Evaluating markers of microsatellite instability in an ethnically diverse patient cohort

PR-7

Background: African Americans (AAs) are affected by colorectal cancer (CRC) at disproportionate levels with both higher incidence and mortality rates than other ethnicities. Reasons for disparities not only include socioeconomic status, lifestyle and screening rates, but possibly tumor biology/genetics. MLH1, MSH2, MSH6 and PMS2 are proteins expressed by mismatch repair (MMR) genes that are responsible for repairing nucleotide mispairs and small insertions or deletions caused by the DNA replication machinery. Loss of expression and function of these proteins are associated with the Lynch syndrome or hereditary nonpolyposis colorectal cancer (HNPCC). Many characteristics of AA CRCs resemble those of HNPCC patients, such as younger age at presentation, a higher incidence of proximal-located tumors, and a higher prevalence of the microsatellite instability (MSI) - high tumor phenotype. The purpose of this study is to determine if correlations exist between MMR protein expression and MSI in CRC tumors, using immunohistochemical (IHC) and MSI analyses, and further if either of these events correlates with race/ethnicity.
 Design: Paraffin-embedded sections of 952 nonconsecutive lesions from 434 CRC patients were constructed into tissue microarrays and then immunostained with MLH1 mouse monoclonal antibody (BD Pharmingen, 1:100), MSH2 mouse monoclonal antibody (Oncogene Research Products, 1:100), MSH6 mouse monoclonal antibody (BD Transduction Laboratories, 1:25) and PMS2 mouse monoclonal antibody (BD Pharmigen, 1:500). The patient sections examined consisted of normal colonic epithelium, adenoma, invasive carcinoma and metastatic CRC located in the lymph node or liver. The MSI molecular study was conducted anonymously. DNA was extracted from 50-micron sections of normal and tumor tissue, then genotyped using standard microsatellite markers BAT25, BAT26 and BAT40 to determine MSI status. Statistical analysis was performed using Stata 9.2.
 Results: Our study sample consisted of 50% AAs, 46% Caucasians, 2% Hispanics, 1% other ethnicities and 53% females. The mean age at diagnosis was 68.5 years old and the median survival time was 6.4 years. The majority (62%) of the tumors was moderately differentiated and the mean tumor size was 4.7 cm. Loss of MLH1, MSH2, MSH6 and PMS2 protein expression was observed in 23% of our ethnically diverse cohort, with no significant differences between AAs and Caucasians. An association between MMR-deficiency and female gender was observed in Caucasians but not among AAs. Adjusted for age, grade, and stage, AAs had a nearly 40% increase in the hazard of death compared to Caucasians. This finding appeared to be isolated to those with MMR-proficient tumors. Of the 20 patient samples with loss of MLH1, MSH2, MSH6 or PMS2 protein expression tested to date, 78% were microsatellite unstable tumors.
 Conclusions: Loss of expression of MLH1, MSH2, MSH6 and PMS2 was observed in 23% of our ethnically diverse cohort. The interaction between treatment, genetic susceptibility, biomarkers of risk and race/ethnicity will be evaluated in this cohort as a means to understanding the disparities that exist in colorectal cancer outcomes.