The results of the current work do not provide support for a gene-level relationship between RXRA and CASR and risk for colorectal cancer. However, the associations observed for one CASR SNP and two RXRA SNPs selected a priori are of interest. Carriage of at least one A allele at RXRA rs7861779 was shown to be associated with higher risk of proximal colorectal cancer, whereas the presence of the A allele at RXRA rs12004589 was linked with higher risk of MSI-high cancers. Additionally, the high-interest CASR SNP rs1801726 was significantly related to reduced risk of rectal cancer.
While the study of genetic variation in nuclear receptors has most often focused on the heterodimer partners of RXR such as VDR and PPAR, evidence supports a role for RXR itself in carcinogenesis. Ligands for RXRα, an isoform expressed in the colonic epithelium, enhance the interaction between RXR and β-catenin and induce β-catenin degradation (25
). In combination with PPAR ligands, RXR agonists act synergistically to inhibit growth in colon cancer cell lines (27
). RXR ligands also alter the cellular response to the VDR ligand 1,25(OH)2
. In Caco-2 cells, the addition of RXR agonists increased the proliferative response, whereas in HT-29 cells, the response was blocked (28
). Hence, experimental work indicates that RXR has an active role in carcinogenesis, and this role may vary by a complex network of potential heterodimer partner selections.
In previous work, carriage of at least one A
allele for RXRA
rs7861779 was shown to be associated with reduced odds of proximal colorectal adenoma (11
), whereas the results of the current work show increased risk of proximal colorectal cancer with presence of at least one of these alleles. Any mechanism for explaining why RXRA
would have different roles in adenoma as compared with cancer is purely speculative, particularly as rs7861779 is located within an intron. Although intronic SNPs can potentially affect alternative splicing of RNA (29
), it is possible that this SNP is in LD with a site of genetic variation, which exerts a conformational change in the resultant protein, which in turn has biological effects. If this were the case, the functional change might be related to the capacity of RXRα to heterodimerize with specific nuclear receptor partners such as VDR and PPAR, both of which are of interest to colorectal carcinogenesis. The RXR/PPAR heterodimer is permissive and therefore can be activated by ligands for either receptor; provision of ligands simultaneously exerts a synergistic effect of cell growth inhibition and apoptosis induction above the activity of the ligands alone (32
). If the structure of RXRα was changed such that it exhibited decreased affinity for PPAR heterodimerization and/or preference for an alternate receptor, this might lead to cancer progression. In turn, the same conformational change might be protective from the formation of new adenomas in the colorectum by increasing the affinity of RXR for VDR and thus activation of the latter, which may have a critical role in the maintenance of normal colon tissue after removal of adenomas (33
). Overall, while the results of the present study were contrary to what was expected based on prior work involving adenomas, the repeated finding of an association for this particular SNP and proximal colorectal neoplasia indicates that it may be marking a region of RXRA
that has the potential for activity in proximal colon carcinogenesis.
While it is unclear as to why RXRA
rs7861779 has been found to be associated with proximal and not distal lesions, neoplasia of these colorectal subsites may represent biologically disparate pathologies with potentially distinct genetic etiologies (35
). PPARγ, one isoform of an RXRA
heterodimer partner, exhibits different patterns of expression in the proximal and distal colon and regulates different genes within each colorectal subsite (36
). In addition, chromosomal instability is more often described in the distal region, and genetic instability and MSI are more often found in proximal neoplasia (37
). Interestingly, an RXRA
SNP in strong LD with rs7861779 and rs12004589 was significantly associated with MSI-high lesions in the current work. MSI is associated with defective mismatch repair (38
), most often because of methylation of mutL humolog 1, a mismatch repair protein (19
). Previous work has shown that RXR interacts with the T:G mismatch-specific thymine-DNA glycolylase, which has a critical role in initiating the repair of methylated DNA (39
). Though the potential mechanism of action is currently unknown, this finding provides more evidence that these SNPs may be marking a functional change that affects colorectal neoplasia of a particular biological pathway, possibly involving MSI.
With regard to CASR
, many epidemiological studies have reported that calcium intake is related to a reduced risk for colorectal neoplasia (2
). In a randomized, placebo-controlled intervention trial, Baron et al.
) demonstrated that daily supplementation with calcium carbonate significantly reduced the risk of adenoma recurrence compared with the placebo group. The CASR is critical for maintenance of circulating calcium concentrations. This receptor is present in colon epithelial tissue, but in colon cancer, this expression is decreased (15
). The promoter region of CASR
contains vitamin D response elements (12
), supporting a mechanistic relationship between vitamin D and calcium, and genetic variation in CASR
may increase or reduce calcium absorption and therefore have implications in colorectal carcinogenesis (3
). There have been several prior studies of variation in this gene and risk for colorectal cancers. In work by Dong et al.
), the authors reported that whereas there were no associations with colorectal cancer overall, several CASR
SNPs were significantly associated with proximal lesions (rs10934578, rs12485716, rs4678174 and rs2270916); while in the current work, these SNPs were not related to colorectal cancer. Another high-interest SNP in CASR
is rs1801725; two studies reported a significant link with colorectal cancer (16
), whereas others found no relationship (13
). Specifically, Speer et al.
) reported an association between this SNP and more advanced rectal tumors, and the results of the current work also reveal a significant association for this SNP that was observed only with rectal cancers. When conducting analyses of all CASR
SNPs and adjusting for multiple comparisons, rs17203502 was significantly associated with risk for proximal colorectal cancer for the recessive mode of inheritance (supplementary Table I
is available at Carcinogenesis
Online). In another study of colorectal cancer, this intronic SNP was not associated with any cancer or proximal lesions; however, in that work, only one mode of inheritance was investigated (13
Whereas there are several published reports for CASR
and colorectal neoplasia, to our knowledge only one study, discussed above (11
), has evaluated association of RXRA
and colorectal lesions. However, there are two published papers investigating RXRA
and other cancer sites. In one study, genetic variation in RXRA
and risk for prostate cancer was investigated and no relationship was observed (42
), though only 11 SNPs in RXRA
were included in the work. In another study, no significant associations between two RXRA
SNPs (rs1536475 and rs1805343) and biliary tract cancers were detected (43
); these SNPs were not significantly related to colorectal cancer risk in the current work, nor with metachronous colorectal adenoma in another study (11
A limitation of this work includes the lack of statistical power for investigating racial or ethnic differences in the reported associations. Because of relatively low numbers of non-white study participants, we could not conduct robust stratified analyses. Given the strong relationship between vitamin D and race/ethnicity, replication of these findings in a more diverse population is necessary.
In summary, the current work provides support for a role of CASR and RXRA in the development of colorectal cancer. The results for CASR support prior findings of a possible role in rectal cancers specifically, whereas those for RXRA provide further evidence of an important functional role in the proximal colon, which may be related in part to tumor MSI status.