Genetic variation determines VEGF-A plasma levels in cancer patients

Abstract

Angiogenesis is essential in tumor biology and is regulated by vascular endothelial growth factor (VEGF) ligands and receptors. Here we aimed to discover genetic variants associated with levels of circulating angiogenic proteins in cancer patients. Plasma was collected at baseline in 216 pancreatic and 114 colorectal cancer patients. Thirty-one angiogenic proteins were measured by ELISA. 484,523 Single Nucleotide Polymorphisms (SNP) were tested for association with plasma levels for each protein in pancreatic cancer patients. Three top-ranked hits were then genotyped in colorectal cancer patients, where associations with the same proteins were measured. The results demonstrated rs2284284 and MCP1 (P-value = 6.7e-08), rs7504372 and VEGF-C (P-value = 9.8e-09), and rs7767396 and VEGF-A (P-value = 5.8e-09) were SNP-protein pairs identified in pancreatic cancer patients. In colorectal cancer patients, only rs7767396 (A > G) and VEGF-A was validated (P-value = 5.18e-05). The AA genotype of rs7767396 exhibited 2.04-2.3 and 2.7-3.4-fold higher VEGF-A levels than those with AG and GG genotypes. The G allele of rs7767396 reduces binding of the NF-AT1 transcription factor. In conclusion, a common genetic variant predicts the plasma levels of VEGF-A in cancer patients through altered binding of NF-AT1.

Figure 1

CONSORT chart for CALGB 80303. Abbreviations: MAF, minor allele frequency; HWE, Hardy-Weinberg Equilibrium.

Figure 2

CONSORT chart for CALGB 80203. Abbreviations: MAF, minor allele frequency; HWE, Hardy-Weinberg Equilibrium.

Figure 3

Associations between rs2284284 and MCP1 levels, rs7504372 and VEGF-C levels, and rs7767396 and VEFG-A levels in both CALGB 80303 and 80203. Boxes represent 25^th (Q1) and 75^th (Q3) percentiles; Horizontal lines indicate the medians; Upper whiskers indicate min(max(x), Q3 + 1.5 * IQR); Lower whiskers indicate max(min(x), Q1–1.5 * IQR). Points indicate observations.

Figure 4

rs7767396 disrupts NF-AT1-DNA binding in vitro. Plot shows the NF-AT1 8-mer PBM data for a 21-bp genomic region centered at rs7767396. The reference allele is bound specifically by the protein, as indicated by the fact that it overlaps two 8-mers with a binding score above 0.37 (dotted line), which corresponds to a false discovery rate (FDR) of 0.001 for calling transcription factor binding sites. The alternate allele is not bound specifically by NF-AT1 in vitro, as indicated by the fact that all 8-mers overlapping the allele have low binding scores.