(5-(2-4-bis((3s)-3-methylmorpholin-4-yl)pyrido(2-3-d)pyrimidin-7-yl)-2-methoxyphenyl)methanol and Pancreatic-Neoplasms

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

The mammalian target of rapamycin (mTOR) is dysregulated in diverse cancers and contributes to tumor progression and drug resistance. The first generation of mTOR inhibitors have failed to show clinical efficiency in treating pancreatic cancers due in part to the feedback relief of the insulin-like growth factor-1 receptor (IGF-1R)-AKT signaling pathway. The second generation of mTOR inhibitors, such as AZD8055, could inhibit AKT activation upon mTOR complex 2 (mTORC2) inhibition. However, whether this generation of mTOR inhibitors can obtain satisfactory activities in pancreatic cancer therapy remains unclear. In this study, we found AZD8055 did not show great improvement compared with everolimus, AZD8055 induced a temporal inhibition of AKT kinase activities and AKT was then rephosphorylated. Additionally, we found that AZD8055-induced transient AKT inhibition increased the expression and activation of epidermal growth factor receptor (EGFR) by releasing its transcriptional factors Fork-head box O 1/3a (FoxO1/3a), which might contribute to cell resistance to AZD8055. The in vitro and in vivo experiments further indicated the combination of AZD8055 and erlotinib synergistically inhibited the mTORC1/C2 signaling pathway, EGFR/AKT feedback activation, and cell growth, as well as suppressed the progression of pancreatic cancer in a xenograft model. This study provides a rationale and strategy for overcoming AZD8055 resistance by a combined treatment with the EGFR inhibitor erlotinib in pancreatic cancer therapy.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; ErbB Receptors; Erlotinib Hydrochloride; Female; Forkhead Transcription Factors; Humans; Morpholines; Pancreatic Neoplasms; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Quinazolines; Signal Transduction; TOR Serine-Threonine Kinases; Tumor Burden; Up-Regulation; Xenograft Model Antitumor Assays

Tyrosine kinase inhibitor BMS-777067 is an inhibitor of RON/MET receptor tyrosine kinases currently under clinical trials. Here, we report the synergistic activity of BMS-777607 in combination with mTOR inhibitor AZD8055 in killing chemoresistant pancreatic cancer and cancer stem cells. Treatment of pancreatic cancer L3.6pl cells with BMS-777607 alone inhibited clonogenic growth and moderately induced apoptotic death. However, BMS-777607 caused extensive polyploidy in L3.6pl cells through inhibition of aurora kinase B activity, independent of RON expression. In contrast, L3.6pl-derived cancer stem cells were highly resistant to BMS-777607-induced growth inhibition and apoptosis. The effect of BMS-777607 on induction of cancer stem cell polyploidy was also weak. BMS-777607-induced polyploidy features a predominant cell population with 8N chromosome content in both L3.6pl and cancer stem cells. These cells also showed decreased sensitivity toward chemotherapeutics by increased survival of IC(50) values in response to doxorubicin, cisplatin, methotrexate, 5-fluorouracial, and gemcitabine. Among a panel of chemical inhibitors that target different signaling proteins, we found that BMS-777607 in combination with mTOR inhibitor AZD8055 exerted synergistic effects on L3.6pl and cancer stem cells. More than 70% of L3.6pl and cancer stem cells lost their viability when both inhibitors were used. Specifically, BMS-777607 in combination with inhibition of mTORC2, but not mTORC1, was responsible for the observed synergism. Our findings demonstrate that BMS-777607 at therapeutic doses exerts inhibitory activities on pancreatic cancer cells but also induces polyploidy insensitive to chemotherapeutics. Combination of BMS-777607 with AZD8055 achieves the maximal cytotoxic effect on pancreatic cancer and cancer stem cells.

Topics: Aminopyridines; Apoptosis; Cell Line, Tumor; Drug Synergism; Humans; Morpholines; Neoplastic Stem Cells; Pancreatic Neoplasms; Polyploidy; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-met; Pyridones; Signal Transduction

Radiation exerts direct antitumor effects and is widely used in clinics, but the efficacy is severely compromised by tumor resistance. Therefore uncovering the mechanism of radioresistance might promote the development of new strategies to overcome radioresistance by manipulating activity of the key molecules.. Immunohistochemistry were used to find whether mTOR were over-activated in radioresistant patients' biopsies. Then Western blot, real-time PCR and transfection were used to find whether radiotherapy regulates the expression and activity of mTOR by modulating its targeting microRNA in human pancreatic cancer cell lines PANC-1, Capan-2 and BxPC-3. Finally efficacy of radiation combined with mTOR dual inhibitor AZD8055 was assessed in vitro and in vivo.. Ionizing radiation promoted mTOR expression and activation in pancreatic cancer cells through reducing miR-99b expression, which negatively regulated mTOR. Novel mTOR inhibitor, AZD8055 (10 nM, 100 nM, 500 nM) synergistically promoted radiation (0-10 Gy) induced cell growth inhibition and apoptosis. In human pancreatic cancer xenografts, fractionated radiation combined with AZD8055 treatment further increased the anti-tumor effect, the tumor volume was shrinked to 278 mm3 after combination treatment for 3 weeks compared with single radiation (678 mm3) or AZD8055 (708 mm3) treatment (P < 0.01).. Our data provide a rationale for overcoming radio-resistance by combined with mTOR inhibitor AZD8055 in pancreatic cancer therapy.

Topics: Animals; Apoptosis; Cell Cycle; Cell Line, Tumor; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Humans; Mice, Nude; MicroRNAs; Morpholines; Pancreatic Neoplasms; Phosphorylation; Radiation Tolerance; Radiation, Ionizing; TOR Serine-Threonine Kinases; Treatment Outcome; Up-Regulation; Xenograft Model Antitumor Assays