Gain-of-Function Mutations in the Extracellular Domain of KIT Are Common in Canine Mast Cell Tumors

Abstract

In the current study, we examined the types and frequency of KIT mutations in mast cell tumors from 191 dogs. Sequencing of reverse transcription-PCR products revealed alterations in 50 (26.2%) of the dogs. Most mutations were in exon 11 (n = 32), and of these, most were internal tandem duplications (n = 25) between residues 571 and 590. Within exon 11, there were two hotspots for mutations at codons 555-559 and 571-590. In addition, nine dogs had mutations in exon 8 and eight had mutations in exon 9. We selected the two most common mutants and two representative exon 11 mutants for further analysis. When expressed in Ba/F3 cells, they were constitutively tyrosine phosphorylated and induced growth factor -independent cell proliferation. AG1296, a tyrosine kinase inhibitor, dose dependently inhibited both the tyrosine phosphorylation of these mutants and their induction of growth factor -independent proliferation. This study shows that activating mutations in not only exon 11 but also exons 8 and 9 are common in canine mast cell tumors. These results also show that Ba/F3 cells can be used for the direct characterization of canine KIT mutants, eliminating the need to make equivalent mutations in the mouse or human genes.

Concordance of c‐kit Mutational Status in Matched Primary and Metastatic Cutaneous Canine Mast Cell Tumors at Baseline

“…Two already known SNPs were found in exon 8 and 113, 30; furthermore, 3 novel mutations (1 in exon 8 and 2 in exon 11), with unknown clinical relevance, were found.…”

“…Furthermore, 2 already known silent SNPs were detected in exon 8 (1275G > A)3 and in exon 11 (1759C > T) 30. The relative frequencies were 33% (7/21) and 5% (1/21), respectively.…”

“…Exons 8, 9, and 11, considered the hot spot regions for activating protein mutations, were screened by PCR and direct sequencing 3, 22. To amplify either c‐kit exons 8, 9, and 11 (starting from cDNA) or exon 11 (from DNA), previously published primers pairs and PCR conditions were used 23.…”

Pediatric mastocytosis–associated KIT extracellular domain mutations exhibit different functional and signaling properties compared with KIT-phosphotransferase domain mutations

“…11,20,23,24 Ba/F3 cells were transfected with different c-kit constructs as described. 23 TF1 cells were lentivirally transduced as described in supplemental data (available on the Blood Web site; see the Supplemental Materials link at the top of the online article). Both Rat2 and EML cells were transduced using the Phoenix retroviral expression system (ATCC, LGC-Standards).…”

Relação da expressão de fatores de crescimento celular (IGF-1) e (SCF) com fatores prognósticos e o alvo da rapamicina em mamíferos (m-TOR) em mastocitomas cutâneos caninos

“…Estudos indicam que, cerca de 15-40% dos MTCs contém mutações no gene KIT que codifica o receptor tirosina quinase c-KIT (Letard, et al 2008, Welle et al 2008. Seu ligante é o fator de célula tronco (SCF), conhecido também como fator de crescimento para mastócitos que é um fator de crescimento expresso em diversos tipos de tecidos (Linnekin, Jelacic, Shivakrupa 2003, Roskoski 2005.…”