Polymorphic variation at the BAT-25 and BAT-26 loci in individuals of African origin. Implications for microsatellite instability testing

Abstract

Instability in the repeat size of microsatellite sequences has been described in both hereditary nonpolyposis and sporadic colorectal cancers. Tumors expressing microsatellite instability are identified through the comparison of the repeat sizes at multiple microsatellite loci between tumor and matched normal tissue DNA. The use of a five-marker panel including two mononucleotide repeat microsatellites, BAT-25 and BAT-26, has recently been suggested for the clinical determination of tumor microsatellite instability. The BAT-25 and BAT-26 loci included in this panel have both demonstrated sensitivity to microsatellite instability and normal quasimonomorphic allelic patterns, which has simplified the distinction between normal and unstable alleles. However, in this study, we identified allelic variations in the size of the poly(A) tract at BAT-26 in 12.6% of 103 healthy African-Americans screened. In addition, 18.4% exhibited allelic size variations in the poly(T) tract at BAT-25. Finally, 2.9% showed variant alleles at both BAT-25 and BAT-26 loci. Screening a small population of Nigerians confirmed the polymorphic nature of both loci and the ethnic origin of alleles not identified in other populations studied thus far. Our results dispute the quasimonomorphic nature of both BAT-25 and BAT-26 in all populations and support the need for thorough population studies to define the different allelic profiles and frequencies at microsatellite loci.

Figure 1.

Results from the amplification of the BAT-26 locus in normal male lymphocyte DNA (Lane A), colorectal tumor DNA exhibiting MSI (Lane B), and lymphocyte DNA from five representative healthy African-Americans in three different PCR reactions (Lanes C-G). The banding pattern in Lane A shows the allelic profile of 26 adenine repeats previously described for BAT-26 in normal MSS DNA. Lane B (MSI-positive tumor) illustrates the characteristic banding pattern of MSI seen at BAT-26 in which one band corresponds to alleles with shortened mononucleotide repeats (lower band) compared to a second band demonstrating an allele of wild-type repeat length (upper band). Lane C and E depict the banding pattern seen in African-Americans with one allele of 26 repeats (upper bands) and one polymorphic 16-repeat allele (lower bands). Lanes F and G show the patterns seen in African-Americans with one 26 repeat allele (upper bands) and either a 22-repeat (Lane F) or a 4-repeat (Lane G) lower allele, respectively. Lane D shows the banding pattern from an African-American with both BAT-26 alleles within the previously described allelic profile.

Figure 2.

Results from the amplification of the BAT-25 locus from normal male lymphocyte DNA (Lane A), colorectal tumor DNA exhibiting MSI (Lane B) , and lymphocyte DNA from one representative healthy African-American (Lane C). The banding pattern in Lane A depicts the allelic pattern of 25 thymine repeats previously described for BAT-25 in normal MSS DNA. Lane B (MSI-positive tumor) illustrates the characteristic banding pattern of MSI seen at BAT-25 with one band corresponding to alleles with shortened mononucleotide repeats (lower band) compared to a second band demonstrating an allele of wild-type repeat length (upper band). Lane C depicts the banding pattern seen in blood DNA from one representative African-American with one allele within the previously described range of size (upper band) and one polymorphic 17-repeat allele (lower band).

Figure 3.

Results from the amplification of BAT-25 from the lymphocyte DNA from three Nigerians. The banding patterns seen in Lanes A and B demonstrate the range of size previously characterized for the BAT-25 locus at both alleles. Lane C demonstrates the banding pattern seen in one sample from a Nigerian patient with two polymorphic BAT-25 alleles, both with reduced mononucleotide tracts.