Mutation Analysis of Gastrointestinal Stromal Tumors in a Pathology Laboratory with 42 Cases of Formalin-Fixed Paraffin-Embedded SpecimensNilufar Lokman1, Yoshimi Suzuki1, Hiroshi Kawachi1, Masaki Sekine2, Asuka Furukawa1, Keisuke Uchida1, Yukichi Hara1, Hideki Ishizu3, Hisataka Uchima4, Takumi Akashi2 and Yoshinobu Eishi1,2*
- *Corresponding Author:
- Yoshinobu Eishi
Department of Human Pathology, Director
Division of Surgical Pathology
Tokyo Medical and Dental University, 1-5-45
Yushima, Bunkyo-ku, Tokyo, 113-8510, Japan
E-mail: [email protected]
Received Date: July 17, 2014; Accepted Date: August 14, 2014; Published Date: August 16, 2014
Citation: Lokman N, Suzuki Y, Kawachi H, Sekine M, Furukawa A, et al. (2014) Mutation Analysis of Gastrointestinal Stromal Tumors in a Pathology Laboratory with 42 Cases of Formalin-Fixed Paraffin-Embedded Specimens. J Clin Exp Pathol 4:185. doi: 10.4172/2161-0681.1000185
Copyright: © 2014 Lokman N, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Background: This study aimed to establish a standard gene analysis procedure for successful mutation analysis in pathology laboratories using formalin-fixed paraffin-embedded gastrointestinal stromal tumor (GIST) specimens.
Methods: Twenty-six cases of GIST were retrospectively collected and subjected to polymerase chain reaction (PCR) for exons 9, 11, 13, and 17 of KIT and exons 12 and 18 of PDGFRA genes, comparing four groups of previously reported primer sets with one group of novel primer sets. Amplified DNA was directly sequenced with or without subsequent subcloning. The standardized procedure established in the retrospective study was used to prospectively analyze 16 additional cases of GIST.
Results: The novel primer sets provided the highest percentages (92%-96%) of successful amplification of all the exons, except for KIT exon 9. In total, 15 double-band samples on electrophoresis after PCR for KIT exon 11 carried a deletion- or insertion-type mutation. Nine single-band samples presented superimposed consecutive double peaks on direct sequencing, and subcloning confirmed a deletion- or insertion-type mutation. Fourteen single-band samples carried a point mutation that presented single base-pair double peaks on direct sequencing. Six single-band samples carried no mutation in any of the exons. In the prospective study, we found KIT-negative GISTs, simultaneous mutations of both KIT and PDGFRA genes, and phenotypic and genotypic changes in pre- and post-imanitib treated GIST lesions.
Conclusions: The validity of the standardized procedure was confirmed in the prospective study. This standardized procedure can make GIST mutation analysis more readily available to pathology laboratories. (243 words).