Dataset Information

Molecular characterization of metastatic exon 11 mutant gastrointestinal stromal tumors (GIST) beyond KIT/PDGFR? genotype evaluated by next generation sequencing (NGS).

ABSTRACT: About 85% of GISTs are associated with KIT and PDGFR? gene mutations, which predict response to tyrosine kinase inhibitors. Although the outcomes in patients affected by GIST have dramatically improved, tumor progression control still remains a challenge. The aim of this study is the genomic characterization of individual metastatic KIT-exon 11-mutant GIST to identify additional aberrations and simultaneous molecular events representing potential therapeutic targets.Seven patients with metastatic GIST were studied with whole transcriptome sequencing and copy number analysis. Somatic single nucleotide variations were called; however, no shared mutated genes were detected except KIT. Almost all patients showed loss of genomic regions containing tumor suppressor genes, sometimes coupled with single nucleotide mutation of the other allele. Additionally, six fusion transcripts were found and three patients showed amplifications involving known oncogenes.Evaluating the concordance between CN status and mRNA expression levels, we detected overexpression of CCND2 and EGFR and silencing of CDKN2A, CDKN2C, SMARCB1, PTEN and DMD. Altered expression of these genes could be responsible for aberrant activation of signaling pathways that support tumor growth. In this work, we assessed the effect of Hedgehog pathway inhibition in GIST882 cells, which causes decrement of cell viability associated with reduction of KIT expression.Additional genomic alterations not previously reported in GIST were found even if not shared by all samples. This contributes to a more detailed molecular understanding of this disease, useful for identification of new targets and novel therapeutics and representing a possible point of departure for a truly individualized clinical approach.

SUBMITTER: Saponara M

PROVIDER: S-EPMC4747222 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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Molecular characterization of metastatic exon 11 mutant gastrointestinal stromal tumors (GIST) beyond KIT/PDGFRα genotype evaluated by next generation sequencing (NGS).

Saponara Maristella M Urbini Milena M Astolfi Annalisa A Indio Valentina V Ercolani Giorgio G Del Gaudio Massimo M Santini Donatella D Pirini Maria Giulia MG Fiorentino Michelangelo M Nannini Margherita M Lolli Cristian C Mandrioli Anna A Gatto Lidia L Brandi Giovanni G Biasco Guido G Pinna Antonio Daniele AD Pantaleo Maria Abbondanza MA

Oncotarget 20151201 39

About 85% of GISTs are associated with KIT and PDGFRα gene mutations, which predict response to tyrosine kinase inhibitors. Although the outcomes in patients affected by GIST have dramatically improved, tumor progression control still remains a challenge. The aim of this study is the genomic characterization of individual metastatic KIT-exon 11-mutant GIST to identify additional aberrations and simultaneous molecular events representing potential therapeutic targets.Seven patients with metastati ...[more]

PMID: 26544626

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Project description:Gastrointestinal stromal tumors (GISTs) are common mesenchymal neoplasms in the gastrointestinal tract of humans and dogs. Little is known about the pathogenesis of these tumors. This study evaluated the role of c-KIT in canine GISTs; specifically, we investigated activating mutations in exons 8, 9, 11, 13, and 17 of c-KIT and exons 12, 14, and 18 of platelet-derived growth factor receptor, alpha polypeptide (PDGFRA), all of which have been implicated in human GISTs.Seventeen canine GISTs all confirmed to be positive for KIT immunostaining were studied. Exons 8, 9, 11, 13 and 17 of c-KIT and exons 12, 14, and 18 of PDGFRA, were amplified from DNA isolated from formalin-fixed paraffin-embedded samples.Of these seventeen cases, six amplicons of exon 11 of c-KIT showed aberrant bands on gel electrophoresis. Sequencing of these amplicons revealed heterozygous in-frame deletions in six cases. The mutations include two different but overlapping six base pair deletions. Exons 8, 9, 13, and 17 of c-KIT and exons 12, 14, and 18 of PDGFRA had no abnormalities detected by electrophoresis and sequencing did not reveal any mutations, other than synonymous single nucleotide polymorphisms (SNPs) found in exon 11 of c-KIT and exons 12 and 14 of PDGFRA.The deletion mutations detected in canine GISTs are similar to those previously found in the juxtamembrane domain of c-KIT in canine cutaneous mast cell tumors in our laboratory as well as to those reported in human GISTs. Interestingly, none of the other c-KIT or PDGFRA exons showed any abnormalities in our cases. This finding underlines the critical importance of c-KIT in the pathophysiology of canine GISTs. The expression of KIT and the identification of these activating mutations in c-KIT implicate KIT in the pathogenesis of these tumors. Our results indicate that mutations in c-KIT may be of prognostic significance and that targeting KIT may be a rational approach to treatment of these malignant tumors. This study further demonstrates that spontaneously occurring canine GISTs share molecular features with human GISTs and are an appropriate model for human GISTs.

Project description:Background and aimsConsidering the indication of adjuvant therapy, the recurrence risk for primary gastrointestinal stromal tumor (GIST) after surgery needs to be accurately estimated. However, current risk stratification schemes may still have room for improvement. This study seeks to analyze prognostic factors for primary GISTs from 3 aspects, including clinicopathological parameters, immunohistochemical biomarkers, and gene mutational status, and attempts to find novel valuable factors predicting the malignancy potential of GISTs.MethodsRetrospective data from 114 primary GIST patients after R0 resection were collected. Clinicopathological data was obtained from medical records and re-evaluated. Immunohistochemical analysis was performed using the Tissue Microarray method for Ki67, p16, p27, p53, SKP2, CD133, and actin. KIT gene exons 9, 11, 13, and 17 and PDGFR? gene exons 12 and 18 were tested for mutations using PCR.ResultsUnivariate analysis revealed the following factors as poor prognostic indicators for relapse-free survival with a median follow-up of 50 months: male gender, gastrointestinal bleeding, mitotic index >5/50HPFs, tumor size >5 cm, non-gastric site, necrosis, epithelioid or mixed cell type, surrounding tissue invasion, Ki67>5%, p16>20%, p53 index >10, SKP2>10%, and KIT exon 11 deletion. Besides mitotic index, tumor size and site, SKP2 high expression (RR = 2.91, 95% CI: 1.41-5.99, P = 0.004) and KIT exon 11 deletion (RR = 2.73, 95% CI: 1.04-7.16, P = 0.041) were also independent risk factors in multivariate analysis, with gastrointestinal bleeding also showing a trend towards significance (RR = 1.88, 95% CI: 0.98-3.64, P = 0.059). In addition, gastrointestinal bleeding and SKP2 high expression showed a good ability to stratify high-risk patients further.ConclusionOur results show that gastrointestinal bleeding, SKP2 high expression, and KIT exon 11 deletions may be useful indicators of high recurrence risk for primary GIST patients.

Project description:Gastrointestinal stromal tumors (GISTs) are the most common Mesenchymal Neoplasm of the gastrointestinal tract. The tumorigenesis of GISTs has been associated with the gain-of-function mutation and abnormal activation of the stem cell factor receptor (c-KIT) and platelet-derived growth factor receptor alpha (PDGFR?) kinases. Hence, inhibitors that target c-KIT and PDGFR? could be a therapeutic option for the treatment of GISTs. The available approved c-KIT/PDGFR? inhibitors possessed low efficacy with off-target effects, which necessitated the development of potent inhibitors. We performed computational studies of 48 pyrazolopyridine derivatives that showed inhibitory activity against c-KIT and PDGFR? to study the structural properties important for inhibition of both the kinases. The derivative of phenylurea, which has high activities for both c-KIT (pIC50 = 8.6) and PDGFR? (pIC50 = 8.1), was used as the representative compound for the dataset. Molecular docking and molecular dynamics simulation (100 ns) of compound 14 was performed. Compound 14 showed the formation of hydrogen bonding with Cys673, Glu640, and Asp810 in c-KIT, and Cys677, Glu644, and Asp836 in PDGFR?. The results also suggested that Thr670/T674 substitution in c-KIT/PDGFR? induced conformational changes at the binding site of the receptors. Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were developed based on the inhibitors. Contour map analysis showed that electropositive and bulky substituents at the para-position and the meta-position of the benzyl ring of compound 14 was favorable and may increase the inhibitory activity against both c-KIT and PDGFR?. Analysis of the results suggested that having bulky and hydrophobic substituents that extend into the hydrophobic pocket of the binding site increases the activity for both c-KIT and PDGFR?. Based on the contour map analysis, 50 compounds were designed, and the activities were predicted. An evaluation of binding free energy showed that eight of the designed compounds have potential binding affinity with c-KIT/PDGFR?. Absorption, distribution, metabolism, excretion and toxicity (ADMET) and synthetic feasibility tests showed that the designed compounds have reasonable pharmaceutical properties and synthetic feasibility. Further experimental study of the designed compounds is recommended. The structural information from this study could provide useful insight into the future development of c-KIT and PDGFR? inhibitors.

Dataset Information

Molecular characterization of metastatic exon 11 mutant gastrointestinal stromal tumors (GIST) beyond KIT/PDGFR? genotype evaluated by next generation sequencing (NGS).

Publications

Molecular characterization of metastatic exon 11 mutant gastrointestinal stromal tumors (GIST) beyond KIT/PDGFRα genotype evaluated by next generation sequencing (NGS).

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