PMID- 25865047 OWN - NLM STAT- MEDLINE DCOM- 20151019 LR - 20220316 IS - 1528-0012 (Electronic) IS - 0016-5085 (Print) IS - 0016-5085 (Linking) VI - 149 IP - 2 DP - 2015 Aug TI - Platelet-Derived Growth Factor Receptor-alpha Regulates Proliferation of Gastrointestinal Stromal Tumor Cells With Mutations in KIT by Stabilizing ETV1. PG - 420-32.e16 LID - S0016-5085(15)00497-7 [pii] LID - 10.1053/j.gastro.2015.04.006 [doi] AB - BACKGROUND & AIMS: In gastrointestinal muscles, v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) is predominantly expressed by interstitial cells of Cajal (ICC) and platelet-derived growth factor receptor-alpha (PDGFRA) polypeptide is expressed by so-called fibroblast-like cells. KIT and PDGFRA have been reported to be coexpressed in ICC precursors and gastrointestinal stromal tumors (GISTs), which originate from the ICC lineage. PDGFRA signaling has been proposed to stimulate growth of GISTs that express mutant KIT, but the effects and mechanisms of selective blockade of PDGFRA are unclear. We investigated whether inhibiting PDGFRA could reduce proliferation of GIST cells with mutant KIT via effects on the KIT-dependent transcription factor ETV1. METHODS: We studied 53 gastric, small intestinal, rectal, or abdominal GISTs collected immediately after surgery or archived as fixed blocks at the Mayo Clinic and University of California, San Diego. In human GIST cells carrying imatinib-sensitive and imatinib-resistant mutations in KIT, PDGFRA was reduced by RNA interference (knockdown) or inhibited with crenolanib besylate (a selective inhibitor of PDGFRA and PDGFRB). Mouse ICC precursors were retrovirally transduced to overexpress wild-type Kit. Cell proliferation was analyzed by methyltetrazolium, 5-ethynyl-2'-deoxyuridine incorporation, and Ki-67 immunofluorescence assays; we also analyzed growth of xenograft tumors in mice. Gastric ICC and ICC precursors, and their PDGFRA(+) subsets, were analyzed by flow cytometry and immunohistochemistry in wild-type, Kit(+/copGFP), Pdgfra(+/eGFP), and NOD/ShiLtJ mice. Immunoblots were used to quantify protein expression and phosphorylation. RESULTS: KIT and PDGFRA were coexpressed in 3%-5% of mouse ICC, 35%-44% of ICC precursors, and most human GIST samples and cell lines. PDGFRA knockdown or inhibition with crenolanib efficiently reduced proliferation of imatinib-sensitive and imatinib-resistant KIT(+)ETV1(+)PDGFRA(+) GIST cells (50% maximal inhibitory concentration = 5-32 nM), but not of cells lacking KIT, ETV1, or PDGFRA (50% maximal inhibitory concentration >230 nM). Crenolanib inhibited phosphorylation of PDGFRA and PDGFRB, but not KIT. However, Kit overexpression sensitized mouse ICC precursors to crenolanib. ETV1 knockdown reduced KIT expression and GIST proliferation. Crenolanib down-regulated ETV1 by inhibiting extracellular-signal-regulated kinase (ERK)-dependent stabilization of ETV1 protein and also reduced expression of KIT and PDGFRA. CONCLUSIONS: In KIT-mutant GIST, inhibition of PDGFRA disrupts a KIT-ERK-ETV1-KIT signaling loop by inhibiting ERK activation. The PDGFRA inhibitor crenolanib might be used to treat patients with imatinib-resistant, KIT-mutant GIST. CI - Copyright (c) 2015 AGA Institute. Published by Elsevier Inc. All rights reserved. FAU - Hayashi, Yujiro AU - Hayashi Y AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota. FAU - Bardsley, Michael R AU - Bardsley MR AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota. FAU - Toyomasu, Yoshitaka AU - Toyomasu Y AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota. FAU - Milosavljevic, Srdjan AU - Milosavljevic S AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota. FAU - Gajdos, Gabriella B AU - Gajdos GB AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota. FAU - Choi, Kyoung Moo AU - Choi KM AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota. FAU - Reid-Lombardo, K Marie AU - Reid-Lombardo KM AD - Department of Surgery, Mayo Clinic, Rochester, Minnesota. FAU - Kendrick, Michael L AU - Kendrick ML AD - Department of Surgery, Mayo Clinic, Rochester, Minnesota. FAU - Bingener-Casey, Juliane AU - Bingener-Casey J AD - Department of Surgery, Mayo Clinic, Rochester, Minnesota. FAU - Tang, Chih-Min AU - Tang CM AD - Division of Surgical Oncology, Moores Cancer Center, University of California, San Diego, California. FAU - Sicklick, Jason K AU - Sicklick JK AD - Division of Surgical Oncology, Moores Cancer Center, University of California, San Diego, California. FAU - Gibbons, Simon J AU - Gibbons SJ AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota. FAU - Farrugia, Gianrico AU - Farrugia G AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Center for Individualized Medicine, Mayo Clinic, Rochester, Minnesota. FAU - Taguchi, Takahiro AU - Taguchi T AD - Division of Human Health and Medical Science, Graduate School of Kuroshio Science, Kochi University, Kochi, Japan. FAU - Gupta, Anu AU - Gupta A AD - Departments of Pathology and Molecular Genetics, Lerner Research Institute and Taussig Cancer Center, Cleveland Clinic, Cleveland, Ohio. FAU - Rubin, Brian P AU - Rubin BP AD - Departments of Pathology and Molecular Genetics, Lerner Research Institute and Taussig Cancer Center, Cleveland Clinic, Cleveland, Ohio. FAU - Fletcher, Jonathan A AU - Fletcher JA AD - Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts. FAU - Ramachandran, Abhijit AU - Ramachandran A AD - AROG Pharmaceuticals, LLC, Dallas, Texas. FAU - Ordog, Tamas AU - Ordog T AD - Enteric Neuroscience Program, Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, Minnesota; Gastroenterology Research Unit, Mayo Clinic, Rochester, Minnesota; Center for Individualized Medicine, Mayo Clinic, Rochester, Minnesota. Electronic address: ordog.tamas@mayo.edu. LA - eng GR - R01DK058185/DK/NIDDK NIH HHS/United States GR - R01 DK057061/DK/NIDDK NIH HHS/United States GR - R01DK057061/DK/NIDDK NIH HHS/United States GR - P30CA015083/CA/NCI NIH HHS/United States GR - P01 DK068055/DK/NIDDK NIH HHS/United States GR - U54 CA168512/CA/NCI NIH HHS/United States GR - P01DK068055/DK/NIDDK NIH HHS/United States GR - R01 DK058185/DK/NIDDK NIH HHS/United States GR - P50CA127003/CA/NCI NIH HHS/United States GR - K08 CA168999/CA/NCI NIH HHS/United States GR - U54CA168512/CA/NCI NIH HHS/United States GR - P30DK084567/DK/NIDDK NIH HHS/United States GR - P30 CA015083/CA/NCI NIH HHS/United States GR - K08CA168999/CA/NCI NIH HHS/United States GR - P30 DK084567/DK/NIDDK NIH HHS/United States GR - P50 CA127003/CA/NCI NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, Non-U.S. Gov't DEP - 20150409 PL - United States TA - Gastroenterology JT - Gastroenterology JID - 0374630 RN - 0 (Benzamides) RN - 0 (Benzimidazoles) RN - 0 (Biomarkers, Tumor) RN - 0 (DNA-Binding Proteins) RN - 0 (Etv1 protein, mouse) RN - 0 (Nucleic Acid Precursors) RN - 0 (Piperazines) RN - 0 (Piperidines) RN - 0 (Pyrimidines) RN - 0 (Transcription Factors) RN - 8A1O1M485B (Imatinib Mesylate) RN - EC 2.7.10.1 (Proto-Oncogene Proteins c-kit) RN - EC 2.7.10.1 (Receptor, Platelet-Derived Growth Factor alpha) RN - EC 2.7.10.1 (Receptor, Platelet-Derived Growth Factor beta) RN - LQF7I567TQ (crenolanib) SB - IM MH - Animals MH - Benzamides/metabolism MH - Benzimidazoles/metabolism MH - Biomarkers, Tumor/metabolism MH - Cell Line, Tumor MH - Cell Proliferation/*genetics MH - DNA-Binding Proteins/*genetics MH - Flow Cytometry MH - Gastrointestinal Stromal Tumors/genetics/*metabolism MH - Gene Knockdown Techniques/methods MH - Humans MH - Imatinib Mesylate MH - Immunohistochemistry MH - Mice MH - Mice, Inbred BALB C MH - Mutation MH - Nucleic Acid Precursors/genetics MH - Phosphorylation/genetics MH - Piperazines/metabolism MH - Piperidines/metabolism MH - Proto-Oncogene Proteins c-kit/genetics/*metabolism MH - Pyrimidines/metabolism MH - Receptor, Platelet-Derived Growth Factor alpha/genetics/*metabolism MH - Receptor, Platelet-Derived Growth Factor beta/metabolism MH - Signal Transduction/*genetics MH - Transcription Factors/*genetics PMC - PMC4516576 MID - NIHMS679747 OTO - NOTNLM OT - Cancer OT - Receptor Tyrosine Kinase OT - Signal Transduction OT - Stem Cells EDAT- 2015/04/14 06:00 MHDA- 2015/10/20 06:00 PMCR- 2016/08/01 CRDT- 2015/04/14 06:00 PHST- 2014/08/13 00:00 [received] PHST- 2015/04/02 00:00 [revised] PHST- 2015/04/06 00:00 [accepted] PHST- 2015/04/14 06:00 [entrez] PHST- 2015/04/14 06:00 [pubmed] PHST- 2015/10/20 06:00 [medline] PHST- 2016/08/01 00:00 [pmc-release] AID - S0016-5085(15)00497-7 [pii] AID - 10.1053/j.gastro.2015.04.006 [doi] PST - ppublish SO - Gastroenterology. 2015 Aug;149(2):420-32.e16. doi: 10.1053/j.gastro.2015.04.006. Epub 2015 Apr 9.