Jornal de Patologia Clínica e Experimental

Abstrato

Mutation Analysis of Gastrointestinal Stromal Tumors in a Pathology Laboratory with 42 Cases of Formalin-Fixed Paraffin-Embedded Specimens

Nilufar Lokman, Yoshimi Suzuki, Hiroshi Kawachi, Masaki Sekine, Asuka Furukawa, Keisuke Uchida, Yukichi Hara, Hideki Ishizu, Hisataka Uchima, Takumi Akashi and Yoshinobu Eishi

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).