Introduction: Recent experimental evidence shows that intestinal barrier disruption initiates colon inflammation, which may promote colorectal carcinogenesis. In experimental models, vitamin D and calcium improved gut barrier function; however, this has not been assessed in humans. Method: To test the effects of supplemental calcium and vitamin D3 on circulating biomarkers of intestinal mucosal damage (intestinal fatty-acid binding protein, IFABP) and exposure to bacterial products due to impaired gut barrier (lipopolysaccharide-binding protein, LBP), we conducted an “adjunct biomarker study” to a larger 11-center, randomized, placebo-controlled, partial 2 × 2 factorial chemoprevention clinical trial, the Vitamin D/Calcium Polyp Prevention Study. Participants were randomized into four different treatment groups: placebo, 1,200 mg/day calcium, 1,000 IU/day vitamin D3, and 1,200 mg/day calcium plus 1,000 IU/day vitamin D3. Circulating concentrations of LBP and IFABP were measured at baseline and 1-year follow-up using the Meso Scale Discovery electrochemiluminescence assays in a subset of 118 participants included in the adjunct biomarker study. Result: Following one year of vitamin D3 treatment, in the vitamin D3 plus calcium and vitamin D3 groups versus placebo and calcium groups, LBP decreased 10% (316 ng/ml; P=0.23) and IFABP 18% (77 ng/ml; P=0.06). A similar decrease of 20% (P=0.10) was observed for IFABP, but not LBP, in the vitamin D3 plus calcium versus the calcium group. There were no appreciable biomarker changes with calcium treatment. In stratified analyses, we found possible effect modifications by baseline serum 25-OH-vitamin D, total calcium intake, body mass index, and interleukin 6 (IL-6) concentration on estimated vitamin D3 treatment effects on LBP and/or IFABP. Conclusion: These preliminary results are consistent with vitamin D’s potential protective effects on intestinal mucosal barrier and support further research of vitamin D against gut barrier disruption and colorectal neoplasms. Citation Format: Sangji Lee, Veronika Fedirko, Roberd M. Bostick, Bradley Pearce, Elizabeth L. Barry, Robin E. Rutherford, March E. Seabrook. Effects of supplemental calcium and vitamin D on circulating biomarkers of gut barrier function in colorectal adenoma patients: A randomized controlled trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 766.

AbstractGut barrier dysfunction promotes chronic inflammation, contributing to several gastrointestinal diseases, including colorectal cancer. Preliminary evidence suggests that vitamin D and calcium could prevent colorectal carcinogenesis, in part, by influencing gut barrier function. However, relevant human data are scarce. We tested the effects of supplemental calcium (1,200 mg/day) and/or vitamin D3 (1,000 IU/day) on circulating concentrations of biomarkers of gut permeability (anti-flagellin and anti-lipopolysaccharide IgA and IgG, measured via ELISA) from baseline to 1 and 3 or 5 years postbaseline among 175 patients with colorectal adenoma in a randomized, double-blinded, placebo-controlled clinical trial. We also assessed factors associated with baseline concentrations of these biomarkers. We found no appreciable effects of supplemental vitamin D3 and/or calcium on individual or aggregate biomarkers of gut permeability. At baseline, a combined permeability score (the summed concentrations of all four biomarkers) was 14% lower among women (P = 0.01) and 10% higher among those who consumed >1 serving per day of red or processed meats relative to those who consumed none (Ptrend = 0.03). The permeability score was estimated to be 49% higher among participants with a body mass index (BMI) > 35 kg/m2 relative to those with a BMI < 22.5 kg/m2 (Ptrend = 0.17). Our results suggest that daily supplemental vitamin D3 and/or calcium may not modify circulating concentrations of gut permeability biomarkers within 1 or 3–5 years, but support continued investigation of modifiable factors, such as diet and excess adiposity, that could affect gut permeability.Prevention Relevance:Calcium and vitamin D may be involved in regulating and maintaining the integrity of the intestinal mucosal barrier, the dysfunction of which results in exposure of the host to luminal bacteria, endotoxins, and antigens leading to potentially cancer-promoting endotoxemia and chronic colon inflammation. While our results suggest that daily supplementation with these chemopreventive agents does not modify circulating concentrations of gut permeability biomarkers, they support continued investigation of other potential modifiable factors, such as diet and excess adiposity, that could alter gut barrier function, to inform the development of treatable biomarkers of risk for colorectal neoplasms and effective colon cancer preventive strategies.

Table S1. pIKKα/β, TLR4, and TLR5 Expression in the Full Length of Crypts and Stroma in the Differentiation Zone (Upper 40%) in the Normal-Appearing Rectal Mucosa of the Adjunct Biomarker Study Table S2. pIKKα/β, TLR4, and TLR5 Expression in the Full Length of Crypts and Stroma in the Proliferation Zone (Lower 60%) of Normal-Appearing Rectal Mucosa of the Adjunct Biomarker Study Participants (n = 105) During the Triala. Table S3. pIKKα/β, TLR4, and TLR5 Expression in the Ratio (ϕh) of the Differentiation Zone over the Full Length of the Crypt in Normal-Appearing Rectal Mucosa of the Adjunct Biomarker Study Participants (n = 105) During the Triala. Table S4. The Ratios of (pIKKα/β)/TLR4 and (pIKKα/β)/TLR5 in the Crypts in the Normal-Appearing Rectal Mucosa of the Adjunct Biomarker Study Participants (n=105)a. Table S5. Geometric Means, 95% CIs, and Proportional Differences in pIKKα/β, TLR4 and TLR5 Expression in the Full Length of the Crypts and Stroma, by Categories of Baseline Participant Characteristics (n = 105)a.

Abnormal expression of Wnt pathway and DNA mismatch repair proteins is common during colorectal carcinogenesis. To investigate cross-sectional associations of lifestyle, dietary, and other participant characteristics with the expression of such proteins, we measured APC, β-catenin, E-cadherin, and MSH2 colorectal crypt expression in biopsies of normal-appearing colorectal mucosa from 104 sporadic colorectal adenoma patients using automated immunohistochemistry and quantitative image analysis. We used multivariable general linear models to compare adjusted mean biomarker expression across categories of participant characteristics. Example findings include that among women relative to men, mean APC expression in whole crypts, the upper 40% of crypts (differentiation zone), and the lower 60% of crypts (proliferation zone) was 322.9% higher (p<0.01), 296.7% higher (p<0.01), and 399.1% higher (p<0.01), respectively. Among participants with higher alcohol consumption, APC expression in the crypt differentiation zone was estimated to be 15.9% lower (p=0.08). Among those with higher total meat consumption, β-catenin expression in whole crypts and the crypt proliferation zone was estimated to be 20.5% higher (p=0.07) and 19.6% higher (p=0.06), respectively, and MSH2 expression in the crypt differentiation zone was estimated to be 64.4% lower (p=0.10). Among those with a higher body mass index, MSH2 expression in the crypt differentiation zone was estimated to be 87.5% lower (p=0.15). These pilot study findings suggest that being male, higher adiposity, and higher alcohol and meat consumption may be unfavorably associated with biomarkers of colorectal carcinogenesis pathway proteins in the normal-appearing colorectal mucosa of sporadic colorectal adenoma patients and support further investigation in larger studies.

Increased COX-2 and decreased 15-hydroxyprostaglandin dehydrogenase (15-HPGD) expression promote prostaglandin-mediated inflammation and colorectal carcinogenesis. Experimental studies suggest that vitamin D and calcium may inhibit these pathways, but their effects on colorectal tissue COX-2 and 15-HPGD expression in humans are unknown. We tested the effects of supplemental vitamin D(1,000 IU/day) and/or calcium (1,200 mg/day) on COX-2 and 15-HPGD expression in the morphologically normal rectal mucosa from 62 paients with colorectal adenoma in a placebo-controlled chemoprevention trial. We measured biomarker expression using automated IHC and quantitative image analysis at baseline and 1-year follow-up, and assessed treatment effects using mixed linear models. The primary outcome was the COX-2/15-HPGD expression ratio, because these enzymes function as physiologic antagonists. After 1 year of treatment, the mean COX-2/15-HPGD expression ratio in full-length crypts proportionately decreased 47% in the vitamin D group (P = 0.001), 46% in the calciumgroup (P = 0.002), and 34% in the calcium thorn vitamin D group (P = 0.03), relative to the placebo group. Among individuals with the functional vitamin D-binding protein isoform DBP2 (GC rs4588*A), the COX-2/15-HPDG ratio decreased 70% (P = 0.0006), 75% (P = 0.0002), and 60% (P = 0.006) in the vitamin D, calcium, and combined supplementation groups, respectively, relative to placebo. These results show that vitamin D and calcium favorably modulate the balance of expression of COX-2 and 15-HPGD-biomarkers of inflammation that are strongly linked to colorectal carcinogenesis-in the normal-appearing colorectal mucosa of patients with colorectal adenoma (perhaps especially those with the DBP2 isoform). Prevention Relevance: Supplemental calcium and vitamin D reduce indicators of cancer-promoting inflammation in normal colorectal tissue in humans, thus furthering our understanding of how they may help prevent colorectal cancer.

The physical gut barrier, comprised of a thick mucus layer and the epithelium, plays an important role in defense against microbes and foreign antigens. Calcium and vitamin D may be involved in maintaining the integrity of the intestinal mucosal barrier, the dysfunction of which may lead to endotoxemia and inflammation, and contribute to colorectal carcinogenesis. We investigated supplemental calcium (1200 mg, daily) and/or vitamin D-3 (1000 IU daily) effects on intestinal barrier function-related biomarkers in a subset of 105 participants from a large colorectal adenoma recurrence chemoprevention clinical trial. We assessed expression of the tight junction proteins claudin-1 (CLDN1), occludin (OCLD), and mucin-12 (MUC12) in the normal-appearing colorectal mucosa using standardized, automated immunohistochemistry and quantitative image analysis. Following 1 year of treatment, in the calcium relative to the no calcium group, the CLDN1, OCLD, and MUC12 expression increased by 14% (P = 0.17), 23% (P = 0.11), and 22% (P = 0.07), respectively. In secondary analyses, the estimated calcium treatment effects were greater among participants with baseline serum 25-OH-vitamin D concentrations below the median value of 22.69 ng/mL (CLDN1: 29%, P = 0.04; OCLD: 36%, P = 0.06; MUC12: 35%, P = 0.05). There were no biomarker expression changes in the vitamin D-3 alone group; however, modest increases were found in the combined calcium/vitamin D-3 group. At baseline, obesity, history of a sessile-serrated adenoma, colorectal MIB-1/Ki-67 expression, and a family history of colorectal cancer were associated with CLDN1, OCLD, and MUC12 expression. Our study supports continued investigation of factors that could affect intestinal mucosal barrier integrity relevant to colorectal carcinogenesis.

To clarify the roles of vitamin D and calcium as potential chemopreventive agents against colorectal cancer in humans, and to develop “treatable”, pre-neoplastic, phenotypic biomarkers of risk for colorectal neoplasms, we estimated the effects of supplemental vitamin D3 (1,000 IU/day [25 μg/day]) and calcium (1,200 mg/day), alone and in combination, on biomarkers of proliferation (mib-1), differentiation (p21), and apoptosis (bax [apoptosis-promoting] and bcl-2 [apoptosis-inhibiting]), in the normal-appearing rectal mucosa in a subsample of participants (n = 104) in a larger randomized, double-blind, placebo-controlled clinical trial among colorectal adenoma patients. The biomarkers were measured in rectal biopsies at baseline and after one year of follow up, using automated immunohistochemistry and quantitative image analysis. In the vitamin D plus calcium group relative to control, in the crypt differentiation zone (upper 40% of crypts), mib-1 expression decreased 24% (P = 0.28); p21 expression alone and relative to mib-1 expression increased 29% (P = 0.06) and 73% (P = 0.06), respectively; and bax expression relative to mib-1 expression increased 58% (P = 0.21). The estimated vitamin D alone treatment effects were similar but of lesser magnitudes, and those for calcium alone were mixed. All estimated treatment effects on bcl-2 expression were close to the null. These pilot study results support further investigation of whether 1) vitamin D and calcium promote colorectal epithelial cell differentiation, reduce proliferation, and promote apoptosis in the normal-appearing human colorectal mucosa, 2) vitamin D and calcium act as chemopreventive agents against colorectal neoplasms, and 3) mib-1, p21, and bax are potential “treatable”, pre-neoplastic, biomarkers of risk for colorectal neoplasms.

Abnormal expression of the DNA mismatch repair protein MSH2 and autocrine/paracrine transforming growth factors TGF alpha (growth promoter) and TGF beta(1) (growth inhibitor) is common during colorectal carcinogenesis. To estimate vitamin D and calcium effects on these biomarkers in the normal-appearing colorectal mucosa of sporadic colorectal adenoma patients, we conducted a pilot, randomized, double-blinded, placebo-controlled, modified 2 x 2 factorial chemoprevention clinical trial (N = 104) of supplemental vitamin D-3 (1000 IU daily) and calcium (1200 mg daily), alone and in combination, versus placebo over 1 year. The expression of the three biomarkers and Ki-67/mib-1 in colorectal crypts in biopsies of normal-appearing rectal mucosa were detected using automated immunohistochemistry and quantified using image analysis. In the vitamin D-3 and vitamin D-3 plus calcium groups, relative to their reference groups, in the upper 40% (differentiation zone) of crypts, it was estimated that, respectively, the MSH2/mib-1 ratio increased by 47% (P = 0.14) and 62% (P = 0.08), TGF beta(1) expression increased by 41% (P = 0.25) and 78% (P = 0.14), and the TGF alpha/TGF beta(1) ratio decreased by 25% (P = 0.31) and 44% (P = 0.13). Although not statistically significant, these results support further research into (i) whether supplemental vitamin D-3, alone or in combination with calcium, may increase DNA mismatch repair relative to proliferation, increase TGF beta(1) expression, and decrease autocrine/paracrine growth promotion relative to growth inhibition in the colorectal epithelium, all hypothesized to reduce risk for colorectal carcinogenesis; and (ii) the expression of MSH2 relative to mib-1, TGF beta(1) alone, and TGF alpha relative to TGF beta(1) in the normal-appearing rectal mucosa as potential modifiable, pre-neoplastic markers of risk for colorectal neoplasms.

Abstract Chronic inflammation in the colorectum, a significant contributor to colorectal carcinogenesis, can be triggered by the activation of proinflammatory signaling pathways such as those initiated by Toll-like receptors (TLR) and nuclear factor κB (NF-κB). Although experimental evidence supports calcium and vitamin D potentially modifying these proinflammatory pathways in the colorectum, human data in these regards are scarce. We investigated supplemental calcium (1,200 mg daily) and/or vitamin D3 (1,000 IU daily) effects on inflammatory signaling pathway-related biomarkers in a subset of 105 participants from a colorectal adenoma recurrence chemoprevention clinical trial. We assessed expression of TLR4 and TLR5, which recognize the bacterial components lipopolysaccharides and flagellin, respectively, and phospho-IKKα/β (pIKKα/β), a biomarker of inflammation, in the normal-appearing rectal crypt epithelium and stroma using standardized, automated immunohistochemistry and quantitative image analysis. Following 1 year of treatment, TLR4, TLR5, and pIKKα/β expression in the rectal mucosa did not statistically significantly change with vitamin D or calcium supplementation, taken alone or in combination. Several baseline participant characteristics, including body mass index, history of sessile serrated adenomas, high red/processed meat intake, and high levels of rectal epithelial cell proliferation (as measured by MIB-1/Ki-67), were associated with higher baseline expression of TLRs or pIKKα/β. Our findings suggest that vitamin D and calcium may have no substantial effect on the investigated biomarkers. However, several modifiable lifestyle factors may be associated with TLRs and pIKKα/β expression in the normal rectal mucosa, supporting their future investigation as potentially treatable, preneoplastic risk factors for colorectal neoplasms. Cancer Prev Res; 11(11); 707–16. ©2018 AACR.

APC/β‐catenin pathway malfunction is a common and early event in colorectal carcinogenesis. To assess calcium and vitamin D effects on the APC/β‐catenin pathway in the normal‐appearing colorectal mucosa of sporadic colorectal adenoma patients, nested within a larger randomized, double‐blind, placebo‐controlled, partial 2 × 2 factorial chemoprevention clinical trial of supplemental calcium (1200 mg daily) and vitamin D (1000 IU daily), alone and in combination versus placebo, we assessed APC, β‐catenin, and E‐cadherin expression in colon crypts in normal‐appearing rectal mucosa biopsies from 104 participants at baseline and 1‐yr follow up using standardized, automated immunohistochemistry and quantitative image analysis. For vitamin D versus no vitamin D, the ratio of APC expression to β‐catenin expression in the upper 40% (differentiation zone) of crypts (APC/β‐catenin score) increased by 28% (P = 0.02), for calcium versus no calcium it increased by 1% (P = 0.88), and for vitamin D + calcium versus calcium by 35% (P = 0.01). Total E‐cadherin expression increased by 7% (P = 0.35) for vitamin D versus no vitamin D, 8% (P = 0.31) for calcium versus no calcium, and 12% (P = 0.21) for vitamin D + calcium versus calcium. These results support (i) that vitamin D, alone or in combination with calcium, may modify APC, β‐catenin, and E‐cadherin expression in humans in directions hypothesized to reduce risk for colorectal neoplasms; (ii) vitamin D as a potential chemopreventive agent against colorectal neoplasms; and (iii) the potential of APC, β‐catenin, and E‐cadherin expression as treatable, pre‐neoplastic risk biomarkers for colorectal neoplasms. © 2016 Wiley Periodicals, Inc.

Compared to whites, blacks have higher colorectal cancer incidence and mortality rates and are at greater risk for early-onset disease. The reasons for this racial disparity are poorly understood, but one contributing factor could be differences in access to high-quality screening and medical care.

BACKGROUND:Epidemiologic and preclinical data suggest that higher intake and serum levels of vitamin D and higher intake of calcium reduce the risk of colorectal neoplasia. To further study the chemopreventive potential of these nutrients, we conducted a randomized, double-blind, placebo-controlled trial of supplementation with vitamin D, calcium, or both for the prevention of colorectal adenomas.METHODS:We recruited patients with recently diagnosed adenomas and no known colorectal polyps remaining after complete colonoscopy. We randomly assigned 2259 participants to receive daily vitamin D3 (1000 IU), calcium as carbonate (1200 mg), both, or neither in a partial 2×2 factorial design. Women could elect to receive calcium plus random assignment to vitamin D or placebo. Follow-up colonoscopy was anticipated to be performed 3 or 5 years after the baseline examinations, according to the endoscopist's recommendation. The primary end point was adenomas diagnosed in the interval from randomization through the anticipated surveillance colonoscopy.RESULTS:Participants who were randomly assigned to receive vitamin D had a mean net increase in serum 25-hydroxyvitamin D levels of 7.83 ng per milliliter, relative to participants given placebo. Overall, 43% of participants had one or more adenomas diagnosed during follow-up. The adjusted risk ratios for recurrent adenomas were 0.99 (95% confidence interval [CI], 0.89 to 1.09) with vitamin D versus no vitamin D, 0.95 (95% CI, 0.85 to 1.06) with calcium versus no calcium, and 0.93 (95% CI, 0.80 to 1.08) with both agents versus neither agent. The findings for advanced adenomas were similar. There were few serious adverse events.CONCLUSIONS:Daily supplementation with vitamin D3 (1000 IU), calcium (1200 mg), or both after removal of colorectal adenomas did not significantly reduce the risk of recurrent colorectal adenomas over a period of 3 to 5 years. (Funded by the National Cancer Institute; ClinicalTrials.gov number, NCT00153816.).

Increased colorectal epithelial cell proliferation is an early, common event in colorectal carcinogenesis. We conducted a pilot, colonoscopy-based case-control study (n=49 cases, 154 controls) of incident, sporadic colorectal adenoma to investigate endogenous cell growth factors and receptor, as well as the balance of growth factors, as potential modifiable pre-neoplastic biomarkers of risk for colorectal neoplasms. We measured transforming growth factor alpha (TGF), TGF(1), and TGF receptor II (TGFRII) expression in normal-appearing mucosa from the rectum, sigmoid colon, and ascending colon using automated immunohistochemistry and quantitative image analysis. Diet and lifestyle were assessed via questionnaires. The mean ratio of rectal TGF to TGF(1) expression and mean rectal TGF expression were, respectively, 110% (P=0.02) and 49% (P=0.04) higher in cases than in controls, and associated with a more than two-fold (OR 2.42, 95% CI 0.85-6.87) and a 62% (OR 1.62, 95% CI 0.63-4.19) higher risk of colorectal adenoma. TGF(1) and TGFRII expression were 6.7% (P=0.75) and 7.2% (P=0.49), respectively, lower in cases than in controls. The TGF/TGF(1) expression ratio was 105% higher among smokers than among non-smokers (P=0.03). These preliminary data suggest that the balance of TGF and TGF(1) expression, and to a lesser extend TGF alone, in the normal-appearing rectal mucosa may be directly associated with risk for incident, sporadic colorectal neoplasms, as well as with modifiable risk factors for colorectal neoplasms. (c) 2014 Wiley Periodicals, Inc.

Kane, Sunanda MD, MSPH, FACG; Leighton, Jonathan MD, FACG; Kefalas, Costas MD, FACG; Cohen, Lawrence MD, FACG; Katz, Philip MD, FACG; Rizk, Maged MD, FACG; Pike, Irving MD, FACG; Lashner, Bret MD, MPH, FACG; Ho, Immanuel MD, FACG; Pochapin, Mark MD, FACG; Seabrook, March MD, FACG; Greenwald, David MD, FACG; DeMarco, Daniel MD, FACG; Johnson, David MD, FACG Author Information

Background: Malfunctioning of the adenomatous polyposis coli (APC)/β-catenin signaling pathway is both an early and common event in sporadic colorectal cancer. To assess the potential of APC/β-catenin signaling pathway markers as treatable, preneoplastic biomarkers of risk for colorectal neoplasms, we conducted a pilot colonoscopy-based case–control study (51 cases and 154 controls) of incident, sporadic colorectal adenoma. Methods: We evaluated APC, β-catenin, and E-cadherin expression in normal mucosa from the rectum and ascending and sigmoid colon using automated immunohistochemical and quantitative image analysis. Diet, lifestyle, and medical history were assessed with validated questionnaires. Results: In the normal rectal mucosa, the ratio of the proportion of APC expression in the upper 40% of crypts with total β-catenin expression (APC/β-catenin score) was 14.3% greater in controls than in cases [P = 0.02; OR, 0.40; 95% confidence interval (CI), 0.14–1.14]. Compared with controls, in cases, APC expression was 3.2% lower, β-catenin expression was 3.0% higher, and E-cadherin expression was 0.7% lower; however, none of these differences were statistically significant. The APC/β-catenin score statistically significantly differed according to categories of plausible risk factors for colorectal cancer [e.g., it was 17.7% higher among those with 25(OH) vitamin D3 concentrations ≥ 27 ng/mL]. Conclusions: These preliminary data suggest that the combined expression of APC and β-catenin in the normal rectal mucosa may be associated with risk for incident, sporadic colorectal neoplasms, as well as with modifiable risk factors for colorectal neoplasms. Impact: Our results may help advance the development of treatable, preneoplastic biomarkers of risk for colorectal neoplasms. Cancer Epidemiol Biomarkers Prev; 21(6); 969–79. ©2012 AACR.

Background: Transforming growth factor-α (TGF-α), a stimulatory growth factor and member of the epidermal growth factor family, is a mediator of oncogenesis and malignant progression in colorectal carcinogenesis. Limited evidence suggests its utility as a growth-related biomarker of risk for colorectal cancer. Methods: We measured expression of TGF-α in biopsies of normal-appearing colorectal mucosa using automated immunohistochemistry and quantitative image analysis in a subsample of 29 cases and 31 controls from a colonoscopy-based case-control study (n = 203) of biomarkers of risk for incident sporadic colorectal adenoma. Diet, lifestyle, and medical history were assessed with validated questionnaires. Results: TGF-α expression in the rectum was 51% higher in cases compared with controls (P = 0.05) and statistically significantly associated with accepted risk factors for colorectal neoplasms (36% lower among nonsteroidal anti-inflammatory drug users, 49% lower among women using hormone replacement therapy, 79% higher among persons with a family history of colorectal cancer). Conclusions: TGF-α expression in the normal-appearing rectal mucosa shows promise as an early, potentially modifiable biomarker of risk for colorectal cancer. (Cancer Epidemiol Biomarkers Prev 2009;18(1):65–73)

To characterize the expression of the mismatch repair gene MutL-homolog 1 (MLH1) in normal colorectal crypts in humans, and assess parameters of its expression as a potential biomarker of risk for colorectal neoplasms, we conducted a pilot, colonoscopy-based case-control study (51 cases, 154 controls) of incident, sporadic colorectal adenoma. Biopsies of normal-appearing rectal, sigmoid, and ascending colon mucosa were procured, immunohistochemically processed for MLH1 protein, and analyzed using custom quantitative image analysis procedures. MLH1 expression in the ascending colon was, on average, 49% proportionally lower in cases than controls (P = 0.03), but there was little evidence for case-control differences in the rectum and sigmoid colon. In cases and controls, average MLH1 expression in the ascending colon tended to be lower with increased age [by 56% (P = 0.02) and 25% (P = 0.16), respectively, for those > or =55 years], and with a history of colorectal cancer in a first-degree relative (by 22% [P = 0.56] and 34% [P = 0.16], respectively). Among cases, but not controls, average MLH1 expression tended to be higher with current alcohol consumption, regular aspirin use, and higher total intakes of calcium, vitamin D, and folate. There was little indication of similar differences in the rectum. These preliminary data suggest that lower MLH1 expression in the normal colonic mucosa, at least in the ascending colon, may be associated with increased risk of incident, sporadic colorectal adenoma, as well as with modifiable risk factors for colorectal neoplasms, thus supporting further investigation of MLH1 expression as a potential "treatable" biomarker of risk for colorectal neoplasms.

To characterize the expression of the mismatch repair gene MSH2 in normal colorectal crypts in humans and assess parameters of its expression as a potential modifiable biomarker of risk for colorectal neoplasms, we conducted a pilot, colonoscopy-based case-control study (51 cases and 154 controls) of incident, sporadic colorectal adenoma. Biopsies of normal-appearing rectal, sigmoid, and ascending colon mucosa were procured, immunohistochemically processed for MSH2 protein, and analyzed using custom quantitative image analysis procedures. MSH2 expression in adenoma cases was lower than in controls by 49% (P = 0.01) and 23% (P = 0.06) in the ascending colon and rectum, respectively, but not in the sigmoid colon. MSH2 expression in the rectum was 39% (P = 0.04) higher in subjects who regularly took a nonsteroidal anti-inflammatory drug than in those who did not, and it tended to be lower in those with adenomas in the right colon and those who had an adenoma with more advanced characteristics. These preliminary data suggest that lower MSH2 expression in the normal colonic mucosa, at least in the ascending colon and rectum, may be associated with increased risk of incident, sporadic colorectal adenoma as well as with modifiable risk factors for colorectal neoplasms, thus supporting further investigation of MSH2 expression as a potential modifiable biomarker of risk for colorectal neoplasms. (Cancer Epidemiol Biomarkers Prev 2009;18(11):2965–73)