Project description:In addition to KIT and PDGFRA mutations, sequential accumulation of other genetic events is involved in the development and progression of gastrointestinal stromal tumors (GISTs). Until recently, the significance of these other alterations has not been thoroughly investigated. The combination of gene expression profiling and high-resolution genomic copy number analysis offers a detailed molecular portrait of GISTs, providing an essential comprehensive knowledge necessary to guide the discovery of novel target genes involved in tumor development and progression. Fresh tissue specimens from 25 patients with GIST were collected and high-resolution genomic copy number analyses were performed using Affymetrix SNP array 6.0. GIST tumor samples from mutated (KIT or PDGFRA) or Wild Type patients were labeled for hybridization on Affymetrix microarrays. Copy number analysis of Affymetrix SNP6.0 arrays was performed for 25 GIST samples, then compared to gene expression data.
Project description:Activating mutation of KIT is well known as a key molecular event for the development of gastrointestinal stromal tumors(GISTs). Dysregulation of microRNAs(miRNA) might elucidate KIT mutation, KIT overexpression and the resulting tumorigenesis in GIST. Herein we identified miRNA expression profiles that associated with KIT mutation and KIT overexpression in GIST by miRNA microarrays and Real-time PCR in GISTs. The potentially target genes of selected miRNAs were analyzed by bioinformatic techniques with GO and KEGG pathway analysis. We showed that 6 miRNAs were differentially expressed in CD117IHC+/KITmutant GISTs compared to CD117IHC-/wild-type GISTs. Of these, 2 miRNAs including miR-483-3p and miR-589 were up-regulated, while the other 4 miRNAs including miR-140-5p, miR-148b-3p, miR-1587 and miR-4507 were down-regulated. GO and KEGG analysis demonstrated that miRNAs with significant change were involved in regulation of target genes related to the development of GIST. Among the candidate miRNAs studied, miR-148b-3p and miR-140-5p may be involved in GIST tumorigenesis via targeting mutant KIT or via intermediate molecules of PDGFRA, PI3K-AKT and MAPK pathway, such as AKT2, MAPK1, MAPK10, STAT5A, SMAD4, SMAD5 and PTEN. Furthermore, the reduced expression of miR-140-5p and miR-148b-3p were inversely correlated with high-risk grade, recurrence and metastasis of GIST. The current findings indicated that miR-148b-3p and miR-140-5p were not only involved in tumorigenesis of GIST, but might also participate in the progression of GIST and could be considered as novel biomarkers for potentially predicting the prognosis of GIST.
Project description:Purpose: Management of gastrointestinal stromal tumor (GIST) has been revolutionized by the identification of activating mutations in KIT and PDGFRA and the clinical application of receptor tyrosine kinase (RTK) inhibitors in the advanced disease setting. Stratification of GIST into molecularly defined subsets provides insight into clinical behavior and response to approved targeted therapies. Although these RTK inhibitors are effective in the majority of GIST, resistance to these agents remains a significant clinical obstacle. Development of effective treatment strategies for refractory GIST 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 that are essential in GIST. Experimental Design: Using Multiplexed Inhibitor Beads and Mass Spectrometry paired with a super-SILAC kinome standard, we explored the majority of the kinome in GIST specimens from three GIST subtypes (KIT-mutant, PDGFRA-mutant and succinate dehydrogenase-deficient GIST) to identify novel kinase targets. In vitro and in vivo studies were performed to evaluate the utility of targeting the identified kinases in GIST. Results: Kinome profiling revealed distinct signatures in three GIST subtypes. PDGFRA-mutant GIST had elevated tumor associated macrophage (TAM) kinases and immunohistochemical analysis confirmed increased TAMs present in these tumors. Kinome profiling with loss-of-function assays revealed a significant role for G2-M tyrosine kinase, Wee1, in GIST survival. In vitro and in vivo studies revealed significant efficacy of MK-1775 (Wee1 inhibitor) in combination with avapritinib in both KIT and PDGFRA-mutant GIST cell lines, as well as notable efficacy of MK-1775 as a single agent in the PDGFRA-mutant line. Conclusions: These studies provide strong preclinical justification for the use of MK-1775 in GIST.
Project description:Secondary mutation of KIT/PDGFRA is the main reason that contributes to the multi-drug resistance phenotype of advanced GIST in the clinical setting. SHP2 is a specific signal transducer of KIT. Therefore, the combination of approved KIT/PDGFRA kinase inhibitors and SHP2 inhibitors is a promising treatment strategy for advanced GIST patients. Here, we treated GIST cells with the SHP2 inhibitor SHP099 and sequenced the total mRNA to analyze the effect of SHP2 inhibition on GIST cells.
Project description:In addition to KIT and PDGFRA mutations, sequential accumulation of other genetic events is involved in the development and progression of gastrointestinal stromal tumors (GISTs). Until recently, the significance of these other alterations has not been thoroughly investigated. The combination of gene expression profiling and high-resolution genomic copy number analysis offers a detailed molecular portrait of GISTs, providing an essential comprehensive knowledge necessary to guide the discovery of novel target genes involved in tumor development and progression. Fresh tissue specimens from 25 patients with GIST were collected and high-resolution genomic copy number analyses were performed using Affymetrix SNP array 6.0.