Purpose: Gastrointestinal stromal tumors (GIST) generally harbor activating mutations in the receptor tyrosine kinase KIT or in the related platelet-derived growth factor receptor alpha (PDGFRA). GIST treated with imatinib mesylate or second-line therapies that target mutant forms of these receptors generally escape disease control and progress over time. Inhibiting additional molecular targets may provide more substantial disease control. Recent studies have implicated the PI3K/AKT pathway in the survival of imatinib mesylate–resistant GIST cell lines and tumors. Experimental Design: Here, we performed in vitro and in vivo studies evaluating the novel combination of imatinib mesylate with the AKT inhibitor MK-2206 in GIST. Whole-transcriptome sequencing (WTS) of xenografts was performed to explore the molecular aspects of tumor response to this novel combination and to potentially identify additional therapeutic targets in GIST. Results: This drug combination demonstrated significant synergistic effects in a panel of imatinib mesylate–sensitive and -resistant GIST cell lines. Furthermore, combination therapy provided significantly greater efficacy, as measured by tumor response and animal survival, in imatinib mesylate–sensitive GIST xenografts as compared with treatment with imatinib mesylate or MK-2206 alone. WTS implicated two neural genes, brain expressed X-linked 1 and neuronal pentraxin I, whose expression was significantly upregulated in combination-treated tumors compared with tumors treated with the two monotherapies. Conclusions: These studies provide strong preclinical justification for combining imatinib mesylate with an AKT inhibitor as a front-line therapy in GIST. In addition, the WTS implicated the BCL-2/BAX/BAD apoptotic pathway as a potential mechanism for this enhanced combination effect. Clin Cancer Res; 23(1); 171–80. ©2016 AACR.

Approximately 10–15 % of gastrointestinal stromal tumors (GISTs) lack gain of function mutations in the KIT and platelet-derived growth factor receptor alpha (PDGFRA) genes. An alternate mechanism of oncogenesis through loss of function of the succinate-dehydrogenase (SDH) enzyme complex has been identified for a subset of these “wild type” GISTs. Paired tumor and normal DNA from an SDH-intact wild-type GIST case was subjected to whole exome sequencing to identify the pathogenic mechanism(s) in this tumor. Selected findings were further investigated in panels of GIST tumors through Sanger DNA sequencing, quantitative real-time PCR, and immunohistochemical approaches. A hemizygous frameshift mutation (p.His2261Leufs*4), in the neurofibromin 1 (NF1) gene was identified in the patient’s GIST; however, no germline NF1 mutation was found. A somatic frameshift mutation (p.Lys54Argfs*31) in the MYC associated factor X (MAX) gene was also identified. Immunohistochemical analysis for MAX on a large panel of GISTs identified loss of MAX expression in the MAX-mutated GIST and in a subset of mainly KIT-mutated tumors. This study suggests that inactivating NF1 mutations outside the context of neurofibromatosis may be the oncogenic mechanism for a subset of sporadic GIST. In addition, loss of function mutation of the MAX gene was identified for the first time in GIST, and a broader role for MAX in GIST progression was suggested.

The insulin-like growth factor (IGF) pathway is implicated in the pathogenesis of hepatocellular carcinoma (HCC) and may be important in nonalcoholic fatty liver disease (NAFLD). The aim of this study is to determine expression levels of IGFs and receptors in NAFLD-associated HCC.

Purpose: Dedifferentiated liposarcoma (DDL) and leiomyosarcoma (LMS) are two common subtypes of soft-tissue sarcoma, a rare group of diseases for which new treatments are needed. Chemotherapy remains the standard option for advanced disease. Targeting cyclin-dependent kinase 4 and 6 (CDK4/6) in DDL and mTOR in LMS is of biologic interest. When combined, the CDK4 inhibitor ribociclib and the mTOR inhibitor everolimus have shown synergistic growth inhibition in multiple tumor models, suggesting that this combination could be beneficial in patients. Patients and Methods: This was a single arm, open label, multicenter phase II study of the combination of ribociclib and everolimus. Patients were enrolled into one of two cohorts: DDL or LMS with intact Rb. The primary endpoint was progression-free rate (PFR) at 16 weeks. Secondary endpoints included progression-free survival (PFS) and overall survival, safety and biomarker analyses. Results: In the DDL cohort, 33.3% [95% confidence interval (CI), 15.6%–55.3%] of patients were progression-free at 16 weeks. Median PFS in this cohort was 15.4 weeks (95% CI, 8–36 weeks) with 2 partial responses. In the LMS cohort the PFR at 16 weeks was 29.2% (95% CI, 12.6%–51.1%). Median PFS in this cohort was 15.7 weeks (95% CI, 7.7–NA). Most common toxicities included fatigue (66.7%), anorexia (43.8%), and hyperglycemia (43.8%). Concordance between Rb testing methodologies was poor. Conclusions: The combination of ribociclib and everolimus demonstrates activity in DDL with prolonged stable disease (≥16 weeks) meeting the primary endpoint. Notably partial responses were observed. The primary endpoint was not reached in the LMS cohort. The combination was well tolerated with expected side effects.

Most gastrointestinal stromal tumors (GISTs) harbor mutually exclusive gain of function mutations in the receptor tyrosine kinase (RTK) KIT (70–80%) or in the related receptor PDGFRA (~10%). These GISTs generally respond well to therapy with the RTK inhibitor imatinib mesylate (IM), although initial response is genotype-dependent. An alternate mechanism leading to GIST oncogenesis is deficiency in the succinate dehydrogenase (SDH) enzyme complex resulting from genetic or epigenetic inactivation of one of the four SDH subunit genes (SDHA, SDHB, SDHC, SDHD, collectively referred to as SDHX). SDH loss of function is generally seen only in GIST lacking RTK mutations, and SDH-deficient GIST respond poorly to imatinib therapy. Tumor and normal DNA from a GIST case carrying the IM-resistant PDGFRA D842V mutation was analyzed by whole exome sequencing (WES) to identify additional potential targets for therapy. The tumors analyzed were separate recurrences following progression on imatinib, sunitinib, and the experimental PDGFRA inhibitor crenolanib. Tumor sections from the GIST case and a panel of ~75 additional GISTs were subjected to immunohistochemistry (IHC) for the SDHB subunit. Surprisingly, a somatic, loss of function mutation in exon 4 of the SDHB subunit gene (c.291_292delCT, p.I97Mfs*21) was identified in both tumors. Sanger sequencing confirmed the presence of this inactivating mutation, and IHC for the SDHB subunit demonstrated that these tumors were SDH-deficient. IHC for the SDHB subunit across a panel of ~75 GIST cases failed to detect SDH deficiency in other GISTs with RTK mutations. This is the first reported case of a PDGFRA mutant GIST exhibiting SDH-deficiency. A brief discussion of the relevant GIST literature is included.

Although imatinib mesylate (IM) has transformed the treatment of gastrointestinal stromal tumors (GIST), many patients experience primary/secondary drug resistance. In a previous study, we identified a gene signature, consisting mainly of Kruppel-associated box (KRAB) domain containing zinc finger (ZNF) transcriptional repressors that predict short-term response to IM. To determine if these genes have functional significance, a siRNA library targeting these genes was constructed and applied to GIST cells in vitro. These screens identified seventeen “IM sensitizing genes” in GIST cells (sensitization index (SI) <0.85 ratio of drug/vehicle) with a false discovery rate (FDR) <15%, including twelve ZNF genes, the majority of which are located within the HSA19p12–13.1 locus. These genes were shown to be highly specific to IM and another tyrosine kinase inhibitor (TKI), sunitinib, in GIST cells. In order to determine mechanistically how these ZNFs might be modulating response to IM, RNAi approaches were used to individually silence genes within the predictive signature in GIST cells and expression profiling was performed. Knockdown of the 14 IM-sensitizing genes (10 ZNFs) universally led to downregulation of six genes, including TGFb3, periostin, and NEDD9. These studies implicate a role of KRAB-ZNFs in modulating response to TKIs in GIST.