Background Soluble mesothelin-related peptide (SMRP) is a promising diagnostic biomarker for malignant pleural mesothelioma (MPM), but various confounders hinder its usefulness in surveillance programmes. We previously showed that a single nucleotide polymorphism (SNP) within the 3’untranslated region (3’UTR) of the mesothelin (MSLN) gene could affect the levels of SMRP.
Objectives To focus on SNPs located within MSLN promoter as possible critical genetic variables in determining SMRP levels.
Methods The association between SMRP and SNPs was tested in 689 non-MPM subjects and 70 patients with MPM. Reporter plasmids carrying the four most common haplotypes were compared in a dual luciferase assay, and in silico analyses were performed to investigate the putative biological role of the SNPs.
Results We found a strong association between serum SMRP and variant alleles of rs3764247, rs3764246 (in strong linkage disequilibrium with rs2235504) and rs2235503 in non-MPM subjects. Inclusion of the genotype information led to an increase in SMRP specificity from 79.9% to 85.5%. Although not statistically significant, the group with MPM showed the same trend of association. According to the in vitro luciferase study, rs3764247 itself had a functional role. In silico approaches showed that the binding sites for transcription factors such as Staf and ZNF143 could be affected by this SNP. The other SNPs were shown to interact with each other in a more complex way.
Conclusions These data support the suggestion that SMRP performance is affected by individual (ie, genetic) variables and that MSLN expression is influenced by SNPs located within the promoter regulatory region.
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Contributors The manuscript was made possible thanks to the following contributions. SL, FG, AC and LM conceived the idea at the basis of the study. AB, RF, and AC from the Occupational Medicine of Cisanello Hospital in Pisa contributed with essential biological samples and the related clinical features. CDS, MC, EB, EP, AC, IL, ID and LP carried out the DNA extractions for the genotyping. CDS, MC and SL performed the statistical analyses on the association between genotypes and SMRP levels. CDS and PP obtained the plasmids via cloning and mutagenesis and performed the in vitro luciferase assay on Met-5A cells, with the contribution of OM. RS and VV carried out the luciferase assays on Mero-14 MPM cells. CDS wrote and revised the paper. FG and PV helped to evaluate, edit and revise the manuscript. SL, AC and LM supervised the development of work, managed the data interpretation and revised the manuscript.
Funding This work was supported by GIME (Gruppo Italiano Mesotelioma) ONLUS and by Ministero della Salute-Bando Ricerca Finalizzata 2009 (RF-2009–1529895).
Competing interests None declared.
Provenance and peer review Not commissioned; externally peer reviewed.
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