing clinical experience suggests a relevant antiproliferative effect of mammalian target of rapamycin inhibitors, also named proliferation signal inhibitors (PSIs). We report the case of a cardiac allograft recipient with an impressive and consolidated reduction of recurrent NMSC, observed after conversion from CNI-therapy to a PSI-based protocol.

Topics: Calcineurin Inhibitors; Carcinoma, Squamous Cell; Cell Proliferation; Drug Therapy, Combination; Everolimus; Heart Transplantation; Humans; Immunosuppressive Agents; Male; Middle Aged; Neoplasm Recurrence, Local; Photochemotherapy; Signal Transduction; Sirolimus; Skin Neoplasms; Tacrolimus; Time Factors; TOR Serine-Threonine Kinases; Transplantation, Homologous; Treatment Outcome

An estimated 34,000 cases of squamous cell carcinomas of the head and neck (HNSCC) will be diagnosed in 2007 with 7500 estimated deaths. Radiation is commonly used to treat these patients. Preclinical studies have suggested that sirolimus may be an effective radiosensitizer in HNSCC.. The present case report describes a patient, status post liver transplant, who was switched to sirolimus for immunosupression. The patient subsequently underwent radiation therapy for a T2N0M0 SCC of the larynx.. The patient had an unusually early response to radiation, with a clinical complete response after 7 fractions of radiation. However, the patients also had toxicity earlier than expected and required a break from radiation after 11 fractions.. To the authors' knowledge, this is the first observation to suggest that sirolimus is an effective radiosensitizer in patients with HNSCC. We hope that our results will create interest in future clinical studies.

Topics: Carcinoma, Squamous Cell; Dose Fractionation, Radiation; Humans; Immunosuppressive Agents; Laryngeal Neoplasms; Liver Transplantation; Male; Middle Aged; Radiation-Sensitizing Agents; Sirolimus

The enzyme mammalian target of rapamycin (mTOR) integrates many different cellular signals to control cell growth and proliferation, protein synthesis and breakdown, and other processes. Dysregulation of mTOR is implicated in a range of human diseases, including cancers and cardiovascular disorders. To date, there has been no report on the expression of protein kinase B (AKT)/mTOR cell signalling in epidermal tumours.. This study was designed to investigate the activation of the mTOR signalling pathway in epidermal tumours and to correlate this with cyclin-dependent kinase 2 (CDK2) expression.. Immunohistological staining was performed with phosphorylated (p-) AKT, p-mTOR, p-4E-binding protein 1 (p-4EBP1), p-ribosomal protein S6 (p-S6), p-p70 ribosomal protein S6 kinase 1 (p-p70S6K1) and CDK2 in 15 cases each of seborrhoeic keratosis, actinic keratosis, keratoacanthoma and Bowen's disease (BD), and 25 cases of squamous cell carcinoma (SCC). Fifteen normal skin (NS) samples served as control.. Among 85 tumours, 40 (47%) were positive for p-AKT, 31 (36%) for p-mTOR, 44 (52%) for p-4EBP1, 38 (45%) for p-S6, and 39 (46%) for p-p70S6K1. CDK2 immunostaining was positive in all cases of SCC and BD, and in 67% of benign tumours. All of these markers were stained much more frequently in malignant tumours than in benign tumours or NS. p-AKT, p-mTOR, p-4EBP1, p-p70S6K1 and p-S6 each showed high correlation with CDK2.. Constitutive activation of the AKT/mTOR pathway was frequent in epidermal tumours, especially in malignant tumours. Activation was highly correlated with CDK2 expression, suggesting that the AKT/mTOR pathway may induce the malignant transition through CDK2 in epidermal tumours.

Topics: Bowen's Disease; Carcinoma, Squamous Cell; Cyclin-Dependent Kinase 2; Epidermis; Humans; Keratosis, Actinic; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; Skin Neoplasms

Topics: Carcinoma, Squamous Cell; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Everolimus; Humans; Liver Transplantation; Male; Middle Aged; Proteinuria; Sirolimus

The increased molecular understanding of cancerous growth may now afford the opportunity to develop novel therapies targeting specific dysregulated molecular mechanisms contributing to the progression of each cancer type. In this regard, the aberrant activation of Akt/mammalian target of rapamycin (mTOR) pathway is a frequent event in head and neck squamous cell carcinomas (HNSCC), thus representing a potential molecular target for the treatment of HNSCC patients. The ability to translate this emerging body of information into effective therapeutic strategies, however, has been hampered by the limited availability of animal models for oral malignancies. Here, we show that the administration in the drinking water to mice of 4-nitroquinoline-1 oxide, a DNA adduct-forming agent that serves as a surrogate of tobacco exposure, leads to the progressive appearance of preneoplastic and tumoral lesions in the tongue and oral mucosa, with 100% incidence after only 16 weeks of carcinogen exposure. Remarkably, many of these lesions evolve spontaneously into highly malignant SCCs few weeks after 4-nitroquinoline-1 oxide withdrawal. In this model, we have observed that the activation of the Akt-mTOR biochemical route represents an early event, which is already detectable in dysplastic lesions. Furthermore, we show that the inhibition of mTOR by the chronic administration of rapamycin halts the malignant conversion of precancerous lesions and promotes the regression of advanced carcinogen-induced SCCs. Together, these findings support the contribution of the mTOR signaling pathway to HNSCC progression and provide a strong rationale for the early evaluation of mTOR inhibitors as a molecular-targeted strategy for HNSCC chemoprevention and treatment.

Topics: 4-Nitroquinoline-1-oxide; Animals; Antibiotics, Antineoplastic; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Disease Progression; Female; Gene Expression; Head and Neck Neoplasms; Immunohistochemistry; Mice; Mice, Inbred C57BL; Precancerous Conditions; Protein Kinases; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

Head and neck squamous cell carcinomas (HNSCC), the majority of which occur in the oral cavity, remain a significant cause of morbidity and mortality worldwide. A major limitation in HNSCC research has been the paucity of animal models to test the validity of current genetic paradigms of tumorigenesis and to explore the effectiveness of new treatment modalities and chemopreventive strategies. Here, we have developed an inducible oral-specific animal tumor model system, which consists in the expression of a tamoxifen-inducible Cre recombinase (CreER(tam)) under the control of the cytokeratin 14 (K14) promoter (K14-CreER(tam)) and mice in which the endogenous K-ras locus is targeted (LSL-K-ras(G12D)), thereby causing the expression of endogenous levels of oncogenic K-ras(G12D) following removal of a stop element. Surprisingly, whereas K14-CreER(tam) can also target the skin, K14-CreER(tam)/LSL-K-ras(G12D) mice developed papillomas exclusively in the oral mucosa within 1 month after tamoxifen treatment. These lesions were highly proliferative but never progressed to carcinoma. However, when crossed with p53 conditional knockout (p53(flox/flox)) mice, mice developed SCCs exclusively on the tongue as early as 2 weeks after tamoxifen induction, concomitant with a remarkable activation of the mammalian target of rapamycin (mTOR) signaling pathway. The availability of this ras and p53 two-hit animal model system recapitulating HNSCC progression may provide a suitable platform for exploring novel molecular targeted approaches for the treatment of this devastating disease. Indeed, we show here that mTOR inhibition by the use of rapamycin is sufficient to halt tumor progression in this genetically defined oral cancer model system, thereby prolonging animal survival.

Topics: Animals; Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Cell Division; Disease Models, Animal; Genes, p53; Genes, ras; Head and Neck Neoplasms; Humans; Integrases; Mice; Mouth Mucosa; Mouth Neoplasms; Papilloma; Sirolimus; Tamoxifen

Immunosuppression favors the development of skin cancer. Experimental data suggest that sirolimus (SRL) has antitumoral and antiangiogenic properties. An investigation was undertaken into the effects of SRL on squamous cell carcinoma (SCC) developing in organ transplant recipients (OTR) receiving immunosuppressive treatments, with special emphasis on vascularization.. SCC that developed in eight OTR before and after conversion from calcineurin inhibitors (CNI) to SRL were compared for thickness, differentiation, ulceration, perineural invasion, density of peritumoral infiltrate, peritumoral vascularization, density of T-regulatory cells and of intratumoral Langerhans cells and growth fraction.. SCC developing under SRL showed lower peritumoral vascularization and thickness, and higher growth fraction and density of peritumoral T-regulatory cells.. Conversion from CNI to SRL at clinically relevant doses is associated in vivo with a reduced vascularization and thickness of post-transplant human cutaneous SCC. This effect could account for the beneficial effect of SRL on immunosuppression-induced skin carcinogenesis in humans.

Topics: Adult; Aged; Angiogenesis Inhibitors; Calcineurin Inhibitors; Carcinoma, Squamous Cell; Cohort Studies; Cyclosporine; Female; Graft Rejection; Heart Transplantation; Humans; Immunoenzyme Techniques; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Neovascularization, Pathologic; Postoperative Complications; Sirolimus; Skin Neoplasms; Treatment Outcome

To determine if the mammalian target of rapamycin (mTOR) inhibitor CCI-779 can sensitize head and neck squamous cell carcinoma (HNSCC) to radiotherapy (XRT) and compare the radiosensitizing effects to cisplatin with its known considerable toxicity. Radiosensitizing effects of CCI-779 were assayed on HNSCC cell lines in vitro. CCI-779 (5 mg/kg), cisplatin (1 mg/kg), and XRT (2 Gy) alone and in combination were evaluated for antitumor activity in mice bearing FaDu and SCC40 xenografts. Effects of CCI-779 on radiation-induced activation of the Akt/mTOR pathway were analyzed. Although CCI-779 did not sensitize HNSCC cells to ionizing radiation in vitro, combination of CCI-779 and XRT significantly augmented the in vivo tumor growth-inhibitory effects of XRT and CCI-779 (P < 0.05). In addition, CCI-779 + XRT suppressed tumor growth more effectively than cisplatin + XRT (P < 0.05). CCI-779 + XRT significantly improved survival compared with XRT alone in both cisplatin-sensitive FaDu (P < 0.01) and cisplatin-resistant SCC40 (P < 0.05) xenograft mice. There were no additional benefits of adding cisplatin to CCI-779 + XRT. CCI-779 significantly attenuated irradiation-induced up-regulation of the mTOR pathway, increased apoptosis and displayed potent antiangiogenic activity in FaDu xenografts that was further enhanced by its combination with XRT (P < 0.05), which may explain the mechanism of its selective radiosensitizing effects in vivo and not in vitro. Antitumor activity of XRT was enhanced when combined with CCI-779 in HNSCC xenograft model. CCI-779 + XRT showed antitumor activity superior to conventional chemoradiotherapy with cisplatin. These results pave the way for clinical trials using molecular targeted therapy with CCI-779 in combination with XRT for HNSCC treatment.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cisplatin; Combined Modality Therapy; Head and Neck Neoplasms; Mice; Mice, Inbred Strains; Protein Kinases; Radiation-Sensitizing Agents; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

The cytotoxic effects and mechanism of action of cisplatin and the mTOR inhibitor rapamycin on Hep-2 laryngeal cancer cells were investigated. Hep-2 cells were cultured in the presence of different concentrations of rapamycin, cisplatin, or the two combined. Inhibition of cell growth, apoptosis, and AKT, mTOR, S6K, and ERCC1 protein levels were assessed. All combinations of rapamycin and cisplatin resulted in synergistic inhibition of cell growth (as indicated by q values determined using Jin's formula > 1.15). Rapamycin inhibited Hep-2 cell growth, induced G1 arrest, and when combined with cisplatin, enhanced apoptosis. p-mTOR and S6K expressions were significantly downregulated by rapamycin. ERCC1 expression was significantly upregulated with cisplatin treatment. Combined cisplatin and rapamycin treatment resulted in significant downregulated p-mTOR and S6K expression, but no change in ERCC1 expression. Rapamycin and cisplatin act in a synergistic manner, increasing the cytotoxic effect on Hep-2 cells. Rapamycin may facilitate increased Hep-2 cell apoptosis with cisplatin via inhibiting downstream expression of proteins in the AKT-mTOR signaling pathway.

Topics: Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cisplatin; DNA-Binding Proteins; Drug Synergism; Endonucleases; Humans; Laryngeal Neoplasms; Protein Kinases; Ribosomal Protein S6 Kinases; Sirolimus; TOR Serine-Threonine Kinases

Traditional immunosuppressive regimens make laryngeal transplantation in cancer patients prohibitive because of the increased risk of recurrence. Everolimus, a recently developed immunosuppressant, has demonstrated significant antitumor properties. The purpose of this study was to examine the effects of everolimus alone and in combination with other immunosuppressants on tumor growth in a combined laryngeal transplantation and tumor model.. Animal, prospective, randomized, controlled, and blinded.. One million squamous cell carcinoma cells (SCC-158) were injected intravenously into a total of 40 rats 1 day before laryngeal transplantation. Rats were divided into four groups differing by immunosuppressive regimens. Lung surface metastases were counted 21 days after inoculation, and numerical transplantation rejection scores were recorded. A separate experiment for comparison was performed with no transplant on 24 rats, but with the same immunosuppressive treatment groups.. The median number of lung surface metastases were: a) control (i.e., no immunosuppression): 85; b) everolimus 1.0 mg/kg: 25; c) tacrolimus 1.2 mg/kg: 1650; d) everolimus 1.0 mg/kg + tacrolimus 0.05 mg/kg: 1300. Rats receiving everolimus alone showed a statistically significant decrease in pulmonary surface metastases compared with the other groups. Transplanted rats had no difference in their outcomes when compared with non-transplanted rats.. Everolimus significantly decreases SCC-158 growth in our combined transplantation and tumor model compared with controls and other immunosuppressants.

Topics: Administration, Oral; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Dose-Response Relationship, Drug; Drug Therapy, Combination; Everolimus; Graft Survival; Immunosuppressive Agents; Laryngeal Neoplasms; Laryngectomy; Larynx; Lung Neoplasms; Male; Neoplasm Recurrence, Local; Neoplasm Transplantation; Rats; Rats, Inbred F344; Sirolimus; Tacrolimus

To evaluate the effect of combined use of rapamycin and cisplatin in the chemotherapy of Hep-2 cells in vitro.. The inhibitory effects of rapamycin and cisplatin, used alone or in combination, on the proliferation of Hep-2 cells were measured with MTT assay and median-effect plot analysis. The cell cycle changes after the treatment were analyzed using flow cytometry and Hoechst 33258 immunofluorescence staining.. The IC50 of rapamycin and cisplatin for inducing growth arrest of Hep-2 cells was 11.03 nmol/L and 8.81 micromol/L, respectively. Rapamycin alone caused cell cycle arrest of the Hep-2 cells in G1 phase. Rapamycin and cisplatin showed synergistic effects in the chemotherapy of Hep-2 cells (q > 1.15, King's Formula), causing significantly increased apoptosis ratio and growth inhibition rate of Hep-2 cells.. Combined use of rapamycin and cisplatin significantly improves the chemotherapeutic effect against Hep-2 cells.

Topics: Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Proliferation; Cisplatin; Drug Synergism; Humans; Laryngeal Neoplasms; Sirolimus; Tumor Cells, Cultured

Emerging evidence supporting the activation of the Akt-mammalian target of rapamycin (mTOR) signaling network in head and neck squamous cell carcinoma (HNSCC) progression has provided the rationale for exploring the therapeutic potential of inhibiting this pathway for HNSCC treatment. Indeed, rapamycin, a clinically relevant mTOR inhibitor, promotes the rapid regression of HNSCC-tumor xenografts in mice. However, rapamycin does not affect the growth of HNSCC cells in vitro, thus raising the possibility that, as for other cancer types, rapamycin may not target cancer cells directly but may instead act on a component of the tumor microenvironment, such as tumor-associated vasculature. Here, we used a retroinhibition approach to assess the contribution of cancer cell-autonomous actions of rapamycin to its antitumor activity in HNSCC. A rapamycin-resistant form of mTOR (mTOR-RR) was expressed in HNSCC cells while retaining the wild-type (rapamycin-sensitive) mTOR (mTOR-WT) alleles in host-derived endothelial and stromal cells. Expression of mTOR-RR prevented the decrease in phospho-S6 levels caused by rapamycin through mTOR in HNSCC cells but not in stromal cells, and rendered HNSCC xenografts completely resistant to the antitumoral activity of rapamycin. This reverse pharmacology strategy also enabled monitoring the direct consequences of inhibiting mTOR in cancer cells within the complex tumor microenvironment, which revealed that mTOR controls the accumulation of hypoxia-inducible factor-1 alpha (HIF-1 alpha) and the consequent expression of vascular endothelial growth factor and a glucose transporter, Glut-1, in HNSCC cells. These findings indicate that HNSCC cells are the primary target of rapamycin in vivo, and provide evidence that its antiangiogenic effects may represent a downstream consequence of mTOR inhibition in HNSCC cells.

Topics: Animals; Antibiotics, Antineoplastic; Apoptosis; Carcinoma, Squamous Cell; Cell Growth Processes; Cell Line, Tumor; Endothelial Cells; Female; Head and Neck Neoplasms; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Mice; Mice, Nude; Mutation; Protein Kinases; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Topics: Adult; Carcinoma; Carcinoma, Squamous Cell; Fatal Outcome; Graft Rejection; Humans; Immunosuppressive Agents; Kidney Transplantation; Lymphangitis; Male; Neoplasm Metastasis; Reoperation; Sirolimus; Skin Neoplasms

The mammalian target of rapamycin (mTOR), an important regulator of protein translation and cell proliferation, is activated in various malignancies. In a randomised controlled trial of advanced renal cell carcinoma patients, targeted therapy to mTOR by means of rapamycin analogues has been shown to significantly improve survival. An in vitro study has revealed that mTOR is activated in oesophageal squamous cell carcinoma (OSCC) cell lines and that mTOR expression is inhibited by rapamycin. The objectives of this histological study were to determine the proportion of OSCC tissues with activated mTOR (p-mTOR) expression, thereby assessing the percentage of patients with OSCC that would possibly benefit from neoadjuvant rapamycin therapy, and to identify the clinicopathological features of these potentially rapamycin-sensitive tumours.. The expression of p-mTOR (Ser2448) was immunohistochemically assessed in a validated tissue microarray comprising triplicate tissue biopsy cores of 108 formalin-fixed, paraffin-embedded OSCCs. Staining results were correlated with clinicopathological data.. Normal oesophageal epithelium was negative for p-mTOR. Activated mTOR expression was located in the cytoplasm of oesophageal tumour cells. 26 (25%) of 105 assessable OSCCs showed tumour cells with positive staining for activated mTOR. Activated mTOR expression was associated with a lesser degree of differentiation only (p = 0.024). No correlation was detected between p-mTOR and the proliferation marker Ki-67.. Activated mTOR can be detected in one-quarter of OSCCs. Since this subset of patients may potentially benefit from mTOR inhibiting therapy, a phase II clinical trial of neoadjuvant mTOR-inhibiting therapy in patients with OSCC may be considered.

Topics: Adult; Aged; Antibiotics, Antineoplastic; Biomarkers, Tumor; Carcinoma, Squamous Cell; Esophageal Neoplasms; Female; Humans; Immunoenzyme Techniques; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Proteins; Neoplasm Staging; Protein Kinases; Sirolimus; TOR Serine-Threonine Kinases

Chronic inflammation and cancer development are associated with dysregulated immune responses and the presence of regulatory T cells (T(reg)). To study the role of T(reg) in tumor cell escape from immune surveillance, an in vitro model simulating the tumor microenvironment and promoting the induction and expansion of IL-10(+) T(reg )type 1 (Tr1) was established.. An in vitro co-culture system (IVA) included an irradiated head and neck squamous cell carcinoma cell line, immature dendritic cells (iDC), CD4(+)CD25(- )T cells and cytokines, IL-2 (10 IU/ml), IL-10 (20 IU/ml), IL-15 (20 IU/ml) +/- 1 nM rapamycin. Autologous iDC and CD4(+)CD25(-) T cells were obtained from the peripheral blood of 15 normal donors. Co-cultures were expanded for 10 days. Proliferating lymphocytes were phenotyped by multi-color flow cytometry. Their suppressor function was measured in CFSE inhibition assays +/- neutralizing anti-IL-10 mAb and using transwell cultures. Culture supernatants were tested for IL-4, IL-10, TGF-beta and IFN-gamma in ELISA.. In the IVA, low doses of IL-2, IL-10 and IL-15 promoted induction and expansion of CD3(+)CD4(+)CD25(-)IL2Rbeta(+)IL2Rgamma(+)FoxP3(+)CTLA-4(+)IL-10(+) cells with suppressor activity (mean suppression +/- SD = 58 +/- 12%). These suppressor cells produced IL-10 (mean +/- SD = 535 +/- 12 pg/ml) and TGF-beta (mean +/- SD = 512 +/- 38 pg/ml), but no IL-4 or IFN-gamma. Suppressor function of co-cultures correlated with the percent of expanding IL-10(+) Tr1 cells (r (2 )=( )0.9; P < 0.001). The addition of rapamycin enriched Tr1 cells in all co-cultures. Neutralizing anti-IL-10 mAb abolished suppressive activity. Suppression was cell-contact independent.. The tumor microenvironment promotes generation of Tr1 cells which have the phenotype distinct from that of CD4(+)CD25(high)FoxP3(+) nTreg and mediate IL-10 dependent immune suppression in a cell-contact independent manner. Tr1 cells may play a critical role in cancer progression.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Coculture Techniques; Cytokines; Dendritic Cells; Enzyme-Linked Immunosorbent Assay; Head and Neck Neoplasms; Humans; Interleukin-10; Lymphocyte Activation; Sirolimus; T-Lymphocytes, Regulatory

Molecular therapeutics identifies an aberration in tumors to select patients that benefit from molecular targeted therapy. Overexpression of eIF4E in histologically "tumor-free" surgical margins of head and neck squamous cell cancer (HNSCC) patients is an independent predictor of recurrence and is functionally activated through the Akt/mammalian target of rapamycin (mTOR) pathway. Although mTOR inhibitors are cytostatic agents, best used in combination therapy, we hypothesize that they can be used as long-term single agents in an HNSCC model of minimal residual disease (MRD). CCI-779, an mTOR inhibitor, arrested growth of a phosphatase and tensin homologue deleted on chromosome 10 (PTEN) abnormal HNSCC cell line FaDu, inhibiting phosphorylation of 4E-binding protein 1, resulting in increased association with eIF4E and inhibition of basic fibroblast growth factor and vascular endothelial growth factor. Fluorescence in situ hybridization detected PTEN abnormalities in 68% of patient tumors and 35% of tumor-free margins. CCI-779 inhibited growth of established tumors in nude mice. However, in the MRD model, there were significant differences in the tumor-free rate between the control (4%) and the treatment group (50%), and the median tumor-free time was 7 versus 18 days, respectively (P < 0.0001). In those animals that formed tumors, CCI-779 caused a significant decrease in the tumor volume. The Kaplan-Meier curve showed that CCI-779 significantly increased survival (P < 0.0001). The mTOR pathway was inhibited in peripheral blood mononuclear cells potential surrogate markers of response to therapy. Stable transfection of FaDu with luciferase allowed us to monitor the effects of CCI-779 with bioluminescence imaging in the MRD model. These results pave the way for a clinical trial using targeted molecular therapy with CCI-779 as a single agent for mTOR-activated residual cells.

Topics: Animals; Antineoplastic Agents; Carcinoma, Squamous Cell; Chemotherapy, Adjuvant; Drug Evaluation, Preclinical; Gene Dosage; Head and Neck Neoplasms; Humans; Mice; Neoplasm, Residual; Protein Kinase Inhibitors; Protein Kinases; PTEN Phosphohydrolase; Sirolimus; TOR Serine-Threonine Kinases; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

The epidermal growth factor receptor (EGFR) is a validated target in squamous cell carcinoma (SCC) of the head and neck. Most patients, however, do not respond or develop resistance to this agent. Mammalian target of rapamycin (mTOR) is involved in the pathogenesis of SCC of the head and neck (SCCHN). This study aimed to determine if targeting mTOR in combination with EGFR is effective in SCC, and to develop early pharmacodynamic markers of efficacy. Two SCC cell lines, one resistant (HEP2) and one of intermediate susceptibility (Detroit 562) to EGFR inhibitors, were xenografted in vivo and treated with an mTOR inhibitor (temsirolimus), an EGFR inhibitor (erlotinib) or a combination of both. Temsirolimus exerted superior growth arrest in both cell lines than erlotinib. The combined treatment resulted in synergistic antitumor effects in the Detroit 562 cell line. Immunohistochemical assessment of pharmacodynamic effects in fine-needle aspiration (FNA) biopsies early after treatment using phospho MAPK, Phospho-P70 and Ki67 as end points demonstrated pathway abrogation in the Detroit 562 tumours treated with the combination, the only group where regressions were seen. In conclusion, an mTOR inhibitor showed antitumor activity in EGFR-resistant SCC cell lines. Marked antitumor effects were associated with dual pathway inhibition, which were detected by early FNA biopsies.

Topics: Angiogenesis Inhibitors; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Drug Synergism; ErbB Receptors; Erlotinib Hydrochloride; Female; Gene Expression; Gene Expression Profiling; Growth Inhibitors; Head and Neck Neoplasms; Humans; Immunohistochemistry; Mice; Mice, Nude; Neovascularization, Pathologic; Protein Kinase Inhibitors; Protein Kinases; Quinazolines; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

mTOR/p70S6K pathway is considered a central regulator in various malignant tumors, but its roles in esophageal squamous cell carcinoma (ESCC), which is a common cause of mortality in China, remain unknown. Here, we identify that the mTOR/p70S6K pathway is activated in ESCC; rapamycin and siRNA against mTOR rapidly inhibited expression of mTOR and the phosphorylation of its major downstream effectors, p70S6K and 4E-BP1, arrested cells in the G(0)/G(1) phase and induced apoptosis of ESCC cells. The findings may lay a foundation for making further investigations on the mTOR/p70S6K pathway as a potential target for ESCC therapy.

Topics: Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Esophageal Neoplasms; Humans; Protein Kinase Inhibitors; Protein Kinases; Ribosomal Protein S6 Kinases, 70-kDa; RNA, Small Interfering; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Transfection; Up-Regulation

Circulating human CD4(+)CD25(high)Foxp3(+) T cell populations (Treg) may contain activated CD4(+)CD25(+) T cells interfering with Treg evaluation. To gain insights into the phenotypic and functional characteristics of Treg in patients with cancer, we have analyzed CD4(+)CD25(high) populations at the clonal level. Single-cell sorted (SCS) CD4(+)CD25(high) T cells obtained from PBMC of normal controls (NC) or patients with squamous cell carcinoma of the head and neck (HNSCC) were plated at 1 cell/well in 96 well plates and expanded with anti-CD3/anti-CD28 Abs and 1,000 IU IL-2/mL in the presence or absence of rapamycin (1 nM). All generated clones were evaluated for the phenotype by flow cyometry and suppressor function in CFSE-based proliferation assays. Clones had heterogeneous CD25 expression levels. Cloning efficiency of CD4(+)CD25(high) T cells was low. CD25(high) clones expressed CTLA-4, Foxp3, CD62L, but little GITR and suppressed proliferation of autologous CD4(+)CD25(-) responder cells. Clones of activated CD4(+)CD25(interm./low) cells expressed intermediate to high levels of GITR and HLA-DR and did not suppress proliferation of responder cells. The number, suppressor phenotype and function of CD25(high) Treg clones were significantly enhanced in HNSCC patients relative to NC (p

Topics: Adult; Aged; Antibiotics, Antineoplastic; Carcinoma, Squamous Cell; Case-Control Studies; CD4-Positive T-Lymphocytes; Cell Proliferation; Clone Cells; Female; Flow Cytometry; Forkhead Transcription Factors; Head and Neck Neoplasms; Humans; Interleukin-2 Receptor alpha Subunit; L-Selectin; Male; Middle Aged; Phenotype; Sirolimus; T-Lymphocytes, Regulatory

Cyclooxygenase 2 (COX-2) overexpression and production of prostaglandin E(2) (PGE(2)) by head and neck squamous cell carcinomas (HNSCC) induce type 1 regulatory T (Tr1) cells and contribute to carcinogenesis by creating a tolerogenic milieu. To test this hypothesis, CD4(+)CD25(-) T cells obtained from the peripheral blood of 10 normal donors were cocultured with autologous dendritic cells, irradiated HNSCC cells and cytokines, interleukin 2 (IL-2), IL-10, and IL-15. HNSCC cells were either COX-2 negative, constitutively expressed COX-2, were transfected with COX-2, or had COX-2 expression knocked down by small interfering RNA. Other modifications included coculture plus or minus the COX-inhibitor, Diclofenac, or synthetic PGE(2) in the absence of HNSCC. Lymphocytes proliferating in 10-day cocultures were phenotyped by flow cytometry, studied for cytokine production by ELISA and for suppressor function in CFSE inhibition assays plus or minus anti-IL-10 or anti-transforming growth factor-beta(1) (TGF-beta(1)) monoclonal antibodies (mAb). COX-2(+) HNSCC or exogenous PGE(2) induced outgrowth of Tr1 cells with the CD3(+)CD4(+)CD25(-)IL2Rbeta(+)IL2Rgamma(+)FoxP3(+)CTLA-4(+)IL-10(+)TGF-beta(1)(+)IL-4(-) phenotype and high suppressor functions (range, 46-68%). Small interfering RNA knockout of COX-2 gene in HNSCC led to outgrowth of lymphocytes with decreased IL2Rgamma (P = 0.0001), FoxP3 (P = 0.05), and IL-10 (P = 0.035) expression and low suppressor activity (range, 26-34%). Whereas COX-2(+) cocultures contained IL-10 and TGF-beta(1) (medians, 615 and 824 pg/mL), cytokine levels were decreased (P < 0.0001) in COX-2(-) cocultures. Inhibition of COX-2 enzymatic activity in HNSCC abrogated outgrowth of Tr1 cells. Neutralizing mAbs to IL-10 and/or TGF-beta(1) abolished Tr1-mediated suppression. COX-2 overexpression in HNSCC plays a major role in the induction of Tr1 cells in the tumor microenvironment.

Topics: Carcinoma, Squamous Cell; CD4-Positive T-Lymphocytes; Cell Line, Tumor; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Cytokines; Head and Neck Neoplasms; Humans; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Lymphocyte Activation; RNA, Small Interfering; Sirolimus; T-Lymphocytes, Regulatory; Transfection; Transforming Growth Factor beta1

Immunosuppressive therapies allow long-term patient and transplant survival, but are associated with increased development of UV-induced skin cancers, particularly squamous cell carcinomas. The mechanisms by which CsA, MMF, tacrolimus (TAC) or sirolimus (SRL), alone or in dual combinations, influence tumor development and progression are not completely understood. In the current study, chronically UV-exposed mice treated with SRL alone or in combination with CsA or TAC developed more tumors than mice treated with vehicle or other immunosuppressants, but the tumors were significantly smaller and less advanced. Mice treated with CsA or TAC developed significantly larger tumors than vehicle-treated mice, and a larger percentage in the CsA group were malignant. The addition of MMF to CsA, but not to TAC, significantly reduced tumor size. Immunosuppressant effects on UVB-induced inflammation and tumor angiogenesis may explain these findings. CsA enhanced both UVB-induced inflammation and tumor blood vessel density, while MMF reduced inflammation. Addition of MMF to CsA reduced tumor size and vascularity. SRL did not affect inflammation, but significantly reduced tumor vascularity. Thus the choice of immunosuppressants has important implications for tumor number, size and progression, likely due to the influence of immunosuppressants on UVB-induced inflammation and angiogenesis.

Topics: Animals; Blood Vessels; Carcinoma, Squamous Cell; Cyclosporine; Disease Models, Animal; Drug Therapy, Combination; Female; Immunosuppressive Agents; Inflammation; Mice; Mice, Hairless; Mycophenolic Acid; Neoplasms, Radiation-Induced; Neovascularization, Pathologic; Sirolimus; Skin Neoplasms; Tacrolimus; Ultraviolet Rays

Polymorphisms or mutations in hypoxia inducible factor-1 alpha (HIF-1alpha) that increases its activity and stability under normoxia have recently been identified. Likewise, disruption of the TSC1/TSC2 complex through loss of TSC1 or TSC2 has been shown to result in abnormal accumulation of HIF-1alpha. Here, we investigate the novel polymorphisms in exon 12, that approximate the oxygen-dependent degradation domain of HIF-1alpha in five cell lines and 28 patients with oral squamous carcinomas. Moreover, we assess for the presence of polymorphisms and mutations in TSC1 and TSC2, to ascertain if dysregulation of such might complement HIF-1alpha expression.. Denaturing high pressure liquid chromatography (DHPLC) analysis on PCR fragments in exon 12 of HIF-1alpha from 28 patients with OSCC revealed that 6 of 28 patients had mismatched heteroduplex patterns. Genomic DNA was extracted from peripheral blood leukocytes and direct sequencing showed that in 5 of the six cases these changes represented polymorphisms while, one case was a somatic mutation. Analyses of TSC1 and TSC2 revealed heteroduplexes in exons: TSC1 exon 17; TSC2 exons 36, 40, and 41. The relative levels of HIF-1alpha were significantly greater for tumors possessing a HIF-1alpha polymorphism or mutation within exon 12, whereas tumors possessing a deletion or polymorphism in TSC1/TSC2 displayed a trend for higher levels of HIF-1alpha. Western blot analyses for HIF-1alpha, TSC1 and TSC2 in five SCC cell lines revealed high levels of HIF-1alpha in SCC cells possessing TSC1 and/or TSC2 mutations. Wild-type TSC2 cells targeted with siRNA to TSC2 exhibited increased levels of HIF-1alpha. Transfection of a HIF-1alpha mutant produced higher levels of HIF-1alpha in TSC1/TSC2 mutant cell lines than in wild type cells. TSC1/TSC2 mutant cell lines administered Rapamycin blocked S6 phorphorylation and diminished the levels of HIF-1alpha to those observed in cell lines with wild type TSC1/TSC2.. Dysregulation of the TSC1/TSC2 complex by mutation compliments HIF-1alpha polymorphisms in the expression of HIF-1alpha in SCC of the head and neck, and may provide biomarkers to predict responses to specific therapies and overall disease prognosis.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Female; Head and Neck Neoplasms; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Mouth Neoplasms; Mutation; Polymorphism, Genetic; Protein Kinase Inhibitors; Protein Kinases; RNA Interference; Sirolimus; TOR Serine-Threonine Kinases; Transfection; Tuberous Sclerosis Complex 1 Protein; Tuberous Sclerosis Complex 2 Protein; Tumor Suppressor Proteins

Everolimus (RAD) is an mTOR inhibitor closely related to rapamycin. A potent immunosuppressive agent, it has also shown evidence of antineoplastic properties. SCC VII is a spontaneously arising murine squamous cell carcinoma line. This study examines the effect of everolimus on SCC VII proliferation. The data may provide support for the use of everolimus in transplant recipients with a history of malignancy.. A dose efficacy study was conducted that used a murine model of intradermal tumor growth and pulmonary metastases. The development of intradermal tumors and pulmonary metastases were studied. Of 80 total mice, 40 received intradermal injection of 1 x 10 SCC VII cells and 40 received intravenous injection of 1 x 10 cells to establish pulmonary metastases. Within each group, animals were subdivided into four subgroups that received 1) 1 mg/kg everolimus twice a day, 2) 0.5 mg/kg everolimus twice a day, 3) 7.5 mg/kg cyclosporine per day, and 4) no treatment. Intradermal tumors were measured three times per week. Animals receiving an intravenous tumor injection were killed after 17 days and pulmonary metastases were quantified. Medication trough levels were measured in all treated animals.. Everolimus showed statistically significant tumor inhibition at 1.0 mg/kg twice a day and 0.5 mg/kg twice a day when compared with animals treated with cyclosporine and with untreated animals (P < .0001). Tumor inhibition was evident in both models studied (intradermal tumors and pulmonary metastasis generation).. Everolimus provides potent tumor inhibition in animals inoculated with SCC VII cells. Inhibition of both local and distant spread of disease is evident. Although most immunosuppressives are known to potentiate neoplastic disease, this study supports the use of everolimus immunosuppression in the face of prior malignancy. This data has significant implication for laryngeal transplantation after laryngectomy.

Topics: Animals; Carcinoma, Squamous Cell; Cell Proliferation; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Everolimus; Female; Immunosuppressive Agents; Injections, Intravenous; Lung Neoplasms; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Probability; Random Allocation; Reference Values; Sensitivity and Specificity; Sirolimus; Skin Neoplasms; Tumor Cells, Cultured

Emerging knowledge on how the dysregulated function of signaling networks contributes to the malignant growth of squamous cell carcinoma of the head and neck (HNSCC) can now be exploited to identify novel mechanism-based anticancer treatments. In this regard, we have observed that persistent activation of the serine/threonine kinase Akt is a frequent event in HNSCC, and that blockade of its upstream kinase, 3'-phosphoinositide-dependent kinase 1, potently inhibits tumor cell growth. Akt promotes cell proliferation by its ability to coordinate mitogenic signaling with energy- and nutrient-sensing pathways that control protein synthesis through the atypical serine/threonine kinase, mammalian target of rapamycin (mTOR). This kinase, in turn, phosphorylates key eukaryotic translation regulators, including p70-S6 kinase and the eukaryotic translation initiation factor, 4E binding protein 1. Indeed, we show here that aberrant accumulation of the phosphorylated active form of S6, the most downstream target of the Akt-mTOR-p70-S6 kinase pathway, is a frequent event in clinical specimens from patients with HNSCC and their derived cell lines. Of interest, this enhanced level of the phosphorylated active form of S6 was rapidly reduced in HNSCC cell lines and HNSCC xenograft models at clinically relevant doses of rapamycin, which specifically inhibits mTOR. Furthermore, we observed that rapamycin displays a potent antitumor effect in vivo, as it inhibits DNA synthesis and induces the apoptotic death of HNSCC cells, ultimately resulting in tumor regression. These findings identify the Akt-mTOR pathway as a potential therapeutic target for HNSCC, and may provide the rationale for the early clinical evaluation of rapamycin and its analogues in patients with HNSCC.

Topics: Animals; Antibiotics, Antineoplastic; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; DNA, Neoplasm; Female; Head and Neck Neoplasms; Humans; Mice; Mice, Nude; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Kinases; Proto-Oncogene Proteins c-akt; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Discovery of the common and ubiquitous molecular targets for the disruption of angiogenesis, that are independent of the characteristics of malignant tumors, is desired to develop the more effective antitumor drugs. In this study, we propose that the platelet-derived growth factor receptor-alpha (PDGFRalpha)-p70S6K signal transduction pathway in mesenchymal cells, which is required for functional angiogenesis induced by fibroblast growth factor-2, is the potent candidate. Using murine limb ischemia as a tumor-free assay system, we demonstrated that p70S6K inhibitor rapamycin (RAPA) targets mesenchymal cells to shut down the sustained expression of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF), via silencing of the PDGFRalpha-p70S6K pathway. Irrespective of the varied expression profiles of angiogenic factors in each tumor tested, RAPA constantly led the tumors to dormancy and severe ischemia in the time course, even associated with upregulated expression of VEGF from tumors. Because RAPA showed only a minimal effect to hypoxia-related expression of VEGF in culture, these results suggest that RAPA targets the host-vasculature rather than tumor itself in vivo. Thus, our current study indicates that the PDGFRalpha-p70S6K pathway is an essential regulator for FGF-2-mediated therapeutic neovascularization, as well as for the host-derived vasculature but not tumors during tumor angiogenesis, via controlling continuity of expression of multiple angiogenic growth factors.

Topics: Animals; Carcinoma, Squamous Cell; Cell Hypoxia; Epidermal Growth Factor; Fibroblast Growth Factor 2; Gene Expression Regulation; Hepatocyte Growth Factor; Hindlimb; Humans; Ischemia; Liver Neoplasms, Experimental; Male; Mesoderm; Mice; Mice, Inbred C57BL; Mice, Nude; Mouth Neoplasms; Neoplasm Proteins; Neovascularization, Pathologic; Neovascularization, Physiologic; Platelet-Derived Growth Factor; Receptor, Platelet-Derived Growth Factor alpha; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Sirolimus; Stromal Cells

Interleukin (IL-2) and IL-2Rbeta/gamma have been shown to be expressed in human carcinomas in culture and in situ. Recently, expression of endogenous IL-2 and IL-2R in the cytoplasm was found to be up-regulated in tumour cells undergoing mitosis. This observation suggested that similar to its role in lymphocytes, the IL-2/IL-R pathway is involved in the regulation of carcinoma cell proliferation. Metabolic labelling followed by immunoprecipitation and Western blot results showed that IL-2 in carcinomas was identical to that in human lymphocytes. However, tumour cells did not secrete IL-2 detectable by immunoassays, although membrane-associated IL-2 was detectable on a proportion of these cells cultured in the absence of exogenous IL-2. Antibodies to IL-2 failed to inhibit proliferation of carcinoma cells, but antibodies specific for the ligand-binding site of the IL-2R were growth inhibitory. Growth of tumour cells was also inhibited by the immunosuppressive drugs, cyclosporin A (CsA), FK506 and rapamycin (RPA), known to interfere with the IL-2 pathway in lymphocytes. To further confirm the role of endogenous IL-2 in the growth of carcinomas, tumour cells were incubated with an IL-2-specific antisense oligonucleotide. The treatment was shown to transiently inhibit IL-2 mRNA and IL-2 protein expression as well as proliferation of tumour cells. Tumour cells treated with IL-2-specific antisense oligonucleotide demonstrated increased apoptosis in comparison to untreated or sense oligonucleotide-treated control cells. The data indicate that in human carcinomas, endogenous IL-2 promotes growth and protects tumour cells from apoptosis.

Topics: Antibodies; Apoptosis; Carcinoma, Squamous Cell; Cell Division; Head and Neck Neoplasms; Humans; Interleukin-2; Jurkat Cells; Oligonucleotides, Antisense; Receptors, Interleukin-2; Sirolimus; Stomach Neoplasms; Tacrolimus; Tumor Cells, Cultured