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932 Fecal MicroRNAs: Novel Biomarkers for Colon Cancer Screening
FIT = fecal immunochemical test *Advanced neoplasia = adenoma ≥ 10mm, villous component > 25%, high grade dysplasia and CRC **Threshold for a positive FIT is ≥ 75 ng/mL † P-value 0.005; ‡ P-value 0.002
AGA Abstracts
930 Comparing Three Different Strategies of Double Sampling by Fecal Immunochemical Tests for Detection of Advanced Colorectal Neoplasm's Sietze T. Van Turenhout, Frank A. Oort, Veerle M. Coupe, René W. van der Hulst, Eric C. Wesdorp, Ilhame Ben Larbi, Shannon L. Kanis, Edwin van Hengel, Anneke A. Bouman, Gerrit A. Meijer, Chris J. Mulder
932 Fecal MicroRNAs: Novel Biomarkers for Colon Cancer Screening Alexander Link, Francesc Balaguer, Yan Shen, Takeshi Nagasaka, Juan José Lozano, C. R. Boland, Ajay Goel
Background Fecal Immunochemical Tests (FITs) are widely used for Colorectal Cancer (CRC) screening. Some sampling schemes use one-day FIT testing, whereas others use twoday FIT testing. Data on sensitivity and specificity of two-day FIT testing are lacking. This study aims to compare the difference in sensitivity and specificity of three different strategies of two-day FIT sampling, at different cut-off values. Methods In three hospitals in the Amsterdam area, all subjects ≥18 years who were referred for elective colonoscopy were asked to perform a FIT (OC sensor®) on two consecutive days. Colonoscopy and histology were considered as gold standard. Two-day FIT sampling was defined positive in three different ways: 1) when at least one of two FITs was above the cut-off value (FIT+), 2) when both FITs were above the cut-off value(FIT++), and 3) when the geometric mean of two FITs was above the cut-off value (FITmean). Test performance of all three strategies was evaluated at cut-off values of 50, 75 and 100ng/ml. Results In 1105 subjects with complete colonoscopy, 140 (9,4%) AN were found (34 CRCs and 106 advanced adenomas). Of the CRC cases, 70% were Dukes stage A or B (stage unknown in 2). Positivity rates for FIT+ ranged from 16-23%, for FIT++ from 10-13% and for FITmean from 13-17% (see table). At any cut-off level, FIT++ resulted in the lowest and FIT+ in the highest sensitivity. FIT++ showed a higher specificity than both other strategies (see table). Conclusions The most sensitive strategy in two-day FIT sampling is defining the test positive when at least one of both FITs passes the cut off (FIT+), whereas the highest specificity is obtained when both FITs need to be positive for calling the tests positive (FIT++). At all cut-off values, FITmean offers substantially better specificity than FIT+, while sensitivity is substantially higher than FIT++. Sensitivity and specificity of three methods to use two-day FIT sampling for the detection of advanced neoplasia
Introduction: Colorectal cancer (CRC) is the second leading cause of cancer-related deaths in the United States. Although screening can reduce cancer-related mortality, the noninvasive screening programs have achieved only a modest decrease in mortality. MicroRNAs (miRNAs) play important roles in a wide array of biological processes by regulating gene expression. Expression patterns of miRNAs are commonly dysregulated in neoplasia and unique miRNA expression patterns allow discrimination between various types of cancers. In this work, we evaluated the feasibility of fecal miRNAs as biomarkers for colorectal neoplasia screening. Material and methods: Total RNA was extracted from feces using Qiagen miRNAeasy Mini Kits with minor modifications. Illumina microRNA microarray profiling was performed to evaluate the differences in miRNA expression patterns between normal colonic mucosa tissues and stool samples from healthy subjects. Quantitative RTPCR was performed for a subset of miRNAs (including miR-21, -16, -26b, -17 and -29b) in stool samples from 8 healthy volunteers. We also determined miRNA expression in stool samples collected with FOBT kits from 29 subjects with normal colonoscopies (n=10), adenomas (n=9) or CRCs (n=10). Results: We efficiently extracted miRNAs from stool specimens using our modified protocol. Analysis of two independent extractions showed high reproducibility for miRNA extraction and expression. miRNA expression patterns were similar in stool specimens among healthy volunteers, and the pattern was reproducible in stool samples that were collected serially in time from the same individuals. miRNA expression profiles from stool samples collected with FOBT kits from 29 patients demonstrated significantly higher expression of miR-21 and miR-106a in patients with adenomas and CRCs, compared with individuals free of colorectal neoplasia (miR-21, p<0.02; and miR-106a, p<0.05). Conclusion: Our data indicate that miRNAs can be extracted from stool easily and reproducibly. The stools of patients with colorectal neoplasms have unique and identifiable patterns of miRNA expression. These findings suggest that fecal miRNA analysis is an excellent candidate for the development of a noninvasive screening test for colorectal neoplasia. 933
931
Pregabalin Prevents Development of Esophageal Hypersensitivity to Esophageal Acid Infusion in Healthy Volunteers Yang C. Chua, Kee Seong Ng, Jafar Jafari, Abhishek Sharma, Susan Surguy, Etsuro Yazaki, Peter O. Julu, Charles H. Knowles, Qasim Aziz
Attendance and Diagnostic Yield of One Versus Two-Sample Fecal Immunochemical Test (FIT) Screening; a Comparative Population-Based Colorectal Cancer Trial Aafke H. van Roon, Janneke Wilschut, Marjolein van Ballegooijen, Laura J. Kranenburg, Anneke van Vuuren, Alexandra C. van der Togt-van Leeuwen, Jacqueline C. Reijerink, J. D. Habbema, Ernst J. Kuipers, Monique E. van Leerdam
Background: Esophageal acid infusion in humans induces primary and secondary hyperalgesia due to peripheral and central sensitization respectively (Sarkar 2001). Pregabalin is a centrally-acting modulator of voltage-sensitive calcium channels which reduces neuropathic pain by attenuating the release of multiple neurotransmitters (Taylor 2007) in the CNS. Its efficacy in reducing acid-induced esophageal hypersensitivity has not been explored. Aim: To test the effects of pregabalin on the development of acid-induced esophageal VPH in healthy human subjects. Methods: Single-centre, placebo-controlled, double-blind, randomized, two-period, cross-over study of 15 healthy volunteers (age 21-52 years, 9 males) attending on 3 separate visits. On visit 1, after esophageal manometry, pain thresholds (PTs) to esophageal electrical stimulation were determined using bipolar ring electrodes positioned in the proximal and distal esophagus. Thereafter, a 30-min infusion of HCL was performed in the distal esophagus.PT was measured again at both sites at 30 and 90mins after the acid infusion. The same methods were repeated for 3 visits. Participants were then given either pregabalin or placebo after visits 1 and 2.The intervals were at least;1 week between visits 1 and 2, and 2 weeks between visits 2 and 3 (to allow time for drug washout). The dose of pregabalin used was 75mg bd for 3 days, 150mg bd for 1 day and 150mg od. Results: The median decrease in PT at 30mins after acid, compared to baseline in the proximal oesophagus was significantly less after treatment with pregabalin. This pattern was maintained even at 90mins after acid, but to a lesser degree (Re: Table).The subjective score of pain to acid infusion was also reduced while on placebo compared to pregabalin (VAS median of 3 vs.1, p= 0.027). The baseline PTs before acid infusion at placebo and Pregabalin visits were not significantly different (p=0.42).There were no differences in the change in postacid PT between pregabalin and placebo in the distal esophagus. Conclusion: Pregabalin prevents the development of secondary hyperalgesia in the proximal esophagus after distal esophageal acidification, both on objective and subjective assessments of pain. It has no significant effect on the site of acid exposure, indicating that it acts to reduce central sensitization and is not analgesic. Further studies of pregabalin in patients with acid-induced esophageal hypersensitivity are now warranted. Table: Median change in PT in proximal oesophagus after acid infusion
Introduction: The fecal immunochemical test (FIT) is superior to the guaiac-based fecal occult blood test for the detection of colonic advanced neoplasia. However, comparisons on the optimal number of FITs performed per screening round are lacking. We therefore conducted a population-based colorectal cancer (CRC) screening trial comparing attendance and diagnostic yield of one versus two-sample FIT (1-FIT vs. 2-FITs). Methods: A representative sample of the Dutch population (n=5,693) aged 50-75 years was randomly selected prior to invitation. Individuals were invited by mail to participate in a first CRC screening round. Subjects of the reference group (n=2,493) received 1-FIT and all others (n=3,200) received 2-FITs (OC-Sensor Micro). Invitees from the 2-FITs group were given instructions to collect fecal samples of two consecutive bowel movements. 2-FIT screening was considered positive if at least one test was positive (cut-off value 75 ng Hb/mL). Results: After excluding 239 individuals (4%) who either met exclusion criteria, had died, or moved away, the attendance rate was 64.4% in the reference group (1,539/2,390; 95% CI 62.5-66.3%), and 61.2% in the 2-FITs group (1,874/3,064; CI 60.2-62.2%, p-value 0.01). The positivity rate in the reference group was 6.9% versus in the 2-FITs group 10.7% for at least one positive test, and 3.7% for two positive tests. The detection rate for advanced neoplasia was respectively 2.9%, 3.7% and 2.0% (see table). In a multivariate logistic regression analysis, age > 60 years (OR 1.4; CI 1.17-1.58), female gender (OR 1.2; CI 1.01-1.36), and high socioeconomic status (OR 1.4; CI 1.13-1.58) were associated with higher attendance rates in both groups. Conclusion: Double FIT-screening on two consecutive bowel movements has a slightly lower attendance than single FIT. 2-FIT screening (≥ 1 positive test) detects more advanced neoplasia compared to screening with only 1-FIT. How the trade-off works out between a higher sensitivity, lower specificity, and higher costs in terms of cost-effectiveness has yet to be determined.
AGA Abstracts
S-134
AGA Abstracts
930 Comparing Three Different Strategies of Double Sampling by Fecal Immunochemical Tests for Detection of Advanced Colorectal Neoplasm's Sietze T. Van Turenhout, Frank A. Oort, Veerle M. Coupe, René W. van der Hulst, Eric C. Wesdorp, Ilhame Ben Larbi, Shannon L. Kanis, Edwin van Hengel, Anneke A. Bouman, Gerrit A. Meijer, Chris J. Mulder
932 Fecal MicroRNAs: Novel Biomarkers for Colon Cancer Screening Alexander Link, Francesc Balaguer, Yan Shen, Takeshi Nagasaka, Juan José Lozano, C. R. Boland, Ajay Goel
Background Fecal Immunochemical Tests (FITs) are widely used for Colorectal Cancer (CRC) screening. Some sampling schemes use one-day FIT testing, whereas others use twoday FIT testing. Data on sensitivity and specificity of two-day FIT testing are lacking. This study aims to compare the difference in sensitivity and specificity of three different strategies of two-day FIT sampling, at different cut-off values. Methods In three hospitals in the Amsterdam area, all subjects ≥18 years who were referred for elective colonoscopy were asked to perform a FIT (OC sensor®) on two consecutive days. Colonoscopy and histology were considered as gold standard. Two-day FIT sampling was defined positive in three different ways: 1) when at least one of two FITs was above the cut-off value (FIT+), 2) when both FITs were above the cut-off value(FIT++), and 3) when the geometric mean of two FITs was above the cut-off value (FITmean). Test performance of all three strategies was evaluated at cut-off values of 50, 75 and 100ng/ml. Results In 1105 subjects with complete colonoscopy, 140 (9,4%) AN were found (34 CRCs and 106 advanced adenomas). Of the CRC cases, 70% were Dukes stage A or B (stage unknown in 2). Positivity rates for FIT+ ranged from 16-23%, for FIT++ from 10-13% and for FITmean from 13-17% (see table). At any cut-off level, FIT++ resulted in the lowest and FIT+ in the highest sensitivity. FIT++ showed a higher specificity than both other strategies (see table). Conclusions The most sensitive strategy in two-day FIT sampling is defining the test positive when at least one of both FITs passes the cut off (FIT+), whereas the highest specificity is obtained when both FITs need to be positive for calling the tests positive (FIT++). At all cut-off values, FITmean offers substantially better specificity than FIT+, while sensitivity is substantially higher than FIT++. Sensitivity and specificity of three methods to use two-day FIT sampling for the detection of advanced neoplasia
Introduction: Colorectal cancer (CRC) is the second leading cause of cancer-related deaths in the United States. Although screening can reduce cancer-related mortality, the noninvasive screening programs have achieved only a modest decrease in mortality. MicroRNAs (miRNAs) play important roles in a wide array of biological processes by regulating gene expression. Expression patterns of miRNAs are commonly dysregulated in neoplasia and unique miRNA expression patterns allow discrimination between various types of cancers. In this work, we evaluated the feasibility of fecal miRNAs as biomarkers for colorectal neoplasia screening. Material and methods: Total RNA was extracted from feces using Qiagen miRNAeasy Mini Kits with minor modifications. Illumina microRNA microarray profiling was performed to evaluate the differences in miRNA expression patterns between normal colonic mucosa tissues and stool samples from healthy subjects. Quantitative RTPCR was performed for a subset of miRNAs (including miR-21, -16, -26b, -17 and -29b) in stool samples from 8 healthy volunteers. We also determined miRNA expression in stool samples collected with FOBT kits from 29 subjects with normal colonoscopies (n=10), adenomas (n=9) or CRCs (n=10). Results: We efficiently extracted miRNAs from stool specimens using our modified protocol. Analysis of two independent extractions showed high reproducibility for miRNA extraction and expression. miRNA expression patterns were similar in stool specimens among healthy volunteers, and the pattern was reproducible in stool samples that were collected serially in time from the same individuals. miRNA expression profiles from stool samples collected with FOBT kits from 29 patients demonstrated significantly higher expression of miR-21 and miR-106a in patients with adenomas and CRCs, compared with individuals free of colorectal neoplasia (miR-21, p<0.02; and miR-106a, p<0.05). Conclusion: Our data indicate that miRNAs can be extracted from stool easily and reproducibly. The stools of patients with colorectal neoplasms have unique and identifiable patterns of miRNA expression. These findings suggest that fecal miRNA analysis is an excellent candidate for the development of a noninvasive screening test for colorectal neoplasia. 933
931
Pregabalin Prevents Development of Esophageal Hypersensitivity to Esophageal Acid Infusion in Healthy Volunteers Yang C. Chua, Kee Seong Ng, Jafar Jafari, Abhishek Sharma, Susan Surguy, Etsuro Yazaki, Peter O. Julu, Charles H. Knowles, Qasim Aziz
Attendance and Diagnostic Yield of One Versus Two-Sample Fecal Immunochemical Test (FIT) Screening; a Comparative Population-Based Colorectal Cancer Trial Aafke H. van Roon, Janneke Wilschut, Marjolein van Ballegooijen, Laura J. Kranenburg, Anneke van Vuuren, Alexandra C. van der Togt-van Leeuwen, Jacqueline C. Reijerink, J. D. Habbema, Ernst J. Kuipers, Monique E. van Leerdam
Background: Esophageal acid infusion in humans induces primary and secondary hyperalgesia due to peripheral and central sensitization respectively (Sarkar 2001). Pregabalin is a centrally-acting modulator of voltage-sensitive calcium channels which reduces neuropathic pain by attenuating the release of multiple neurotransmitters (Taylor 2007) in the CNS. Its efficacy in reducing acid-induced esophageal hypersensitivity has not been explored. Aim: To test the effects of pregabalin on the development of acid-induced esophageal VPH in healthy human subjects. Methods: Single-centre, placebo-controlled, double-blind, randomized, two-period, cross-over study of 15 healthy volunteers (age 21-52 years, 9 males) attending on 3 separate visits. On visit 1, after esophageal manometry, pain thresholds (PTs) to esophageal electrical stimulation were determined using bipolar ring electrodes positioned in the proximal and distal esophagus. Thereafter, a 30-min infusion of HCL was performed in the distal esophagus.PT was measured again at both sites at 30 and 90mins after the acid infusion. The same methods were repeated for 3 visits. Participants were then given either pregabalin or placebo after visits 1 and 2.The intervals were at least;1 week between visits 1 and 2, and 2 weeks between visits 2 and 3 (to allow time for drug washout). The dose of pregabalin used was 75mg bd for 3 days, 150mg bd for 1 day and 150mg od. Results: The median decrease in PT at 30mins after acid, compared to baseline in the proximal oesophagus was significantly less after treatment with pregabalin. This pattern was maintained even at 90mins after acid, but to a lesser degree (Re: Table).The subjective score of pain to acid infusion was also reduced while on placebo compared to pregabalin (VAS median of 3 vs.1, p= 0.027). The baseline PTs before acid infusion at placebo and Pregabalin visits were not significantly different (p=0.42).There were no differences in the change in postacid PT between pregabalin and placebo in the distal esophagus. Conclusion: Pregabalin prevents the development of secondary hyperalgesia in the proximal esophagus after distal esophageal acidification, both on objective and subjective assessments of pain. It has no significant effect on the site of acid exposure, indicating that it acts to reduce central sensitization and is not analgesic. Further studies of pregabalin in patients with acid-induced esophageal hypersensitivity are now warranted. Table: Median change in PT in proximal oesophagus after acid infusion
Introduction: The fecal immunochemical test (FIT) is superior to the guaiac-based fecal occult blood test for the detection of colonic advanced neoplasia. However, comparisons on the optimal number of FITs performed per screening round are lacking. We therefore conducted a population-based colorectal cancer (CRC) screening trial comparing attendance and diagnostic yield of one versus two-sample FIT (1-FIT vs. 2-FITs). Methods: A representative sample of the Dutch population (n=5,693) aged 50-75 years was randomly selected prior to invitation. Individuals were invited by mail to participate in a first CRC screening round. Subjects of the reference group (n=2,493) received 1-FIT and all others (n=3,200) received 2-FITs (OC-Sensor Micro). Invitees from the 2-FITs group were given instructions to collect fecal samples of two consecutive bowel movements. 2-FIT screening was considered positive if at least one test was positive (cut-off value 75 ng Hb/mL). Results: After excluding 239 individuals (4%) who either met exclusion criteria, had died, or moved away, the attendance rate was 64.4% in the reference group (1,539/2,390; 95% CI 62.5-66.3%), and 61.2% in the 2-FITs group (1,874/3,064; CI 60.2-62.2%, p-value 0.01). The positivity rate in the reference group was 6.9% versus in the 2-FITs group 10.7% for at least one positive test, and 3.7% for two positive tests. The detection rate for advanced neoplasia was respectively 2.9%, 3.7% and 2.0% (see table). In a multivariate logistic regression analysis, age > 60 years (OR 1.4; CI 1.17-1.58), female gender (OR 1.2; CI 1.01-1.36), and high socioeconomic status (OR 1.4; CI 1.13-1.58) were associated with higher attendance rates in both groups. Conclusion: Double FIT-screening on two consecutive bowel movements has a slightly lower attendance than single FIT. 2-FIT screening (≥ 1 positive test) detects more advanced neoplasia compared to screening with only 1-FIT. How the trade-off works out between a higher sensitivity, lower specificity, and higher costs in terms of cost-effectiveness has yet to be determined.
AGA Abstracts
S-134