Purpose To analyze the genetic variability in a variable number of tandem repeats (VNTR) in the thymidylate synthase (TS) enhancer promoter region and assess the influence of functional alterations in mismatch repair genes by analyzing constitutional and tumoral DNA from patients with colorectal adenocarcinoma with a high microsatellite instability (MSI-H) or microsatellite stability (MSS) status. Patients and methods Patients who underwent surgery for colorectal adenocarcinoma were selected from the colorectal database of our institute and, on the basis of MSI status, assigned to a study group and a control group: group A, MSI-H; group B, MSS. Microsatellite status was investigated using the Bethesda recommended panel (BAT-26, BAT-25, D2S123, D5S346, D17S250). In MSI-H patients an additional analysis was made of the microsatellite loci D18S61 and D18S58, both mapping in the region containing the TS gene (18p11.2–11.32). Based on the number of altered microsatellites (≥2, 1, or 0), tumors were considered as having high (MSI-H) or low (MSI-L) instability or microsatellite stability (MSS), respectively. Genotyping for thymidylate synthase promoter polymorphism was carried out on constitutional and tumor DNA of each patient by PCR amplification of the polymorphic region. Results MSI-H was found in 55 patients (group A) and MSS in 50 patients (group B). In none of the MSI-H patients was microsatellite instability found in the additional D18S61 and D18S58 loci. In five group A and ten group B cases the analysis was not performed because constitutional DNA and/or tumoral DNA were not amplifiable. Homozygotes for the triple repeat variant (3R/3R) displayed only the large PCR product, homozygotes for the double repeat variant (2R/2R) displayed only the smaller PCR product, while heterozygotes (2R/3R) displayed both the larger and smaller PCR products. In 3/50 (6%) group A patients and 5/40 (12%) group B patients repeat variations were found in tumoral DNA. Conclusion Our findings demonstrate that there is genetic homogeneity between constitutional and tumoral DNA but do not support the hypothesis that mismatch repair genes are involved in VNTR recombinant events in TS gene variability.
A Novel Nomenclature for Repeat Motifs in the Thymidylate Synthase Enhancer Region and Its Relevance for Pharmacogenetic Studies