pyrimidinones and Carcinoma--Pancreatic-Ductal

There is paucity of investigations into immunotherapy or targeted therapy for postoperative locally recurrent pancreatic cancer. We aimed to assess the efficacy of stereotactic body radiotherapy (SBRT) plus pembrolizumab and trametinib in these patients.. In this open-label, randomised, controlled, phase 2 study, participants were recruited from Changhai Hospital affiliated to Naval Medical University, Shanghai, China. Eligible patients were aged 18 years or older with histologically confirmed pancreatic ductal adenocarcinoma characterised by mutant KRAS and positive immunohistochemical staining of PD-L1, Eastern Cooperative Oncology Group performance status of 0 or 1, and documented local recurrence after surgery followed by chemotherapy (mFOLFIRINOX or 5-fluorouracil). Eligible participants were randomly assigned (1:1) using an interactive voice or web response system, without stratification, to receive SBRT with doses ranging from 35-40 Gy in five fractions, intravenous pembrolizumab 200 mg once every 3 weeks, and oral trametinib 2 mg once daily or SBRT (same regimen) and intravenous gemcitabine (1000 mg/m. Between Oct 10, 2016, and Oct 28, 2017, 198 patients were screen, of whom 170 patients were enrolled and randomly assigned to receive SBRT plus pembrolizumab and trametinib (n=85) or SBRT plus gemcitabine (n=85). As of the clinical cutoff date (Nov 30, 2020), median follow-up was 23·3 months (IQR 20·5-27·4). Median overall survival was 24·9 months (23·3-26·5) with SBRT plus pembrolizumab and trametinib and 22·4 months (95% CI 21·2-23·6) with SBRT plus gemcitabine (hazard ratio [HR] 0·60 [95% CI 0·44-0·82]; p=0·0012). The most common grade 3 or 4 adverse effects were increased alanine aminotransferase or aspartate aminotransferase (ten [12%] of 85 in SBRT plus pembrolizumab and trametinib group vs six [7%] of 85 in SBRT plus gemcitabine group), increased blood bilirubin (four [5%] vs none), neutropenia (one [1%] vs nine [11%]), and thrombocytopenia (one [1%] vs four [5%]). Serious adverse events were reported by 19 (22%) participants in the SBRT plus pembrolizumab and trametinib group and 12 (14%) in the SBRT plus gemcitabine group. No treatment-related deaths occurred.. The combination of SBRT plus pembrolizumab and trametinib could be a novel treatment option for patients with locally recurrent pancreatic cancer after surgery. Phase 3 trials are needed to confirm our findings.. Shanghai Shenkang Center and Changhai Hospital.. For the Chinese translation of the abstract see Supplementary Materials section.

Topics: Aged; Antibodies, Monoclonal, Humanized; Carcinoma, Pancreatic Ductal; China; Combined Modality Therapy; Deoxycytidine; Female; Gemcitabine; Humans; Male; Middle Aged; Pancreatic Neoplasms; Pyridones; Pyrimidinones; Radiosurgery

Topics: Animals; Antineoplastic Agents; Apoptosis; beta Catenin; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Cell Proliferation; Gene Expression Regulation, Neoplastic; Heterocyclic Compounds, 4 or More Rings; Humans; Mice; Mice, Inbred NOD; Mice, SCID; Mitogen-Activated Protein Kinases; Pancreatic Neoplasms; Pyridones; Pyrimidinones; Tumor Cells, Cultured; Wnt1 Protein; Xenograft Model Antitumor Assays

Over the last decades, cytotoxic chemotherapy has been the cornerstone of metastatic pancreatic adenocarcinoma treatment. In late-stage disease, a range of treatment regimens still offers minor benefits. Molecular profiling studies have shown that pancreatic adenocarcinoma (PDAC) is a mutation-driven tumor type, with KRAS mutations found in approximately 90% of cases, which could partially explain the resistance to chemotherapy. Preclinical data on selective targeting of a downstream point of the RAF-MEK-ERK pathway with a MEK inhibitor along with the concurrent use of an autophagy inhibitor such as hydroxychloroquine appears to be one alternative approach to overcome resistance and inhibit cell proliferation.. We herein aim to investigate the rationale of autophagy inhibitors use and describe the outcomes of patients who received this experimental treatment.. Two patients have received this experimental regimen from January 2020 to the present date, achieving disease stabilization that is clinically meaningful, considering the chemoresistance scenario of the included patients.. Our real-life data regarding KRAS-mutated PDAC patients who received treatment with the MEK inhibitor trametinib combined with hydroxychloroquine after experiencing disease progression are consistent with the preclinical data, pointing to the clinical benefits of this regimen.

Topics: Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Autophagy; CA-19-9 Antigen; Carcinoma, Pancreatic Ductal; Disease Progression; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Female; Humans; Hydroxychloroquine; Male; MAP Kinase Signaling System; Middle Aged; Mutation; Neoplasm Staging; Pancreas; Pancreatic Neoplasms; Positron Emission Tomography Computed Tomography; Proto-Oncogene Proteins p21(ras); Pyridones; Pyrimidinones; Treatment Outcome

Autophagy and cell-cycle checkpoints act in concert to confer cellular radioresistance. We investigated the functional interaction between radiation-induced autophagy and G2 checkpoint activation in highly radioresistant human pancreatic ductal adenocarcinoma (PDAC) cells.. Four human PDAC cell lines (MIA PaCa-2, KP-4, Panc-1, and SUIT-2) were analyzed. These cells were first irradiated using x-rays, and their cell cycle status, autophagy, and cell cycle checkpoint marker expression and ATP production levels were evaluated. Autophagic flux assays and siRNA knockdown were used to evaluate autophagy activity. Double thymidine block experiments were performed to synchronize the cells. Two inhibitors (MK-1775 and SCH 900776) were used to attenuate G2 checkpoint activation. Cell survival assays and animal experiments were performed to evaluate the radiosensitizing effects of the G2 checkpoint inhibitors.. Autophagy and G2/M accumulation were synchronously induced in human PDAC cells with an activated G2 checkpoint at 12 hours after x-ray irradiation of 6 Gy. Radiation-induced autophagy produced the ATP levels required for cell survival. Double thymidine block experiments revealed that no autophagy occurred in cells that were solely in G2 phase. MK-1775 or SCH 900776 exposure attenuated not only G2 checkpoint activation but also postirradiation autophagy, indicating the dependence of radiation-induced autophagy on an activated G2 checkpoint. The inhibitors demonstrated a higher radiosensitizing effect in the PDAC cells than the autophagy inhibitor chloroquine. MK-1775 in combination with x-rays significantly suppressed the tumor growth of MIA PaCa-2 xenografts compared with other treatment groups, including radiation or drug exposure alone, to enhance the radiosensitivity of PDAC cells in vivo.. Biological crosstalk exists between the G2 checkpoint activation and radiation-induced autophagy processes that are believed to independently contribute to the radioresistance of human PDAC cells. These findings have important implications for the development of future radiation therapy strategies for PDAC.

Topics: Adenosine Triphosphate; Animals; Autophagy; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Female; G2 Phase Cell Cycle Checkpoints; Humans; Mice; Mice, Inbred BALB C; Pancreatic Neoplasms; Pyrazoles; Pyrimidinones; Quinolines; Radiation Tolerance; Thiazoles

Pancreatic cancer is a lethal solid malignancy with limited therapeutic options. The development of novel therapeutic drugs requires adequate new cell line models. A new pancreatic cancer cell line, designated PDXPC1, was established from one pancreatic ductal adenocarcinoma (PDAC) patient‑derived xenograft. The PDXPC1 cells were stably cultured for >2 years and had a stable short tandem repeat profile. The PDXPC1 cell line retained the key mutations of the primary tumor, along with the epithelial origin and other important protein expression. The PDXPC1 cells induced rapid in vivo tumor growth, both subcutaneously and orthotopically, in a mouse model with an elevated CA199 level. The PDXPC1 cells showed weak growth, invasion and migration potency compared to another pancreatic cancer cell line, but were relatively resistant to multiple anti‑cancer drugs. Interestingly, the MEK inhibitor trametinib significantly inhibited the proliferation of PDXPC1 cells, and not that of Panc‑1 cells, by inactivating MEK/ERK/MYC signaling and activating the apoptotic pathway via Bcl‑2 degradation. In conclusion, the PDXPC1 cell line, capturing the major characteristics of the primary tumor, may be a suitable tool for studying the underlying mechanisms of chemo‑resistance in PDAC and developing new targeted therapeutic options.

Topics: Animals; Carcinoma, Pancreatic Ductal; Cell Culture Techniques; Cell Proliferation; Cell Survival; Female; Humans; Mice; Mutation; Pancreatic Neoplasms; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Signal Transduction; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Molecular mechanisms underlying adaptive targeted therapy resistance in pancreatic ductal adenocarcinoma (PDAC) are poorly understood. Here, we identify SETD5 as a major driver of PDAC resistance to MEK1/2 inhibition (MEKi). SETD5 is induced by MEKi resistance and its deletion restores refractory PDAC vulnerability to MEKi therapy in mouse models and patient-derived xenografts. SETD5 lacks histone methyltransferase activity but scaffolds a co-repressor complex, including HDAC3 and G9a. Gene silencing by the SETD5 complex regulates known drug resistance pathways to reprogram cellular responses to MEKi. Pharmacological co-targeting of MEK1/2, HDAC3, and G9a sustains PDAC tumor growth inhibition in vivo. Our work uncovers SETD5 as a key mediator of acquired MEKi therapy resistance in PDAC and suggests a context for advancing MEKi use in the clinic.

Topics: Animals; Apoptosis; Carcinoma, Pancreatic Ductal; Cell Proliferation; Chromatin; Drug Resistance, Neoplasm; Female; Histocompatibility Antigens; Histone Deacetylases; Histone-Lysine N-Methyltransferase; Humans; MAP Kinase Kinase 1; MAP Kinase Kinase 2; Methyltransferases; Mice; Mice, Inbred C57BL; Mice, Inbred NOD; Mice, SCID; Molecular Targeted Therapy; Pancreatic Neoplasms; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Small Molecule Libraries; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Gene Expression Regulation, Neoplastic; Glycogen Synthase Kinase 3 beta; Humans; Inhibitory Concentration 50; Male; Mice; Pancreatic Neoplasms; Pharmacogenomic Testing; Pyridones; Pyrimidines; Pyrimidinones; Signal Transduction; Thiophenes; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

Although rare, NTRK gene fusions are known to be oncogenic drivers in pancreatic ductal adenocarcinoma (PDAC). We report the response of a metastatic CTRC-NTRK1 gene fusion-positive PDAC to targeted treatment with the oral tropomyosin receptor kinase (TRK) inhibitor larotrectinib and the eventual development of resistance to treatment.. A 61-year-old woman presented with a 2.5-cm mass in the body of the pancreas and a 1.2-cm liver lesion on routine follow-up for endometrial cancer that was in complete remission. Liver biopsy confirmed a primary PDAC unrelated to the endometrial cancer. The patient was treated with gemcitabine, nab-paclitaxel and ADI-PEG 20 for 12 months until disease progression and toxicity emerged [best overall response (BOR): partial response (PR)]. The patient switched to a modified regimen of folinic acid, fluorouracil, irinotecan and oxaliplatin for 4 months until neuropathy occurred. Oxaliplatin was withheld until disease progression 6 months later (BOR: stable disease). Despite recommencing oxaliplatin, the disease continued to progress. At this time, somatic profiling of the liver lesion revealed a CTRC-NTRK1 gene fusion. Treatment with larotrectinib 100 mg twice daily was commenced with BOR of PR at 2 months. The patient progressed after 6 months and was re-biopsied. Treatment was switched to the investigational next-generation TRK inhibitor selitrectinib (BAY 2731954, LOXO-195) 100 mg twice daily. After 2 months, the disease progressed and dabrafenibtrametinib combination therapy was initiated due to existence of a BRAF-V600E mutation. However, the cancer continued to progress and the patient died 2 months later.. Targeted TRK inhibition with larotrectinib in PDAC harbouring a CTRC-NTRK1 gene fusion is well tolerated and can improve quality of life for the patient. However, acquired resistance to therapy can emerge in some patients. Next-generation TRK inhibitors such as selitrectinib are currently in development to overcome this resistance (NCT02576431; NCT03215511).

Topics: Antineoplastic Combined Chemotherapy Protocols; Aza Compounds; Carcinoma, Pancreatic Ductal; Chymotrypsin; Female; Humans; Imidazoles; Middle Aged; Oncogene Proteins, Fusion; Oximes; Pancreatic Neoplasms; Protein Kinase Inhibitors; Protein Kinases; Pyrazoles; Pyridones; Pyrimidines; Pyrimidinones; Receptor, trkA

Clinical trials of EGFR inhibitors in combination with gemcitabine for the treatment of pancreatic ductal adenocarcinoma (PDAC) have generated mixed results partially due to the poorly defined effectiveness of EGFR inhibitors in PDAC. Here, we studied a panel of PDAC cell lines to compare the IC50s of the EGFR inhibitors gefitinib and cetuximab. We found that gefitinib induced biphasic inhibition in over 50% of PDAC cells, with the initial growth inhibition occurring at nanomolar concentrations and a second growth inhibition occurring outside the clinical range. In contrast to gefitinib, cetuximab produced a single phase growth inhibition in a subset of PDAC cells. Using this sensitivity data, we screened for correlations between cell morphology proteins and EGFR ligands to EGFR inhibitor sensitivity, and found that mesothelin and the EGFR ligand TGF-α have a strong correlation to gefitinib and cetuximab sensitivity. Analysis of downstream signaling pathways indicated that plc-γ1 and c-myc were consistently inhibited by EGFR inhibitor treatment in sensitive cell lines. While an inconsistent additive effect was observed with either cetuximab or gefitinib in combination with gemcitabine, the cell pathway data indicated consistent ERK activation, leading us to pursue EGFR inhibitors in combination with trametinib, a MEK1/2 inhibitor. Both cetuximab and gefitinib in combination with trametinib produced an additive effect in all EGFR sensitive cell lines. Our results indicate that mesothelin and TGF-α can predict PDAC sensitivity to EGFR inhibitors and a combination of EGFR inhibitors with trametinib could be a novel effective treatment for PDAC.

Topics: Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Cycle; Cell Proliferation; Cetuximab; Drug Resistance, Neoplasm; ErbB Receptors; Gefitinib; GPI-Linked Proteins; Humans; Mesothelin; Pancreatic Neoplasms; Pyridones; Pyrimidinones; Signal Transduction; Transforming Growth Factor alpha; Tumor Cells, Cultured

Standard chemotherapy for pancreatic ductal adenocarcinoma (PDAC), nab-paclitaxel (NPT) plus gemcitabine (Gem), has led to an average survival of 8.5 months. Presently, no therapeutics exist that effectively target the KRAS oncogene, activated in 95% of PDACs, but alternative strategies focus on inhibition of downstream effectors of KRAS signaling. Through combined inhibition of PI3K and MAPK signaling with MK-2206 (MK) and trametinib (Tra), enhancement of NPT + Gem response was evaluated. Median animal survival was significantly improved by the NPT + Gem combination (67% increase). Addition of MK-2206 or trametinib further increased median survival: NPT + Gem + MK (86%), NPT + Gem + Tra (105%), and NPT + Gem + MK + Tra (129%). In cell line-derived xenografts, the net tumor growth (in mm

Topics: Albumins; Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; Deoxycytidine; Drug Synergism; Female; Gemcitabine; Heterocyclic Compounds, 3-Ring; Humans; MAP Kinase Signaling System; Mice; Mice, Inbred NOD; Mice, SCID; Paclitaxel; Pancreatic Neoplasms; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Pyridones; Pyrimidinones; Xenograft Model Antitumor Assays

Advances in therapies targeting proteins and pathways affected by genetic alterations has raised the possibility of personalized cancer treatments.. The efficacy of targeting molecular aberrations was determined in the pancreatic ductal adenocarcinoma (PDAC) cell line, CAPAN2. Two mutations were targeted, KRAS (p.G12V) and ABL1 (p.G1060D), and cells were treated with regorafenib and trametinib, individually and in combination.. Exposure to either drug significantly increased cell death compared to the current standard of care, gemcitabine. Treatment with combinations of the drugs led to significant increases in cell death compared to either monotherapy. Strong additive/synergistic interactions were observed across a range of dosages and ratios, reducing dose requirements with potential clinical relevance.. The data obtained in this PDAC cell model: i) support the use of matched monotherapies; ii) indicate the effectiveness of matched combination therapies; and iii) provide potential proof-of-concept for precision medicine approach to cancer treatment.

Topics: Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Deoxycytidine; Drug Synergism; Gemcitabine; Humans; Pancreatic Neoplasms; Phenylurea Compounds; Piperazines; Protein Kinase Inhibitors; Pyridines; Pyridones; Pyrimidinones

Topics: Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Growth Processes; Drug Synergism; Heterocyclic Compounds, 3-Ring; Humans; MAP Kinase Kinase Kinases; Morpholines; Pancreatic Neoplasms; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Triazines

Activating mutations in the proto-oncogene KRAS are a hallmark of pancreatic ductal adenocarcinoma (PDAC), an aggressive malignancy with few effective therapeutic options. Despite efforts to develop KRAS-targeted drugs, the absolute dependence of PDAC cells on KRAS remains incompletely understood. Here we model complete KRAS inhibition using CRISPR/Cas-mediated genome editing and demonstrate that KRAS is dispensable in a subset of human and mouse PDAC cells. Remarkably, nearly all KRAS deficient cells exhibit phosphoinositide 3-kinase (PI3K)-dependent mitogen-activated protein kinase (MAPK) signaling and induced sensitivity to PI3K inhibitors. Furthermore, comparison of gene expression profiles of PDAC cells retaining or lacking KRAS reveal a role of KRAS in the suppression of metastasis-related genes. Collectively, these data underscore the potential for PDAC resistance to even the very best KRAS inhibitors and provide insights into mechanisms of response and resistance to KRAS inhibition.

Topics: Animals; Antineoplastic Agents; Benzimidazoles; Carcinoma, Pancreatic Ductal; DNA Copy Number Variations; Humans; Immunoblotting; Indazoles; Mice; Morpholines; Pancreatic Neoplasms; Phenylurea Compounds; Piperidines; Proto-Oncogene Mas; Proto-Oncogene Proteins p21(ras); Purines; Pyrimidines; Pyrimidinones; Quinazolinones; Sulfonamides; Thiazoles

Most patients with pancreatic ductal adenocarcinoma (PDAC) die within 5 years following resection plus adjuvant gemcitabine (Gem) from outgrowth of occult metastases. We hypothesized that inhibition of the KRAS pathway with the MEK inhibitor trametinib would inhibit the outgrowth of occult liver metastases in a preclinical model.. Liver metastases harvested from two patients with PDAC (Tumors 608, 366) were implanted orthotopically in mice. Tumor cell lines were derived and transduced with lentiviruses encoding luciferase and injected into spleens of mice generating microscopic liver metastases. Growth kinetics of liver metastases were measured with bioluminescent imaging and time-to-progression (TTP), progression-free survival (PFS), and overall survival (OS) were determined.. Trametinib (0.3 mg/kg BID) significantly prolonged OS versus control (Tumor 608: 114 vs. 43 days, p < 0.001; Tumor 366: not reached vs. 167 days, p = 0.0488). In vivo target validation demonstrated trametinib significantly reduced phosphorylated-ERK and expression of the ERK-responsive gene DUSP6. In a randomized, preclinical trial, mice were randomized to: (1) control, (2) adjuvant Gem (100 mg/kg IP, Q3 days) × 7 days followed by surveillance, or (3) adjuvant Gem followed by trametinib. Sequential Gem-trametinib significantly decreased metastatic cell outgrowth and increased TTP and PFS.. Treatment of mice bearing micrometastases with trametinib significantly delayed tumor outgrowth by effectively inhibiting KRAS-MEK-ERK signaling. In a randomized, preclinical, murine trial adjuvant sequential Gem followed by trametinib inhibited occult metastatic cell outgrowth in the liver and increased PFS versus adjuvant Gem alone. An adjuvant trial of sequential Gem-trametinib is being planned in patients with resected PDAC.

Topics: Animals; Carcinoma, Pancreatic Ductal; Humans; Liver Neoplasms; Mice; Pancreatic Neoplasms; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Pancreatic ductal adenocarcinoma (PDAC) is a highly deadly malignancy. Expression of the stem cell transcription factor SOX2 increases during progression of PDAC. Knockdown of SOX2 in PDAC cell lines decreases growth in vitro; whereas, stable overexpression of SOX2 in one PDAC cell line reportedly increases growth in vitro. Here, we reexamined the role of SOX2 in PDAC cells, because inducible SOX2 overexpression in other tumor cell types inhibits growth. In this study, four PDAC cell lines were engineered for inducible overexpression of SOX2 or inducible knockdown of SOX2. Remarkably, inducible overexpression of SOX2 in PDAC cells inhibits growth in vitro and reduces tumorigenicity. Additionally, inducible knockdown of SOX2 in PDAC cells reduces growth in vitro and in vivo. Thus, growth and tumorigenicity of PDAC cells is highly dependent on the expression of optimal levels of SOX2 - a hallmark of molecular rheostats. We also determined that SOX2 alters the responses of PDAC cells to drugs used in PDAC clinical trials. Increasing SOX2 reduces growth inhibition mediated by MEK and AKT inhibitors; whereas knockdown of SOX2 further reduces growth when PDAC cells are treated with these inhibitors. Thus, targeting SOX2, or its mode of action, could improve the treatment of PDAC.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; Doxorubicin; Female; Heterocyclic Compounds, 3-Ring; Humans; Mice; Mice, Nude; Mitogen-Activated Protein Kinase Kinases; Pancreatic Neoplasms; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins p21(ras); Pyridones; Pyrimidinones; RNA Interference; RNA, Small Interfering; SOXB1 Transcription Factors

Pancreatic ductal adenocarcinoma (PDAC) harbors the worst prognosis of any common solid tumor, and multiple failed clinical trials indicate therapeutic recalcitrance. Here, we use exome sequencing of patient tumors and find multiple conserved genetic alterations. However, the majority of tumors exhibit no clearly defined therapeutic target. High-throughput drug screens using patient-derived cell lines found rare examples of sensitivity to monotherapy, with most models requiring combination therapy. Using PDX models, we confirmed the effectiveness and selectivity of the identified treatment responses. Out of more than 500 single and combination drug regimens tested, no single treatment was effective for the majority of PDAC tumors, and each case had unique sensitivity profiles that could not be predicted using genetic analyses. These data indicate a shortcoming of reliance on genetic analysis to predict efficacy of currently available agents against PDAC and suggest that sensitivity profiling of patient-derived models could inform personalized therapy design for PDAC.

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Dasatinib; Docetaxel; Drug Resistance, Neoplasm; Everolimus; Exome; Humans; Mice; Models, Genetic; Models, Statistical; Pancreatic Neoplasms; Precision Medicine; Prognosis; Pyridones; Pyrimidinones; Taxoids; Xenograft Model Antitumor Assays

Pancreatic ductal adenocarcinoma (PDA) is a lethal disease, in part, because of the lack of effective targeted therapeutic options. MK-1775 (also known as AZD1775), a mitotic inhibitor, has been demonstrated to enhance the anti-tumor effects of DNA damaging agents such as gemcitabine. We evaluated the efficacy of MK-1775 alone or in combination with DNA damaging agents (MMC or oxaliplatin) in PDA cell lines that are either DNA repair proficient (DDR-P) or deficient (DDR-D). PDA cell lines PL11, Hs 766T and Capan-1 harboring naturally selected mutations in DNA repair genes FANCC, FANCG and BRCA2 respectively, were less sensitive to MK-1775 as compared to two out of four representative DDR-P (MIA PaCa2 and PANC-1) cell lines. Accordingly, DDR-P cells exhibit reduced sensitivity to MK-1775 upon siRNA silencing of DNA repair genes, BRCA2 or FANCD2, compared to control cells. Only DDR-P cells showed increased apoptosis as a result of early mitotic entry and catastrophe compared to DDR-D cells. Taken together with other recently published reports, our results add another level of evidence that the efficacy of WEE1 inhibition is influenced by the DNA repair status of a cell and may also be dependent on the tumor type and model evaluated.

Topics: Antineoplastic Agents; Apoptosis; Carcinoma, Pancreatic Ductal; Cell Cycle Proteins; Cell Line, Tumor; DNA Damage; DNA Repair; DNA Repair Enzymes; Drug Resistance, Neoplasm; Drug Synergism; Humans; Inhibitory Concentration 50; Mitomycin; Mitosis; Mutagens; Mutation; Nuclear Proteins; Organoplatinum Compounds; Oxaliplatin; Pancreatic Neoplasms; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones

Recently, the transient receptor potential melastatin 8 (TRPM8) channel has emerged as a putative biomarker for pancreatic ductal adenocarcinoma (PDA). This study aimed to evaluate the expression of TRPM8 and its modulation by specific agonists and antagonists in PDA cells.. We examined the protein expression of TRPM8 in 3 different PDA cell lines and compared it with a nontumoral epithelial cell line of human pancreatic origin using Western blotting and immunocytochemical analysis. To assess the function of TRPM8 channels, we measured the TRPM8 currents in whole-cell mode of the patch clamp technique. To explore the putative involvement of TRPM8 in cell migration, we investigated the motility of PDA cells using the scratch-wound assay.. Pancreatic ductal adenocarcinoma cells express functional plasma membrane TRPM8 channels, which are responsive after exposure to agonists (menthol and icilin) and antagonists N-(3-aminopropyl)-2-{[(3-methylphenyl) methyl]oxy}-N-(2-thienylmethyl)benzamide hydrochloride salt. The silencing of TRPM8 expression by small interfering RNA augments the migration of PDA cells. Conversely, the activated form of TRPM8 inhibits PDA cell motility.. An unglycosylated TRPM8 protein is expressed and is functional in the membrane of PDA cells. Transient receptor potential melastatin 8 inhibits the migration of PDA cells, suggesting a putative role as a biomarker or target for this channel for PDA therapy.

Topics: Adenocarcinoma; Benzamides; Blotting, Western; Carcinoma, Pancreatic Ductal; Cell Line; Cell Line, Tumor; Cell Membrane; Cell Movement; HEK293 Cells; Humans; Immunohistochemistry; Membrane Potentials; Membrane Proteins; Menthol; Pancreatic Neoplasms; Patch-Clamp Techniques; Pyrimidinones; RNA Interference; Thiophenes; TRPM Cation Channels

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer due in part to a lack of highly robust cytotoxic or molecular-based therapies. Recent studies investigating ligand-mediated Wnt/β-catenin signaling have highlighted its importance in pancreatic cancer initiation and progression, as well as its potential as a therapeutic target in PDAC. The small-molecule ICG-001 binds cAMP-responsive element binding (CREB)-binding protein (CBP) to disrupt its interaction with β-catenin and inhibit CBP function as a coactivator of Wnt/β-catenin-mediated transcription. Given its ability to inhibit Wnt/β-catenin-mediated transcription in vitro and in vivo, as well as its efficacy in preclinical models of colorectal cancer and other Wnt-driven diseases, we examined ICG-001 and its potential role as a therapeutic in PDAC. ICG-001 alone significantly inhibited anchorage-dependent and -independent growth of multiple PDAC lines, and augmented in vitro growth inhibition when used in combination with gemcitabine. ICG-001 had only variable modest effects on PDAC apoptosis and instead mediated PDAC growth inhibition primarily through robust induction of G₁ cell-cycle arrest. These effects, however, seemed decoupled from its inhibition of Wnt/β-catenin-mediated transcription. DNA microarrays performed on PDAC cells in the context of ICG-001 treatment revealed ICG-001 altered the expression of several genes with well-established roles in DNA replication and cell-cycle progression, including direct actions on SKP2 and CDKN1A. ICG-001 also significantly prolonged survival in an in vivo orthotopic xenograft model of PDAC, indicating ICG-001 or derived compounds that disrupt CBP activity are potentially useful small-molecule therapeutics for pancreatic cancer.

Topics: Animals; Apoptosis; Bridged Bicyclo Compounds, Heterocyclic; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; CREB-Binding Protein; Female; G1 Phase Cell Cycle Checkpoints; Gene Expression; Humans; Mice; Mice, Nude; Pancreatic Neoplasms; Pyrimidinones; Random Allocation; Tissue Array Analysis; Xenograft Model Antitumor Assays

Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations and epidermal growth factor receptor (EGFR) family signaling are drivers of tumorigenesis in pancreatic ductal adenocarcinoma (PDAC). Previous studies have demonstrated that combinatorial treatment of PDAC xenografts with the mitogen-activated protein kinase-extracellular-signal-regulated kinase (ERK) kinase1/2 (MEK1/2) inhibitor trametinib and the dual EGFR/human epidermal growth factor receptor 2 (HER2) inhibitor lapatinib provided more effective inhibition than either treatment alone. In this study, we have used the therapeutic antibodies, panitumumab (specific for EGFR) and trastuzumab (specific for HER2), to probe the role of EGFR and HER2 signaling in the proliferation of patient-derived xenograft (PDX) tumors. We show that dual anti-EGFR and anti-HER2 therapy significantly augmented the growth inhibitory effects of the MEK1/2 inhibitor trametinib in three different PDX tumors. While significant growth inhibition was observed in both KRAS mutant xenograft groups receiving trametinib and dual antibody therapy (tumors 366 and 608), tumor regression was observed in the KRAS wild-type xenografts (tumor 738) treated in the same manner. Dual antibody therapy in conjunction with trametinib was equally or more effective at inhibiting tumor growth and with lower apparent toxicity than trametinib plus lapatinib. Together, these studies provide further support for a role for EGFR and HER2 in pancreatic cancer proliferation and underscore the importance of therapeutic intervention in both the KRAS-rapidly accelerated fibrosarcoma kinase (RAF)-MEK-ERK and EGFR-HER2 pathways to achieve maximal therapeutic efficacy in patients.

Topics: Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Disease Models, Animal; Drug Therapy, Combination; ErbB Receptors; Humans; Male; Mitogen-Activated Protein Kinases; Mutation; Pancreatic Neoplasms; Panitumumab; Phosphorylation; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Pyridones; Pyrimidinones; Receptor, ErbB-2; Signal Transduction; Trastuzumab; Tumor Burden; Xenograft Model Antitumor Assays

Several lines of evidence indicate that Sirt1, a class III histone deacetylase (HDAC) is implicated in the initiation and progression of malignancies and thus gained attraction as druggable target. Since data on the role of Sirt1 in pancreatic ductal adenocarcinoma (PDAC) are sparse, we investigated the expression profile and prognostic significance of Sirt1 in vivo as well as cellular effects of Sirt1 inhibition in vitro.. Sirt1 expression was analyzed by immunohistochemistry in a large cohort of PDACs and correlated with clinicopathological and survival data. Furthermore, we investigated the impact of overexpression and small molecule inhibition on Sirt1 in pancreatic cancer cell culture models including combinatorial treatment with chemotherapy and EGFR-inhibition. Cellular events were measured quantitatively in real time and corroborated by conventional readouts including FACS analysis and MTT assays.. We detected nuclear Sirt1 expression in 36 (27.9%) of 129 PDACs. SIRT1 expression was significantly higher in poorly differentiated carcinomas. Strong SIRT1 expression was a significant predictor of poor survival both in univariate (p = 0.002) and multivariate (HR 1.65, p = 0.045) analysis. Accordingly, overexpression of Sirt1 led to increased cell viability, while small molecule inhibition led to a growth arrest in pancreatic cancer cells and impaired cell survival. This effect was even more pronounced in combinatorial regimens with gefitinib, but not in combination with gemcitabine.. Sirt1 is an independent prognosticator in PDACs and plays an important role in pancreatic cancer cell growth, which can be levered out by small molecule inhibition. Our data warrant further studies on SIRT1 as a novel chemotherapeutic target in PDAC.

Topics: Aged; Aged, 80 and over; Carcinoma, Pancreatic Ductal; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Follow-Up Studies; Gefitinib; Gene Expression Regulation, Neoplastic; Humans; Middle Aged; Naphthalenes; Neoplasm Grading; Neoplasm Metastasis; Neoplasm Staging; Niacinamide; Pancreatic Neoplasms; Prognosis; Pyrimidinones; Quinazolines; Sirtuin 1

Investigate the efficacy and pharmacodynamic effects of MK-1775, a potent Wee1 inhibitor, in both monotherapy and in combination with gemcitabine (GEM) using a panel of p53-deficient and p53 wild-type human pancreatic cancer xenografts.. Nine individual patient-derived pancreatic cancer xenografts (6 with p53-deficient and 3 with p53 wild-type status) from the PancXenoBank collection at Johns Hopkins were treated with MK-1775, GEM, or GEM followed 24 hour later by MK-1775, for 4 weeks. Tumor growth rate/regressions were calculated on day 28. Target modulation was assessed by Western blotting and immunohistochemistry.. MK-1775 treatment led to the inhibition of Wee1 kinase and reduced inhibitory phosphorylation of its substrate Cdc2. MK-1775, when dosed with GEM, abrogated the checkpoint arrest to promote mitotic entry and facilitated tumor cell death as compared to control and GEM-treated tumors. MK-1775 monotherapy did not induce tumor regressions. However, the combination of GEM with MK-1775 produced robust antitumor activity and remarkably enhanced tumor regression response (4.01-fold) compared to GEM treatment in p53-deficient tumors. Tumor regrowth curves plotted after the drug treatment period suggest that the effect of the combination therapy is longer-lasting than that of GEM. None of the agents produced tumor regressions in p53 wild-type xenografts.. These results indicate that MK-1775 selectively synergizes with GEM to achieve tumor regressions, selectively in p53-deficient pancreatic cancer xenografts.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Pancreatic Ductal; Cell Cycle Proteins; Cell Line, Tumor; Deoxycytidine; Disease Progression; Drug Synergism; Female; Gemcitabine; Genes, p53; Humans; Mice; Mice, Nude; Mutation; Neoplasms; Nuclear Proteins; Pancreatic Neoplasms; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Tumor Burden; Xenograft Model Antitumor Assays