Sensitive Detection of Colorectal Cancer in Peripheral Blood by Septin 9 DNA Methylation Assay

Abstract

BackgroundColorectal cancer (CRC) is the second leading cause of cancer deaths despite the fact that detection of this cancer in early stages results in over 90% survival rate. Currently less than 45% of at-risk individuals in the US are screened regularly, exposing a need for better screening tests. We performed two case-control studies to validate a blood-based test that identifies methylated DNA in plasma from all stages of CRC.Methodology/Principal FindingsUsing a PCR assay for analysis of Septin 9 (SEPT9) hypermethylation in DNA extracted from plasma, clinical performance was optimized on 354 samples (252 CRC, 102 controls) and validated in a blinded, independent study of 309 samples (126 CRC, 183 controls). 168 polyps and 411 additional disease controls were also evaluated. Based on the training study SEPT9-based classification detected 120/252 CRCs (48%) and 7/102 controls (7%). In the test study 73/126 CRCs (58%) and 18/183 control samples (10%) were positive for SEPT9 validating the training set results. Inclusion of an additional measurement replicate increased the sensitivity of the assay in the testing set to 72% (90/125 CRCs detected) while maintaining 90% specificity (19/183 for controls). Positive rates for plasmas from the other cancers (11/96) and non-cancerous conditions (41/315) were low. The rate of polyp detection (>1 cm) was ∼20%.Conclusions/SignificanceAnalysis of SEPT9 DNA methylation in plasma represents a straightforward, minimally invasive method to detect all stages of CRC with potential to satisfy unmet needs for increased compliance in the screening population. Further clinical testing is warranted.

Advances in epigenetic biomarker research in colorectal cancer

“…Bisulfite treatment and methylation specific polymerase chain reaction (PCR) are the two most commonly used techniques. A blood biomarker with a high sensitivity and specificity for CRC can not only be used to segregate high-risk patients for further clinical tests but also be an excellent tool for monitoring CRC recurrence in patients who have undergone tumor resection (Table 1) [20][21][22][23][24][25][26][27][28][29][30] .…”

“…The v2 region of the Septin 9 (SEPT9) promoter has been shown to be methylated in CRC tissue compared with normal colonic mucosa. Using highly sensitive realtime PCR assays, methylated SEPT9 was first detected in the plasma of CRC patients with an overall sensitivity of 72% and a specificity of 90% [24] . Significant validation has been performed for this methylation biomarker, and Warren et al [27] have confirmed a sensitivity of up to 90% and a specificity of up to 88% for SEPT9.…”

A novel gene expression signature in peripheral blood mononuclear cells for early detection of colorectal cancer

“…Several attempts have been made in the past to search for tumour markers in the blood to develop screening tests for CRC. [7][8][9] However, validation data obtained from a large screening-eligible population are still missing or showed mitigated test performances. 10 Searching for blood-borne tumour markers could leverage different concepts such as the release of tumour-derived molecules (proteins, nucleic acids) into the blood stream, the presence of circulating tumour cells or the generation of a host response to tumour-derived signals.…”

Epi proColon®: Use of a Non-Invasive SEPT9 Gene Methylation Blood Test for Colorectal Cancer Screening: A National Laboratory Experience

“…Epigenetic silencing of the SEPT9 gene by promoter methylation in cell-free plasma has been shown to be a biomarker for CRC [12,16,17]. Using high-sensitivity Real-Time PCR methods, hypermethylated SEPT9 DNA signatures can be detected in cellfree plasma [12,[19][20][21][22][23][24][25][26][27]. As an early event and hallmark of human cancers, aberrant DNA methylation biomarkers have clinical roles in screening, diagnosis, prognosis and therapeutic response [11,28].…”