TNFRSF1B Gene Variants in Clinicopathological Aspects and Prognosis of Patients with Cutaneous Melanoma

Abstract

Regulatory T lymphocytes play a critical role in immune regulation and are involved in the aberrant cell elimination by facilitating tumor necrosis factor connection to the TNFR2 receptor, encoded by the TNFRSF1B polymorphic gene. We aimed to examine the effects of single nucleotide variants TNFRSF1B c.587T>G, c.*188A>G, c.*215C>T, and c.*922C>T on the clinicopathological characteristics and survival of cutaneous melanoma (CM) patients. Patients were genotyped using RT-PCR. TNFRSF1B levels were measured using qPCR. Luciferase reporter assay evaluated the interaction of miR-96 and miR-1271 with the 3'-UTR of TNFRSF1B. The c.587TT genotype was more common in patients younger than 54 years old than in older patients. Patients with c.*922CT or TT, c.587TG or GG + c.*922CT or TT genotypes, as well as those with the haplotype TATT, presented a higher risk of tumor progression and death due to the disease effects. Individuals with the c.*922TT genotype had a higher TNFRSF1B expression than those with the CC genotype. miR-1271 had less efficient binding with the 3'-UTR of the T allele when compared with the C allele of the SNV c.*922C>T. Our findings, for the first time, demonstrate that TNFRSF1B c.587T>G and c.*922C>T variants can serve as independent prognostic factors in CM patients.

Keywords: TNFRSF1B; clinicopathological aspects; cutaneous melanoma; single nucleotide variant; survival.

Figure 1

Analysis of genetic variants in the TNFRSF1B gene in survival of patients with cutaneous melanomas, where lower progression-free survival and melanoma-specific survival were seen in patients with the TNFRSF1B c.*922CT or TT genotypes (A,B), the TNFRSF1B c.587TG or GG + c.*922CT or TT genotypes (C,D), and TATT haplotype (E,F) compared to patients with the remaining genotypes and haplotypes.

Figure 2

TNFRSF1B gene expression in peripheral blood leukocyte samples from individuals stratified by SNV c.*922C>T variant genotypes. The expression is higher in (A) individuals with the TT genotype when compared to individuals with CC (2.57 arbitrary units (AUs) ± 0.96 standard deviation (SD) versus 1.96 AUs ± 1.26 SD; p = 0.02) or CT (2.57 AUs ± 0.96 SD versus 1.94 AUs ± 1.21 SD; p = 0.008) genotypes and (B) in carriers of the variant T allele when compared to those with the ancestral C allele (2.28 AUs ± 1.11 SD versus 1.95 AUs ± 1.22 SD; p = 0.04). No difference in gene expression was observed in individuals with CC and CT genotypes (p > 0.05) (t test).

Figure 3

Quantification of relative luciferase activity in groups: pMIR c.*922C (CC genotype) co-transfected with microRNA inhibitor (miR)-1271 (considered 100% luciferase enzyme activity); pMIR c.*922C co-transfected with the mimic sequence of miR-1271; pMIR c.*922T (TT genotype) co-transfected with the inhibitor of miR-1271; and pMIR c.*922T co-transfected with the mimic sequence of miR-1271 in melanoma cell lines SK-MEL-28 (A) and A-375 (B). (*) The relative luciferase activity was higher when comparing the group containing the plasmid with the CC genotype co-transfected with the inhibitory sequence of miR-1271 and the group containing the same plasmid co-transfected with the mimic sequence of miR-1271 (SK-MEL-28: 100.0 versus 71.0%; p = 0.04; A-375: 100.0 versus 54.0%; p = 0.01). (**) The relative luciferase activity was lower when comparing the group containing the plasmid with the CC genotype co-transfected with the mimic sequence of miR-1271 and the group containing the plasmid TT co-transfected with the mimic sequence of miR-1271 (SK-MEL-28: 71.0 versus 107.0%, p = 0.02; A-375: 54.0 versus 72.0%, p = 0.01) (t test).