Functional polymorphisms in the promoter regions of MMP2 and MMP3 are not associated with melanoma progression

Abstract

Background: The matrix metalloproteinases (MMPs) are enzymes that cleave various components of the extracellular matrix (ECM) and basement membranes. MMPs are expressed in melanocytes and their overexpression has been linked to tumor development, progression and metastasis. At the genetic level, the following functional promoter polymorphisms are known to modify the gene transcription: -1306 C/T and -735 C/T in the MMP2 gene, and -1171 5A/6A in the MMP3 gene. Functional polymorphisms in MMP genes' promoter regions may modulate the risk for melanoma progression.

Methods: We evaluated MMP2 and MMP3 germline polymorphisms in a group of 1002 melanoma patients using PCR-based methods, including fragment size analysis and melting temperature profiles. Two-sided Chi-Square, Cochran-Armitage tests for trend, Fisher's exact tests, and Kendall's Tau tests were performed to evaluate the associations between genotype and various clinical and epidemiologic factors. Multivariate analyses were conducted using logistic regression, adjusting for known melanoma confounders such as age, sex, phenotypic index, moles, freckles, and race. Survival estimates were computed using the Kaplan-Meier method and differences in survival were assessed using the log rank test.

Results: All genotypes were in Hardy-Weinberg equilibrium. After adjustment for age, sex and phenotypic characteristics of melanoma risk, no significant associations were identified with the clinical, pathological, and epidemiological variables studied. The melting profile for MMP2 -735 C/T identified a new change in one sample. A new PCR-amplification followed by direct sequencing confirmed a heterozygote G to A substitution at position -729.

Conclusion: This study does not provide strong evidence for further investigation into the role of the MMP2 and MMP3 variants in melanoma progression.

Figure 1

Genotyping of MMP2 -735 C/T SNP by melting temperature analysis. (A) The top panels depict the derivative melting curve plots obtained for (from left to right): MMP2 -735CC, MMP2 -735CT, MMP2 -735TT, and an unexpected profile (UKN) showing peaks between 62 and 66°C (B) The bottom panel depicts the wild type (left) and a new variant (right) at position -729 near the target SNP (white arrow). Reverse sequencing revealed a novel mutation corresponding to a G to A change at position -729 in the sense strand (black arrow).