sirolimus and Papillomavirus-Infections

Activation of the phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is common in head and neck cancers, and it has been demonstrated that inhibition of mTOR complex 1 sensitizes cell lines to platinum and taxane chemotherapy. The authors conducted a phase 1 study to evaluate the addition of oral everolimus to cisplatin and docetaxel as induction chemotherapy for head and neck cancer.. In this single-institution phase 1 study, 3 doses of daily everolimus were explored: 5 mg daily, 7.5 mg daily (administered as 5 mg daily alternating with 10 mg daily), and 10 mg daily of each 21-day cycle. Cisplatin and docetaxel doses were fixed (both were 75 mg/m(2) on day 1 of 21-day cycle) at each dose level with pegfilgrastim support. A standard 3 + 3 dose-escalation plan was used. After induction, patients were removed from protocol.. Eighteen patients were enrolled (15 men, 3 women), and their median Karnofsky performance status was 90. The most common toxicities were hyperglycemia, low hemoglobin, fatigue, and thrombocytopenia. Dose-limiting toxicities (DLTs) were neutropenic fever (1 event at dose level 2, 2 events at dose level 3), and all patients recovered fully from these DLTs. The maximum tolerated dose was exceeded at dose level 3. The progression-free survival rate at 1 year was 87.5% (95% confidence interval, 56.8%-96.7%); and, at 2 years, it was 76.6% (95% confidence interval, 41.2%-92.3%). Activating PI3K catalytic subunit α (PIK3CA) gene mutations were identified in 2 human papillomavirus-associated oropharyngeal cancers.. The phase 2 recommended dose was 7.5 mg daily for everolimus plus cisplatin and docetaxel (both at 75 mg/m(2) on day 1 of a 21-day cycle) given with pegfilgrastim support.

Topics: Adult; Aged; Antineoplastic Combined Chemotherapy Protocols; Cisplatin; Disease-Free Survival; Docetaxel; Drug Administration Schedule; Everolimus; Female; Head and Neck Neoplasms; Humans; Induction Chemotherapy; Male; Maximum Tolerated Dose; Middle Aged; Mutation; Neoplasm Staging; Papillomavirus Infections; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Risk Assessment; Risk Factors; Signal Transduction; Sirolimus; Smoking; Taxoids; TOR Serine-Threonine Kinases; Treatment Outcome

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

Human Papillomavirus (HPV) infection is the main risk factor for the development and progression of cervical cancer. HPV-16 E6 and E7 expression is essential for induction and maintenance of the transformed phenotype. These oncoproteins interfere with the function of several intracellular proteins, including those controlling the PI3K/AKT/mTOR pathway in which Phospolipase D (PLD) and Phosphatidic acid (PA) play a critical role.. PLD activity was measured in primary human keratinocytes transduced with retroviruses expressing HPV-16 E6, E7 or E7 mutants. The cytostatic effect of rapamycin, a well-known mTOR inhibitor with potential clinical applications, was evaluated in monolayer and organotypic cultures.. HPV-16 E7 expression in primary human keratinocytes leads to an increase in PLD expression and activity. Moreover, this activation is dependent on the ability of HPV-16 E7 to induce retinoblastoma protein (pRb) degradation. We also show that cells expressing HPV-16 E7 or silenced for pRb acquire resistance to the antiproliferative effect of rapamycin.. This is the first indication that HPV oncoproteins can affect PLD activity. Since PA can interfere with the ability of rapamycin to bind mTOR, the use of combined strategies to target mTOR and PLD activity might be considered to treat HPV-related malignancies.

Topics: Cell Proliferation; Cells, Cultured; Drug Resistance, Neoplasm; Gene Expression; Human papillomavirus 16; Humans; Keratinocytes; Models, Biological; Papillomavirus E7 Proteins; Papillomavirus Infections; Phospholipase D; Protein Binding; Retinoblastoma Protein; Sirolimus

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

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

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

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

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

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

The use of proliferative signal inhibitors (PSIs) in immunosuppression-related malignancies opens new roads for increasing the survival and quality of life in patients with solid organ transplantation. A 56-year-old female recipient of a living donor renal allograft (1990), who was immunosuppressed with cyclosporine (CsA; Neoral), azathioprine, and steroids, did initially well with acceptable renal function. During the last 5 years she required local therapy due to posterior vaginal lip human papillomavirus (HPV) lesions. In 2000, she discontinued azathioprine and the CsA doses were reduced to 100 mg daily. The local lesion showed a good response to reduced immunosuppression. In February 2005, the lesion reappeared and a biopsy showed malignancy. Local surgery was performed and CsA was replaced by everolimus (EVL; Certican). Two months after treatment initiation, the patient developed cough, dyspnea, and low-grade fever. Chest X-ray showed a lesion at the base of the left lung compatible with pneumonitis. After fiberbronchoscopy a diagnosis of bronchiolitis obliterans organizing pneumonia (BOOP) was obtained. She was treated with increased doses of oral steroids. EVL was never discontinued. The radiological lesion disappeared and the malignancy is currently in remission. In summary, a case of gynecological cancer in a renal transplant recipient was treated by surgical removal. After 1 year of immunosuppression with EVL, no recurrence has been observed. The adverse event (BOOP) was probably related to the PSI treatment and was controlled with an increased dose of steroids without discontinuing EVL.

Topics: Adrenal Cortex Hormones; Azathioprine; Cryptogenic Organizing Pneumonia; Cyclosporine; Everolimus; Female; Humans; Kidney Transplantation; Methylprednisolone; Middle Aged; Papillomavirus Infections; Polycystic Kidney Diseases; Postoperative Complications; Sirolimus; Vaginal Neoplasms

Dermatological complications following transplantation are very common and the majority of immunosuppressed transplant recipients develop some to many warts due to human papillomavirus (HPV) infection. In the setting of immunosuppression, therapeutic management may be disappointing because of the extent of the lesions in patients unable to develop a sufficient immune response directed against HPV. We report here a case of a young liver transplant recipient who developed diffuse recalcitrant HPV-induced warts leading to an impairment of her quality of life. Taking into account the antiproliferative and cytostatic properties of the target-of-rapamycin (TOR) inhibitors, a new class of immunosuppressive drug, we significantly modified the immunosuppressive regimen. Conversion to sirolimus was followed by a rapid improvement of cutaneous state suggesting that this strategy may be useful for recalcitrant cutaneous viral warts in transplant recipient.

Topics: Adolescent; Female; Humans; Immunosuppressive Agents; Liver Transplantation; Papillomavirus Infections; Sirolimus; Skin Diseases, Viral; Warts