Abstract
Gastrointestinal stromal tumors (GISTs) are frequently driven by auto-activated, mutant KIT and have durable response to KIT tyrosine kinase inhibitor. However, acquired resistance is an increasing clinical issue in GIST patients receiving front-line imatinib therapy. Our previous studies showed the colocalization of KIT with DAPI-stained nuclei in GIST cells without knowing the role of nuclear KIT in GIST tumorigenesis. In this article, we first identified the binding of nuclear KIT to the promoter of NFKB inhibitor beta (NFKBIB) by chromatin immunoprecipitation (ChIP) sequencing and ChIP assays, which was accompanied with enhanced NFKBIB protein expression in GIST cells. Clinically, high NCCN risk GISTs had significantly higher mean expression levels of nuclear phospho-KIT and NFKBIB as compared with those of intermediate or low/very low-risk GISTs. Conversely, downregulation of NFKBIB by siRNA led to RELA nuclear translocation that could bind to the KIT promoter region and subsequently reduced KIT transcription/expression and the viability of GIST cells. These findings were further confirmed by either RELA overexpression or NFKB/RELA inducer, valproic acid, treatment to result in reduced KIT expression and relative cell viability of imatinib-resistant GIST cells. Combining valproic acid with imatinib showed significantly better growth inhibitory effects on imatinib-resistant GIST48 and GIST430 cells in vitro, and in the GIST430 animal xenograft model. Taken together, these results demonstrate the existence of a nuclear KIT-driven NFKBIB-RELA-KIT autoregulatory loop in GIST tumorigenesis, which are potential targets for developing combination therapy to overcome imatinib-resistant of KIT-expressing GISTs.
Highlights
Gastrointestinal stromal tumors (GISTs) are the most common type of mesenchymal neoplasm of the gastrointestinal tract [1, 2]
Using chromatin immunoprecipitation sequencing (ChIP-seq) and ChIP assays, we found that nuclear KIT could bind to the NFKB inhibitor beta (NFKBIB) promoter region and regulate NFKBIB expression
The phosphorylation levels of wild-type (WT) KIT induced by its ligand stem cell factor (SCF) for 30 and 60 min were correlated with the KIT expression levels in the nucleus
Summary
Gastrointestinal stromal tumors (GISTs) are the most common type of mesenchymal neoplasm of the gastrointestinal tract [1, 2]. Develop treatment for patients with TKI-resistant, KITexpressing GISTs. Increasing evidence has shown that receptor tyrosine kinases, such as epidermal growth factor receptor (EGFR) and insulin-like growth factor 1 receptor (IGF1R), can translocate into the nucleus and mediate gene expression, resulting in tumorigenesis and drug resistance. We found that KIT colocalized with DAPI-stained nuclei in IM-resistant, mutant KITexpressing GIST48 and GIST430 cells [12, 13]. It is unknown whether KIT can locate in the nucleus. Our results help elucidate the role of nuclear KIT and provide potential therapeutic targets for IM-resistant, KIT-expressing GISTs
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