title: KIT Signaling Promotes Growth of Colon Xenograft Tumors in Mice and Is Up-Regulated in a Subset of Human Colon Cancers.
abstract: OBJECTIVE
  Receptor tyrosine kinase (RTK) inhibitors have advanced colon cancer treatment. We investigated the role of the RTK KIT in development of human colon cancer.
  
  
  METHODS
  An array of 137 patient-derived colon tumors and their associated xenografts were analyzed by immunohistochemistry to measure levels of KIT and its ligand KITLG. KIT and/or KITLG was stably knocked down by expression of small hairpin RNAs from lentiviral vectors in DLD1, HT29, LS174T, and COLO320 DM colon cancer cell lines, and in UM-COLON#8 and POP77 xenografts; cells transduced with only vector were used as controls. Cells were analyzed by real-time quantitative reverse transcription polymerase chain reaction, single-cell gene expression analysis, flow cytometry, and immunohistochemical, immunoblot, and functional assays. Xenograft tumors were grown from control and KIT-knockdown DLD1 and UM-COLON#8 cells in immunocompromised mice and compared. Some mice were given the RTK inhibitor imatinib after injection of cancer cells; tumor growth was measured based on bioluminescence. We assessed tumorigenicity using limiting dilution analysis.
  
  
  RESULTS
  KIT and KITLG were expressed heterogeneously by a subset of human colon tumors. Knockdown of KIT decreased proliferation of colon cancer cell lines and growth of xenograft tumors in mice compared with control cells. KIT knockdown cells had increased expression of enterocyte markers, decreased expression of cycling genes, and, unexpectedly, increased expression of LGR5 associated genes. No activating mutations in KIT were detected in DLD1, POP77, or UM-COLON#8 cells. However, KITLG-knockdown DLD1 cells formed smaller xenograft tumors than control cells. Gene expression analysis of single CD44(+) cells indicated that KIT can promote growth via KITLG autocrine and/or paracrine signaling. Imatinib inhibited growth of KIT(+) colon cancer organoids in culture and growth of xenograft tumors in mice. Cancer cells with endogenous KIT expression were more tumorigenic in mice.
  
  
  CONCLUSIONS
  KIT and KITLG are expressed by a subset of human colon tumors. KIT signaling promotes growth of colon cancer cells and organoids in culture and xenograft tumors in mice via its ligand, KITLG, in an autocrine or paracrine manner. Patients with KIT-expressing colon tumors can benefit from KIT RTK inhibitors.
authors.affiliation: Stanford School of Medicine, Stanford, California.
authors.forename: Evan C
authors.initials: EC
authors.lastname: Chen
authors.affiliation: Division of Gastroenterology and Hepatology, Stanford School of Medicine, Stanford, California.
authors.forename: Taylor A
authors.initials: TA
authors.lastname: Karl
authors.affiliation: Department of Bioengineering, Bar-Ilan University, Ramat Gan, Israel.
authors.forename: Tomer
authors.initials: T
authors.lastname: Kalisky
authors.affiliation: Department of Pathology, Stanford School of Medicine, California.
authors.forename: Santosh K
authors.initials: SK
authors.lastname: Gupta
authors.affiliation: Division of General Surgery, University of Toronto, Toronto, Canada.
authors.forename: Catherine A
authors.initials: CA
authors.lastname: O'Brien
authors.affiliation: Department of Pathology, Stanford School of Medicine, California.
authors.forename: Teri A
authors.initials: TA
authors.lastname: Longacre
authors.affiliation: Department of Pathology, Stanford School of Medicine, California.
authors.forename: Matt
authors.initials: M
authors.lastname: van de Rijn
authors.affiliation: Department of Bioengineering, Stanford School of Medicine, Stanford, California; Howard Hughes Medical Institute, Chevy Chase, Maryland.
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Quake
authors.affiliation: Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford, California.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Clarke
authors.affiliation: Division of Gastroenterology and Hepatology, Stanford School of Medicine, Stanford, California; Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford, California. Electronic address: rmike@stanford.edu.
authors.forename: Michael E
authors.initials: ME
authors.lastname: Rothenberg
mesh_terms: D000818:Animals; D019898:Autocrine Communication; D018938:Caco-2 Cells; D049109:Cell Proliferation / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D019899:Paracrine Communication; D047428:Protein Kinase Inhibitors / Q000627:therapeutic use; D019009:Proto-Oncogene Proteins c-kit / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D015398:Signal Transduction / Q000187:drug effects; D019089:Stem Cell Factor / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D047368:Tumor Burden; D015854:Up-Regulation; D023041:Xenograft Model Antitumor Assays
chemical_list: D047428:Protein Kinase Inhibitors; D019089:Stem Cell Factor; D019009:Proto-Oncogene Proteins c-kit
keywords: CD117; Colorectal Cancer; Enteroids; Stem Cell Factor