Aspirin and Colorectal Cancer According to CTNNB1 Alterations
Background Regular aspirin use reduces the risk for colorectal cancer (CRC), possibly through inhibition of WNT/cadherin-associated protein β1 (CTNNB1 or β-catenin) signaling. The single nucleotide polymorphism (SNP) rs6983267 on chromosome 8q24 is a CRC susceptibility locus that affects binding activity of transcription factor 7 like-2 (TCF7L2) to CTNNB1, thereby altering expression of target oncogenes, including MYC.
Methods We evaluated regular aspirin use and CRC risk according to genotypes of SNP rs6983267 and CTNNB1 expression status in two prospective case–control studies (840 CRC case patients and 1686 age- and race-matched control subjects) nested within the Nurses' Health Study and the Health Professionals Follow-up Study. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) using unconditional logistic regression models. All statistical tests were two-sided.
Results A lower risk of CRC was associated with regular aspirin use and the T allele of rs6983267. The effect of aspirin was confined to individuals with protective T allele of rs6983267 (additive matching factors–adjusted OR for T allele = 0.83; 95% CI = 0.74 to 0.94; Ptrend = .002; Pinteraction = .01). Additionally, the T allele of rs6983267 was associated with a reduced expression of MYC oncogene (Ptrend = .03). Moreover, among individuals with protective T allele, the effect of regular aspirin use was limited to those with positive nuclear CTNNB1 expression. In a functional analysis, in vitro treatment of LS174T cells (a cell line heterozygous for rs6983267) with aspirin was statistically significantly associated with higher G/T allelic ratio of TCF7L2 immunoprecipitated DNA (P = .03).
Conclusions Our results support an influence of aspirin on WNT/CTNNB1 signaling and suggest that aspirin chemoprevention may be tailored according to rs6983267 genotype.
Considerable evidence demonstrates that regular aspirin use reduces the risk of colorectal neoplasms. However, routine use of aspirin for chemoprevention of colorectal cancer is not currently recommended because of uncertainty about its risk–benefit profile. It remains unclear whether there are specific markers that can identify individuals who may obtain differential benefit from aspirin chemoprevention. Substantial data show that the anticancer benefit of aspirin is mediated by inhibition of prostaglandin synthase (PTGS or cyclooxygenase [COX]) enzymes responsible for the conversion of arachidonic acid to prostaglandins. We have previously shown that regular aspirin use is associated with a lower risk of colorectal cancers that overexpress PTGS2 (COX-2) but not risk of cancers that do not overexpress PTGS2 (COX-2). Emerging evidence suggests that aspirin may also modulate WNT/cadherin-associated protein β1 (CTNNB1 or β-catenin) signaling, one of the most essential oncogenic pathways in colorectal cancer. Aspirin inhibits COX-mediated synthesis of prostaglandin E2 (PGE2) that stimulates the CTNNB1 pathway. Through COX-independent pathways, aspirin also induces phosphorylation, ubiquination, and degradation of CTNNB1. Thus, the net result of aspirin is inhibition of CTNNB1 signaling (Figure 1, A and B).
(Enlarge Image)
Figure 1.
Proposed mechanisms of aspirin, the single nucleotide polymorphism rs6983267, and their interaction on the WNT/CTNNB1 pathway. A) Aspirin dampens CTNNB1 signaling through inhibition of PTGS (COX)–mediated synthesis of PGE2. PGE2 stimulates the CTNNB1 pathway through EP2 receptors, coupling the heterotrimeric G proteins of the Gs family and inhibiting the CTNNB1 destruction complex. This leads to release of GSK-3β from the complex, thereby blocking phosphorylation and destruction of CTNNB1. Subsequently, CTNNB1 accumulates in the cytoplasm, eventually translocating into the nucleus and complexing with the TCF/LEF family transcription factor TCF7L2, thereby promoting expression of target oncogenes, including MYC and CCND1, which promote colorectal tumorigenesis. B) Through COX-independent pathways, aspirin also inhibits CTNNB1 signaling by directly inducing phosphorylation and subsequent degradation of CTNNB1. C) The T allele of the single nucleotide polymorphism rs6983267 impairs binding of CTNNB1/TCF7L2, downregulating expression of MYC, the oncogene most proximate to rs6983267. Aspirin lowers level of CTNNB1 (A and B), further impairing binding of CTNNB1/TCF7L2. D) In contrast, the G allele of rs6983267 leads to constitutively active binding of CTNNB1/TCF7L2 and expression of MYC, promoting carcinogenesis. Thus, among individuals with only G allele, binding of CTNNB1/TCF7L2 is less sensitive to nuclear CTNNB1 levels, resulting in relative resistance to the effect of aspirin on inhibition of CTNNB1 accumulation.
Previous genome-wide association studies have consistently identified a single nucleotide polymorphism (SNP) rs6983267 on chromosome 8q24 as a susceptibility locus for colorectal cancer, with the T allele associated with a 15% to 18% lower risk of colorectal cancer. The most proximate gene in this region is MYC, residing 335kb downstream from rs6983267. Previous in vivo and in vitro experiments have demonstrated that the T allele of the SNP rs6983267 impairs binding of WNT/CTNNB1 pathway-related transcription factor 7 like-2 (TCF7L2), inhibiting the MYC promoter (Figure 1, C and D). Most recently, SNP rs6983267 has been shown to functionally reduce MYC expression, inducing resistance to intestinal tumorigenesis.
Based on these results, we examined whether the effect of aspirin use on colorectal cancer risk varied according to rs6983267 genotype. We hypothesized that if aspirin influences colorectal carcinogenesis through modulation of WNT signaling, individuals may derive differential benefit with regular aspirin use according to rs6983267 genotype. If confirmed, such associations would be a novel example of a gene–environment interaction in which the effect of a genome-wide association study susceptibility locus for colorectal cancer was modified by the presence or absence of a known environmental factor (eg, medication use) associated with disease. Moreover, to further validate such a gene–environment interaction, we confirmed its biological plausibility by determining whether rs6983267 is associated with differential MYC expression in tumors and whether the joint effect of aspirin and the rs6983267 T allele may be particularly pronounced for tumors that appear more dependent on CTNNB1 activation for their growth.
Abstract and Study Population
Abstract
Background Regular aspirin use reduces the risk for colorectal cancer (CRC), possibly through inhibition of WNT/cadherin-associated protein β1 (CTNNB1 or β-catenin) signaling. The single nucleotide polymorphism (SNP) rs6983267 on chromosome 8q24 is a CRC susceptibility locus that affects binding activity of transcription factor 7 like-2 (TCF7L2) to CTNNB1, thereby altering expression of target oncogenes, including MYC.
Methods We evaluated regular aspirin use and CRC risk according to genotypes of SNP rs6983267 and CTNNB1 expression status in two prospective case–control studies (840 CRC case patients and 1686 age- and race-matched control subjects) nested within the Nurses' Health Study and the Health Professionals Follow-up Study. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) using unconditional logistic regression models. All statistical tests were two-sided.
Results A lower risk of CRC was associated with regular aspirin use and the T allele of rs6983267. The effect of aspirin was confined to individuals with protective T allele of rs6983267 (additive matching factors–adjusted OR for T allele = 0.83; 95% CI = 0.74 to 0.94; Ptrend = .002; Pinteraction = .01). Additionally, the T allele of rs6983267 was associated with a reduced expression of MYC oncogene (Ptrend = .03). Moreover, among individuals with protective T allele, the effect of regular aspirin use was limited to those with positive nuclear CTNNB1 expression. In a functional analysis, in vitro treatment of LS174T cells (a cell line heterozygous for rs6983267) with aspirin was statistically significantly associated with higher G/T allelic ratio of TCF7L2 immunoprecipitated DNA (P = .03).
Conclusions Our results support an influence of aspirin on WNT/CTNNB1 signaling and suggest that aspirin chemoprevention may be tailored according to rs6983267 genotype.
Study Population
Considerable evidence demonstrates that regular aspirin use reduces the risk of colorectal neoplasms. However, routine use of aspirin for chemoprevention of colorectal cancer is not currently recommended because of uncertainty about its risk–benefit profile. It remains unclear whether there are specific markers that can identify individuals who may obtain differential benefit from aspirin chemoprevention. Substantial data show that the anticancer benefit of aspirin is mediated by inhibition of prostaglandin synthase (PTGS or cyclooxygenase [COX]) enzymes responsible for the conversion of arachidonic acid to prostaglandins. We have previously shown that regular aspirin use is associated with a lower risk of colorectal cancers that overexpress PTGS2 (COX-2) but not risk of cancers that do not overexpress PTGS2 (COX-2). Emerging evidence suggests that aspirin may also modulate WNT/cadherin-associated protein β1 (CTNNB1 or β-catenin) signaling, one of the most essential oncogenic pathways in colorectal cancer. Aspirin inhibits COX-mediated synthesis of prostaglandin E2 (PGE2) that stimulates the CTNNB1 pathway. Through COX-independent pathways, aspirin also induces phosphorylation, ubiquination, and degradation of CTNNB1. Thus, the net result of aspirin is inhibition of CTNNB1 signaling (Figure 1, A and B).
(Enlarge Image)
Figure 1.
Proposed mechanisms of aspirin, the single nucleotide polymorphism rs6983267, and their interaction on the WNT/CTNNB1 pathway. A) Aspirin dampens CTNNB1 signaling through inhibition of PTGS (COX)–mediated synthesis of PGE2. PGE2 stimulates the CTNNB1 pathway through EP2 receptors, coupling the heterotrimeric G proteins of the Gs family and inhibiting the CTNNB1 destruction complex. This leads to release of GSK-3β from the complex, thereby blocking phosphorylation and destruction of CTNNB1. Subsequently, CTNNB1 accumulates in the cytoplasm, eventually translocating into the nucleus and complexing with the TCF/LEF family transcription factor TCF7L2, thereby promoting expression of target oncogenes, including MYC and CCND1, which promote colorectal tumorigenesis. B) Through COX-independent pathways, aspirin also inhibits CTNNB1 signaling by directly inducing phosphorylation and subsequent degradation of CTNNB1. C) The T allele of the single nucleotide polymorphism rs6983267 impairs binding of CTNNB1/TCF7L2, downregulating expression of MYC, the oncogene most proximate to rs6983267. Aspirin lowers level of CTNNB1 (A and B), further impairing binding of CTNNB1/TCF7L2. D) In contrast, the G allele of rs6983267 leads to constitutively active binding of CTNNB1/TCF7L2 and expression of MYC, promoting carcinogenesis. Thus, among individuals with only G allele, binding of CTNNB1/TCF7L2 is less sensitive to nuclear CTNNB1 levels, resulting in relative resistance to the effect of aspirin on inhibition of CTNNB1 accumulation.
Previous genome-wide association studies have consistently identified a single nucleotide polymorphism (SNP) rs6983267 on chromosome 8q24 as a susceptibility locus for colorectal cancer, with the T allele associated with a 15% to 18% lower risk of colorectal cancer. The most proximate gene in this region is MYC, residing 335kb downstream from rs6983267. Previous in vivo and in vitro experiments have demonstrated that the T allele of the SNP rs6983267 impairs binding of WNT/CTNNB1 pathway-related transcription factor 7 like-2 (TCF7L2), inhibiting the MYC promoter (Figure 1, C and D). Most recently, SNP rs6983267 has been shown to functionally reduce MYC expression, inducing resistance to intestinal tumorigenesis.
Based on these results, we examined whether the effect of aspirin use on colorectal cancer risk varied according to rs6983267 genotype. We hypothesized that if aspirin influences colorectal carcinogenesis through modulation of WNT signaling, individuals may derive differential benefit with regular aspirin use according to rs6983267 genotype. If confirmed, such associations would be a novel example of a gene–environment interaction in which the effect of a genome-wide association study susceptibility locus for colorectal cancer was modified by the presence or absence of a known environmental factor (eg, medication use) associated with disease. Moreover, to further validate such a gene–environment interaction, we confirmed its biological plausibility by determining whether rs6983267 is associated with differential MYC expression in tumors and whether the joint effect of aspirin and the rs6983267 T allele may be particularly pronounced for tumors that appear more dependent on CTNNB1 activation for their growth.