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Novel Approach to Fecal Occult Blood Testing by Assay of Erythrocyte-Specific microRNA Markers

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Abstract

Background

Fecal occult blood testing (FOBT) has historically relied on methods to detect hemoglobin with no fundamental innovations in decades.

Aim

To examine microRNA (miRNA) as a new marker class for FOBT.

Methods

Candidate miRNA markers were identified by small RNA sequencing of human whole blood compared to colorectal epithelia. Markers were tested in human blood cell subsets and blood from non-human species. We assessed assay linearity in blood spiking and marker stability in stool over incubation experiments. Levels of candidate erythrocyte markers were explored in stools from colorectal cancer (CRC) cases and controls.

Results

Based on small RNA sequencing and validation RT-qPCR, expression level of each of the top blood-enriched markers (hsa-miR-144-3p, 144-5p, 451a, 486-5p, 363-3p, 20b-5p) could perfectly discriminate blood from colorectal epithelia. All six markers arose from and showed specificity to human erythrocytes. Marker levels increased linearly with erythrocyte concentration in saline or stool and demonstrated a broader dynamic range than did immunochemical test for hemoglobin. Degradation of markers occurred in stool but was reduced with preservative buffers. Erythrocyte marker candidates for stool testing were selected in an exploratory set of stools (20 CRC, 40 normal). Candidates were then further tested in a feasibility set (29 CRC, 31 advanced adenoma, and 115 normal); a miRNA panel (hsa-miR-451a, 144-5p, and 200b-3p as normalizer) yielded an AUC of 0.89 (95% CI 0.82–0.95, P < .0001) for CRC.

Conclusions

A novel miRNA-based approach accurately quantifies fecal blood levels over a broad, clinically relevant range.

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References

  1. Mandel JS, Bond JH, Church TR, et al. Reducing mortality from colorectal cancer by screening for fecal occult blood. Minnesota colon cancer control study. N Engl J Med. 1993;328:1365–1371.

    Article  CAS  PubMed  Google Scholar 

  2. Mandel JS, Church TR, Ederer F, Bond JH. Colorectal cancer mortality: effectiveness of biennial screening for fecal occult blood. J Natl Cancer Inst. 1999;91:434–437.

    Article  CAS  PubMed  Google Scholar 

  3. Ahlquist DA, Young GP. Approach to the patient with occult gastrointestinal bleeding. In: Yamada T, Alpers DH, eds. Principles of clinical gastroenterology. New York: Wiley; 2008:152–169.

    Chapter  Google Scholar 

  4. Boas I. Die lehre von den okkulten blutungen. Leipzig: G. Thieme; 1914.

    Google Scholar 

  5. Barrows GH, Burton RM, Jarrett DD, Russell GG, Alford MD, Songster CL. Immunochemical detection of human blood in feces. Am J Clin Pathol. 1978;69:342–346.

    Article  CAS  PubMed  Google Scholar 

  6. Harewood GC, McConnell JP, Harrington JJ, Mahoney DW, Ahlquist DA. Detection of occult upper gastrointestinal tract bleeding: performance differences in fecal occult blood tests. Mayo Clin Proc. 2002;77:23–28.

    Article  PubMed  Google Scholar 

  7. Rockey DC, Auslander A, Greenberg PD. Detection of upper gastrointestinal blood with fecal occult blood tests. Am J Gastroenterol. 1999;94:344–350.

    Article  CAS  PubMed  Google Scholar 

  8. Ahlquist DA, Zou H, Domanico M, et al. Next-generation stool DNA test accurately detects colorectal cancer and large adenomas. Gastroenterology. 2012;142:248–256. (quiz e225–246).

    Article  CAS  PubMed  Google Scholar 

  9. Imperiale TF, Ransohoff DF, Itzkowitz SH, et al. Multitarget stool DNA testing for colorectal-cancer screening. N Engl J Med. 2014;370:1287–1297.

    Article  CAS  PubMed  Google Scholar 

  10. Lidgard GP, Domanico MJ, Bruinsma JJ, et al. Clinical performance of an automated stool DNA assay for detection of colorectal neoplasia. Clin Gastroenterol Hepatol. 2013;11:1313–1318.

    Article  CAS  PubMed  Google Scholar 

  11. Redwood DG, Asay ED, Blake ID, et al. Stool DNA testing for screening detection of colorectal neoplasia in Alaska Native people. Mayo Clin Proc. 2016;91:61–70.

    Article  CAS  PubMed  Google Scholar 

  12. Ahlquist DA, McGill DB, Fleming JL, et al. Patterns of occult bleeding in asymptomatic colorectal cancer. Cancer. 1989;63:1826–1830.

    Article  CAS  PubMed  Google Scholar 

  13. Heigh RI, Yab TC, Taylor WR, et al. Detection of colorectal serrated polyps by stool DNA testing: comparison with fecal immunochemical testing for occult blood (FIT). PLoS ONE. 2014;9:e85659.

    Article  PubMed  PubMed Central  Google Scholar 

  14. Bibbins-Domingo K, Grossman DC, Curry SJ, et al. Screening for colorectal cancer: US preventive services task force recommendation statement. JAMA. 2016;315:2564–2575.

    Article  CAS  PubMed  Google Scholar 

  15. El-Shami K, Oeffinger KC, Erb NL, et al. American cancer society colorectal cancer survivorship care guidelines. CA Cancer J Clin. 2015;65:428–455.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Arroyo JD, Chevillet JR, Kroh EM, et al. Argonaute2 complexes carry a population of circulating microRNAs independent of vesicles in human plasma. Proc Natl Acad Sci USA. 2011;108:5003–5008.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Wu CW, Ng SC, Dong Y, et al. Identification of microRNA-135b in stool as a potential noninvasive biomarker for colorectal cancer and adenoma. Clin Cancer Res. 2014;20:2994–3002.

    Article  CAS  PubMed  Google Scholar 

  18. Friedlander MR, Mackowiak SD, Li N, Chen W, Rajewsky N. miRDeep2 accurately identifies known and hundreds of novel microRNA genes in seven animal clades. Nucleic Acids Res. 2012;40:37–52.

    Article  PubMed  Google Scholar 

  19. DeLong ER, DeLong DM, Clarke-Pearson DL. Comparing the areas under two or more correlated receiver operating characteristic curves: a nonparametric approach. Biometrics. 1988;44:837–845.

    Article  CAS  PubMed  Google Scholar 

  20. Schwarzenbach H, da Silva AM, Calin G, Pantel K. Data normalization strategies for microRNA quantification. Clin Chem. 2015;61:1333–1342.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Kirschner MB, Edelman JJ, Kao SC, Vallely MP, van Zandwijk N, Reid G. The impact of hemolysis on cell-free microRNA. Biomark Front Genet. 2013;4:94.

    CAS  Google Scholar 

  22. Pritchard CC, Kroh E, Wood B, et al. Blood cell origin of circulating microRNAs: a cautionary note for cancer biomarker studies. Cancer Prev Res. 2012;5:492–497.

    Article  CAS  Google Scholar 

  23. Wu CW, Ng SS, Dong YJ, et al. Detection of miR-92a and miR-21 in stool samples as potential screening biomarkers for colorectal cancer and polyps. Gut. 2012;61:739–745.

    Article  CAS  PubMed  Google Scholar 

  24. Dore LC, Amigo JD, Dos Santos CO, et al. A GATA-1-regulated microRNA locus essential for erythropoiesis. Proc Natl Acad Sci USA. 2008;105:3333–3338.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Rasmussen KD, Simmini S, Abreu-Goodger C, et al. The miR-144/451 locus is required for erythroid homeostasis. J Exp Med. 2010;207:1351–1358.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Park DI, Ryu S, Kim YH, et al. Comparison of guaiac-based and quantitative immunochemical fecal occult blood testing in a population at average risk undergoing colorectal cancer screening. Am J Gastroenterol. 2010;105:2017–2025.

    Article  PubMed  Google Scholar 

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Acknowledgments

Research support was obtained from Eugene and Eva Lane, Mayo Clinic, Clinical Core of Mayo Clinic Center for Cell Signaling in Gastroenterology (P30DK084567) and Exact Sciences.

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Authors and Affiliations

  1. Division of Gastroenterology and Hepatology, Mayo Clinic, 200 First Street SW, Rochester, MN, 55905, USA

    Chung Wah Wu, Xiaoming Cao, Calise K. Berger, Patrick H. Foote, Julie A. Simonson, Bradley W. Anderson, Tracy C. Yab, William R. Taylor, Lisa A. Boardman, John B. Kisiel & David A. Ahlquist

  2. Division of Biomedical Statistics and Informatics, Mayo Clinic, 200 First Street SW, Rochester, MN, 55905, USA

    Douglas W. Mahoney

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  1. Chung Wah Wu
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  2. Xiaoming Cao
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  3. Calise K. Berger
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  4. Patrick H. Foote
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  5. Douglas W. Mahoney
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  6. Julie A. Simonson
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  7. Bradley W. Anderson
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  8. Tracy C. Yab
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  9. William R. Taylor
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  10. Lisa A. Boardman
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  12. David A. Ahlquist
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Corresponding author

Correspondence to David A. Ahlquist.

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Conflict of interest

David A. Ahlquist is a scientific advisor to and research collaborator with Exact Sciences. Other authors declared no conflict of interests.

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Wu, C.W., Cao, X., Berger, C.K. et al. Novel Approach to Fecal Occult Blood Testing by Assay of Erythrocyte-Specific microRNA Markers. Dig Dis Sci 62, 1985–1994 (2017). https://doi.org/10.1007/s10620-017-4627-6

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  • DOI: https://doi.org/10.1007/s10620-017-4627-6

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