Evaluating new therapies in gastrointestinal stromal tumor using in vivo molecular optical imaging

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors in the US. The majority (~85%) of GISTs possess gain-of-function mutations in KIT or PDGFRA, causing constitutive activation of the kinase receptor. GIST management has been transformed by the identification of tumor driver mutations leading to unprecedented disease control of advanced GIST with the introduction of imatinib mesylate (IM). Despite IM’s efficacy, most patients experience primary and/or secondary resistance within 2 y of treatment. Additional therapies and methods to optimize screening of novel approaches in preclinical studies are warranted. Clinically, treatment efficacy is typically assessed using Response Evaluation Criteria In Solid Tumors (RECIST) guidelines or Choi criteria. Both require a period of time on therapy before changes indicative of response can be observed. In addition, neither informs directly about cell death. We evaluated the use of molecular imaging technology in an animal model using near-infrared (NIR) imaging probes together with three-dimensional fluorescence molecular tomography (FMT) for assessing therapeutic response and ultimately optimizing our understanding of the biologic effects of these agents. We determined the potential of NIR probes (PSVue(TM)794 and cell-penetrating KcapQ647) for detecting distinct markers of apoptosis and compare this to tumor size measured by MRI in response to IM treatment in GIST-T1 xenografts. Our studies revealed statistically significant increases in apoptosis due to IM treatment using both probes as early as 24 h post IM treatment which was confirmed by IHC. Molecular imaging will allow for faster and more effective screening of novel therapies in preclinical GIST models

Succinate dehydrogenase deficiency in a PDGFRA mutated GIST

Abstract Background 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. Methods 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. Results 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. Conclusions This is the first reported case of a PDGFRA mutant GIST exhibiting SDH-deficiency. A brief discussion of the relevant GIST literature is included

ZNF-Mediated Resistance to Imatinib Mesylate in Gastrointestinal Stromal Tumor

<div><p>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 <em>in vitro</em>. 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.</p> </div