Evaluation of an assay for methylated BCAT1 and IKZF1 in plasma for detection of colorectal neoplasia

Abstract

BackgroundSpecific genes, such as BCAT1 and IKZF1, are methylated with high frequency in colorectal cancer (CRC) tissue compared to normal colon tissue specimens. Such DNA may leak into blood and be present as cell-free circulating DNA. We have evaluated the accuracy of a novel blood test for these two markers across the spectrum of benign and neoplastic conditions encountered in the colon and rectum.MethodsCirculating DNA was extracted from plasma obtained from volunteers scheduled for colonoscopy for any reason, or for colonic surgery, at Australian and Dutch hospitals. The extracted DNA was bisulphite converted and analysed by methylation specific real-time quantitative PCR (qPCR). A specimen was deemed positive if one or more qPCR replicates were positive for either methylated BCAT1 or IKZF1 DNA. Sensitivity and specificity for CRC were estimated as the primary outcome measures.ResultsPlasma samples were collected from 2105 enrolled volunteers (mean age 62 years, 54 % male), including 26 additional samples taken after surgical removal of cancers. The two-marker blood test was run successfully on 2127 samples. The test identified 85 of 129 CRC cases (sensitivity of 66 %, 95 % CI: 57–74). For CRC stages I-IV, respective positivity rates were 38 % (95 % CI: 21–58), 69 % (95 % CI: 53–82), 73 % (95 % CI: 56–85) and 94 % (95 % CI: 70–100). A positive trend was observed between positivity rate and degree of invasiveness. The colonic location of cancer did not influence assay positivity rates. Gender, age, smoking and family history were not significant predictors of marker positivity. Twelve methylation-positive cancer cases with paired pre- and post-surgery plasma showed reduction in methylation signal after surgery, with complete disappearance of signal in 10 subjects. Sensitivity for advanced adenoma (n = 338) was 6 % (95 % CI: 4–9). Specificity was 94 % (95 % CI: 92–95) in all 838 non-neoplastic pathology cases and 95 % (95 % CI: 92–97) in those with no colonic pathology detected (n = 450).ConclusionsThe sensitivity for cancer of this two-marker blood test justifies prospective evaluation in a true screening population relative to a proven screening test. Given the high rate of marker disappearance after cancer resection, this blood test might also be useful to monitor tumour recurrence.Trial registrationACTRN12611000318987.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1674-2) contains supplementary material, which is available to authorized users.

Diagnostic and prognostic role of cell‐free DNA testing for colorectal cancer patients

“…To date, many studies had reported aberrant methylation in tissue of colon cancer, while relatively few reports studied CRC plasma methylation biomarkers, e. g. APC and RASSF1 , BCAT1 and IKZF1 , AGBL4 , FLI1 , and TWIST1 , PCDH10 , THBD and C9orf50 and CDKN2A genes, with exception of the aberrant methylation of SEPT9 gene. This biomarker is currently included in non‐invasive commercial kit called Epi proColon® by Epigenomics AG (Germany, Berlin), the first and only FDA‐approved blood‐based colorectal cancer screening test.…”

Detection of colorectal neoplasia: Combination of eight blood-based, cancer-associated protein biomarkers

“…The eight protein biomarkers were chosen due to their association with GI malignancy. Additional blood‐based biomarkers such as genomics, epigenomics and metabolomics are currently under intense evaluation in various clinical studies. Methylated genes such as SEPT9 may be valuable in early detection of colorectal neoplasia .…”

Future perspectives of circulating tumor DNA in colorectal cancer

“…[61][62][63][64][65][66][67][68][69] For example, an approach using a BCAT1 and IKZF1 methylation assay identified approximately 70% of CRC. [75][76][77] Detection of tumor recurrence, minimal residual disease, and prognosis value Surgery alone cures a large fraction of patients with localized CRC. However, there are currently no effective means to identify which patients have residual disease that will result in recurrence.…”