Detecting Colorectal Cancer
Blog Nov 28, 2013
VolitionRx presented their first study data at the recent Circulating Nucleic Acids in Plasma and Serum congress in Baltimore. We spoke to coordinator of the trial, Dr Stefan Holdenrieder, to understand more about circulating nucleosomes and the use of VolitionRx's NuQ assays in detecting colorectal cancer.
AB: Could you explain the importance of focusing on circulating nucleosomes in colorectal cancer detection?
Stefan Holdenrieder (SH): Nucleosomes themselves are highly interesting analytes as they are complexes of protein components, the so-called histones, and associated nucleic acids.
Our early studies showed that the amount of nucleosomes is increased in sera of patients with many cancer types, among them also in colorectal cancer, when compared with healthy individuals. This was very encouraging for us. Unfortunately, also patients with benign organ-related diseases, such as inflammations or non-tumor lesions, showed elevated levels limiting the diagnostic value of this parameter if taken as sole criterion. Although some acute diseases like inflammations could be ruled out by other biochemical parameters it seemed us to uncertain to use it as diagnostic tool.
However, we found that the courses of serial nucleosome values during a cytotoxic therapy were very well able to identify therapy response or disease progression. For some entities like in lung, pancreatic, liver and colorectal cancer, the serial nucleosome patterns during the first week of an anticancer treatment could already predict the later non-response or progression in a portion of the patients.
AB: Could you give us an outline of this study and the methods used?
SH: The NuQ-5mc test is one first example of a wide portfolio of VolitionRx’s new immunological assays that detect nucleosome-modifications that occur during the development of a tumor. While the NuQ-5mc test quantifies the methylation status of DNA, other assays measure characteristic groups that are added to regulatory sites of the histones such as acetyl or methyl groups; others measure typical histone variants or proteins that are attached to the nucleosomes and regulate transcription processes. The strategy is to combine several of these different approaches to identify a marker pattern that is highly sensitive and specific for tumor detection and monitoring.
In many tumors DNA becomes less methylated during the disease development making the DNA less stable and more vulnerable to DNA damages, thereby increasing the risk for generating genetic errors and mutations. The NuQ-5mc assay quantifies this loss of overall methylation and may reflect the risk of genetic instability.
AB: How accurate are these tests and how can this be improved?
SH: The quality of the assay was reflected by intra- and interassay reproducibilities of 3.4% and 15.3%, respectively. There was no major influence of preanalytic factors on analyte concentration.
In the clinical testing, the levels of circulating methylated DNA were significantly decreased in CRC and BCD when compared with HC, while there was no difference between BCD and CRC. When compared with healthy controls, the sensitivity for the detection of colorectal cancer was 75% at a specificity of 70%. At 95% specificity, the detection rate was 33%. This is in the range of PSA for prostate cancer screening. The low levels in BCD patients will have to be further investigated in larger cohorts to see whether NuQ-5mc detects also polyps that potentially could progress to colorectal cancer.
The assay will have to be validated in larger patient cohorts comprising relevant subgroups of patients with colorectal cancer in early and late stages as well as patients with colorectal polyps, inflammatory and other diseases. Then it can be seen whether already patients with early stage cancers disease and also those with high-cancer risk adenomatous polyps can be detected with high accuracy. Subsequently it will be tested in a real screening cohort, that is in patients under-going colonoscopy, in order to establish the positive and negative predictive values. Thereby the new assays will be tested in parallel with nowadays used screening markers, such as FOBT or FIT, to show the superiority to conventional markers. Most importantly, the assay will be further analyzed on its analytical and preanalytical properties. In particular it will be tested at various laboratory sites to show its stability, inter-laboratory and long-term comparability.
Then also other cancer and benign diseases, particularly those that can influence the metabolism of the analyte in the blood will have to be tested.
Finally, the assay will be combined with other nucleosome modification assays to further enhance the accuracy of cancer detection. This is important as due to the small portion of cancer cases in a screening cohort, the sensitivity and specificity should be considerably high, to catch most cancer patients and to avoid false-positive results.
In addition, we will also test the performance of the assay for the prediction and monitoring of the response to therapy and the estimation of prognosis in colorectal cancer patients.
The first results are very encouraging for us. We proceed now with further validation steps. If our results are also confirmed for early stage cancer and if the accuracy is enhanced by combination with further nucleosomics markers, they could become a valuable tool for prescreening individuals to identify those who are at risk and will have to undergo further diagnostics, e.g. colonoscopy. It will be important to show that the positive predictive value of the new assays is higher than that of currently used screening markers.
AB: What conclusions can be drawn from this research and how does the future of look for colorectal cancer?
SH: The decrease in mortality of colorectal cancer patients is a result of screening efforts in recent decades. Therefore, screening will probably remain a major focus in the future. Increasing the compliance of individuals to participate at screening programs is the main challenge nowadays. Therefore convenient and meaningful test systems that are cheap and give reliable results are needed. As blood is taken minimal-invasively, sensitive and specific blood assays are “hot” candidates to be introduced as prescreening tools to convince cautious individuals at risk to undergo colonoscopy.