Molecular prediction of therapy with selective analysis of microsatellites in neoadjuvant therapy of rectal cancer

Introduction: Neoadjuvant therapy regimens are increasing popular for the therapy of locally advanced cancer, however until now, only a certain percentage of patients usually benefit from applied therapeutic regimens. Therefore the characterization of specific molecular markers for response prediction is highly desirable. In this study we analysed certain chromosomal regions for loss of heterozygosity (LOH) in rectal cancer of patients treated with neoadjuvant radio-chemotherapy for an association with therapy response. Materials and methods: We analysed locally advanced rectal carcinomas (cT3/4 NX MO) of 47 patients treated with radio-chemotherapy based on the "Münchener Protokoll" (45 Gy, 1.8 Gy daily, 300 mg/m2/d continously). Only patients who received 100% of the radiation and at least 50% of the planned chemotherapy were included in the study. Response was determined by histopathological examination of the resected specimen. Patients with less than 10% of vital tumor cells in the specimens were classified as responder ( 18/47; 38%) and patients with 10% or more of vital tumor cells in the specimen as non-responder. (29/47; 62%). DNA was isolated from tumor and non tumerous tissue from pretherapeutic biopsies after microdissection. The following 8 microsatellite markers which are known to be involved in the carcinogenesis of colorectal carcinomas (gene loci in parenthesis) were amplified by PCR: D5S346 (APC), TP53 (p53), D18S58 (DCC), D2S123 (hMSH2, hMSH6), D17S250 (17q21), D7S1824 (7q34-35), BAT25 (4ql2), BAT26 (2pl6). An allele peak ratio of <0.6, representing an allelic signal reduction of 40% was considered as LOH. The FAL ratio (fractional allelic loss) was calculated by dividing the number of markers showing LOH by the number of informative markers for each tumor. The FAL ratio was divided into three groups: tumors with low FAL (0,00-0,25), medium FAL (0,26-0,50) and high FAL (0,51-1,00). The tumors were also analysed for microsatellite instability (MSI). Results: LOH at TP53 was observed in 78% of informative tumors. LOH at the other chromosomal markers was in the range of 34 to 47%. No significant differences between responder and non-responder were found for any marker. In addition the FAL ratio (11 tumors with low FAL, 22 with medium and 14 with high FAL) did not show significant differences between responder and non-responder. None of the tumors exhibited microsatellite instability. Conclusion: LOH and FAL in rectal carcinomas of patients treated with neoadjuvant radio-chemotherapy did not show an association with therapy response. This is in contrast to our results of microsatellite analysis in gastric carcinomas of patients, treated with neoadjuvant chemotherapy based on cisplatinum and 5-FU. (Gründe! et al., Clin. Cancer Research, 2000). Evaluation of the data in respect to the local recurrence rate and the overall survival of the patients will be done at a two and five year follow-up.