We assume that besides mismatch repair (MMR) genes, next generation sequencing (NGS) of MMR associated genes could improve detection of driver mutattors and clarify the somatic mutation patterns of subtypes CRC classfied by EMAST and MSI-high.
Extracted from a 1505 CRC cases database,81 cases with MSI-high and EMAST+,78 cases with EMAST+ and MSS, and 72 cases with MSI-high but EMAST- were identified. The tumor and WBC DNA were applied and got from Biobank of Taipei-Veteran General Hospital afer approval of IRB. The germline and somatic mutations were analyzied with 16- genes NGS (illumina HiSeq2500 system).
Totally 284 pathological germline mutations were identified in 161 patients with MSI-high or EMAST+. The most common gene mutations were EPCAM (17.3%), MSH6 (16.9%), followed by MLH1 (15.2%), and AXIN2(15.2%). Majority of EMAST and MSI resulted from not only MMR dysfunction, but also germline mutations of AXIN2, POLD1 and TGFBR2. After deduction of 284 germline mutations, there were 1,090 somatic mutations in 161 cases with germline mutations, 445 mutations in 70 cases without germline mutations. Tumors with EMAST+ and MSI-high had signifcantly higher mutation number than those of tumors with only EMAST+ or only MSI-high. Besides germline mutations of AXIN2, germline mutations of other genes were similar. With AXIN2 germline mutations, somatic mutation rate was 187.7 ± 97.8 mut/MB significantly higher than those of without germline mutations (137.8± 84.5 mut/MB p = 0.002). Besides five major MMR genes, Eleven Axin2, eight POLE and six TGFbR2 germline mutations resulted in following MSI-high or EMAST (+) genotype without other accompany germline mutation. Clinically, patients with germline mutation had significantly higher frequency of proximal tumor location and early stage disease.
Our result showed that NGS could enhance detection of familial CRC. Somatic mutation burden might be through MSI or EMAST but not only germline mutation genes themselves. Several genes with germline mutations could explain origin of the familiar CRC. AXIN2 gene deserved to do futher experiment to confirm its role in WNT pathway and as a hypermutator.
Clinical trial identification
Legal entity responsible for the study
The Institutional Review Board of Taipei Veterans General Hospital (number 2017-06-004BC).
Taipei-Veterans General Hospital, Taiwan.
All authors have declared no conflicts of interest.