Association between tumor mutation burden (TMB) and MLH1, PMS2, MSH2, and MSH6 alterations in 395 microsatellite instability-high (MSI-High) gastrointestinal (GI) tumors (2018)

Introduction: Correlation between TMB and objective response to checkpoint inhibitors has been shown. MSI-High tumors usually exhibit high TMB. Relationship between the level of TMB and alterations in the four-mismatch repair deficiency (MMR) genes in gastrointestinal tumors has not been comprehensively studied. Methods: MSI-High status was determined by examining altered microsatellite (MS) loci using NextGen sequencing (cutoff: > =46) on a 592-gene panel. TMB was calculated by enumerating somatic missense mutations. MMR protein expression was evaluated by IHC. ANOVA and chi-square tests were used for comparisons. Results: In total, 395 MSI-High gastrointestinal tumors were examined. High TMB (≥ 17 mutations/megabase [mt/MB]) was seen in 89% of the MSI-High tumors. Overall, MSI-High CRC exhibited the highest TMB compared to “all other MSI-High gastrointestinal cancers” (39 vs. 27 mt/MB, respectively, p < 0.0001). Left-sided MSI-High tumors had a higher TMB compared to right-sided tumors (51 vs. 35 mt/MB, p = 0.01). No significant differences were observed in TMB between BRAF V600 mutant and wild type MSI-High CRC (38.7 vs. 39 mt/MB). Significant association between the microsatellite loci and TMB (Rho=0.60, p < 0.0001) was observed. The mean TMB among MSI-High gastrointestinal tumors were as follow: 39 (mt/MB) CRC, 33 (mt/MB) gastric adenocarcinomas, 31 (mt/MB) small bowel adenocarcinomas, and 29 (mt/MB) in esophageal, GEJ, and pancreatic cancers, and cholangiocarcinoma. The most frequently altered (IHC loss or mutation) MMR genes in the MSI-High gastrointestinal tumors were MLH1 and PMS2. In CRC, MSH2 was more frequently altered in left-sided than right-sided tumors (45% vs. 12%), as was MSH6 (67% vs. 40%), p = 0.01. In fact MLH1, PMS2, MSH2 and MSH6 gene alterations varied significantly across MSI-H gastrointestinal tumors: CRC 72%, 83%, 21% and 49%; left-sided CRC, 47%, 55%, 45%, 67%; right-sided CRC, 80%, 91%, 12%, 40%; gastric adenocarcinomas, 80%, 86%, 13% and 35%; esophageal and GEJ adenocarcinomas, 100%, 100%, 0% and 13%; small bowel adenocarcinomas, 79%, 75%, 29% and 43%; pancreatic adenocarcinomas, 50%, 60%, 50%, 42%; cholangiocarcinoma, 83%, 80%, 33% and 60%; respectively. Generally, MSH2 or MSH6 alterations were associated with higher TMB (49 and 44 mt/MB, respectively) than MLH1 or PMS2 (33 mt/MB for both), P < 0.0001. Furthermore, tumors with a MSH2/MSH6 co-alteration had a significantly higher TMB compared to those with a MLH1/PMS2 co-alteration (51 mt/MB vs. 31, p < 0.0001). However, no significant difference was observed in the level of TMB among MSI-High tumors that exhibited a MSH2/MSH6 co-alteration (52 mt/MB), a MSH2 alteration (51 mt/MB), or a MSH6 (48 mt/MB), p = 0.95. Conclusion: Among MSI-High gastrointestinal cancers, CRC exhibited the highest TMB level, and left-sided tumors exhibited higher TMB than right-sided tumors. MSH2 and/or MSH6 alterations were associated with a significantly higher TMB than MLH1/PMS2 alterations across all gastrointestinal cancer types. Additional analysis to assess the correlation between specific MMR gene alterations and response to checkpoint inhibitors is underway.