mk-1775 and Gastrointestinal-Stromal-Tumors

Management of gastrointestinal stromal tumors (GISTs) has been revolutionized by the identification of activating mutations in KIT and PDGFRA and clinical application of RTK inhibitors in advanced disease. Stratification of GISTs into molecularly defined subsets provides insight into clinical behavior and response to approved targeted therapies. Although these RTK inhibitors are effective in most GISTs, resistance remains a significant clinical problem. Development of effective treatment strategies for refractory GISTs requires identification of novel targets to provide additional therapeutic options. Global kinome profiling has the potential to identify critical signaling networks and reveal protein kinases essential in GISTs. Using multiplexed inhibitor beads and mass spectrometry, we explored the majority of the kinome in GIST specimens from the 3 most common molecular subtypes (KIT mutant, PDGFRA mutant, and succinate dehydrogenase deficient) to identify kinase targets. Kinome profiling with loss-of-function assays identified an important role for G2/M tyrosine kinase, Wee1, in GIST cell survival. In vitro and in vivo studies revealed significant efficacy of MK-1775 (Wee1 inhibitor) in combination with avapritinib in KIT mutant and PDGFRA mutant GIST cell lines as well as notable efficacy of MK-1775 as a monotherapy in the engineered PDGFRA mutant line. These studies provide strong preclinical justification for the use of MK-1775 in GIST.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Cell Cycle Proteins; Cell Line, Tumor; Cell Survival; Drug Resistance, Neoplasm; Female; Gastrointestinal Stromal Tumors; Humans; Male; Mice; Mice, SCID; Mutation; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-kit; Pyrazoles; Pyrimidinones; Pyrroles; Receptor, Platelet-Derived Growth Factor alpha; Signal Transduction; Triazines; Xenograft Model Antitumor Assays

Activating mutation of KIT or PDGFRA is the primary molecular mechanism for gastrointestinal stromal tumors (GISTs). Although imatinib has a revolutionary effect on GIST therapeutics, the benefits are not durable. Increasing reports have demonstrated that cell cycle checkpoint plays critical roles in GIST. Here, we explore the role of WEE1 kinase in GIST progression.. Oncomine public database, western blotting, and immunohistochemistry were used to analyze WEE1 expression in GISTs. Using MTT assays, colony formation analysis, and flow cytometry, we examined the role of WEE1 in GIST cells and the antitumor activity of the inhibitor MK1775 alone, or in combination with imatinib. Cycloheximide chase assay and pharmacological inhibition of autophagy and proteasome pathway were performed to analyze KIT expression. Additionally, autophagic markers Beclin1 and LC3B were detected by western blotting.. Upregulated WEE1 expression was observed in GIST tissues and correlated with tumor size, mitotic count, and risk grade. Inhibition of WEE1 significantly suppressed GIST cell proliferation, induced apoptosis and cell cycle arrest. Imatinib and MK1775 co-treatment markedly enhanced the antitumor activity. Targeting WEE1 decreased the expression of KIT expression. Moreover, WEE1 stabilized KIT protein and KIT reduction observed upon WEE1 inhibition could be reversed by pharmacological inhibition of autophagy, but not proteasome pathway. WEE1 inhibition also increased Beclin1 expression and LC3B II/I ratio in GIST cells.. Our data suggest that WEE1 plays a pivotal role in GIST proliferation. WEE1 inhibition could promote KIT autophagic degradation and, therefore, targeting WEE1 might represent a novel strategy for GIST therapies.

Topics: Aged; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Autophagy; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Female; Gastrointestinal Neoplasms; Gastrointestinal Stromal Tumors; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Imatinib Mesylate; Male; Middle Aged; Molecular Targeted Therapy; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-kit; Pyrazoles; Pyrimidinones