Early cancer detection significantly increases a patient’s chances of a positive outcome and can reduce treatment costs. However, current cancer screening tools face challenges with sensitivity, specificity and high sequencing costs. Due to this, no single multi-cancer detection test has yet been established for widespread use. A report of a new multi-modal approach, currently in pre-print on elifesciences, uses circulating tumour DNA (ctDNA) to allow not only the early detection of multiple cancer types, but to also provide information about the tumour’s location and stage, all at lower costs than previously thought possible.
Spotting a tumour
Despite their enormous potential, previously published ctDNA-based liquid biopsy assays have centred around high-depth sequencing (30X), resulting in high costs and low accessibility for use in widespread screening. A new assay developed by collaborative researchers at Gene Solutions and the Medical Genetics Institute in Vietnam, known as SPOT-MAS (Screening for the Presence of Tumour by Methylation and Size), simultaneously profiles methylomics, fragmentomics, copy number and circulating free DNA (cfDNA) end motifs in a single workflow (Figure 1). By combining several areas of focus, SPOT-MAS uses targeted, shallow depth (0.55X) genome-wide sequencing in combination with innovative machine-learning algorithms to reduce costs while delivering comparable results to other, more expensive assays.
Figure 1. Three step workflow of the SPOT-MAS assay for multi-cancer detection and localization.
Firstly, cfDNA is isolated from peripheral blood. Bisulfite conversion and adapter ligation is used to make a whole-genome bisulfite cfDNA library which is then subjected to hybridization by probes specific for 450 target regions to collect the target capture fraction. The whole-genome fraction is retrieved by collecting the ‘flow-through’, then hybridized with probes specific for adapter sequences of the DNA library. Both the target capture and whole-genome fractions are subjected to massive parallel sequencing and the resulting data is pre-processed into five different features of cfDNA: Target methylation (TM), genome-wide methylation (GWM), fragment length profile (Flen), DNA copy number (CNA) and end motif (EM). Finally, machine learning models and graph convolutional neural networks are adopted for classification of cancer status and identification of the tissue of origin.
Adapted from Nguyen et al., 2023.
Sensitive and specific
As ctDNA is a carrier of genetic and epigenetic modifications from cancer-derived DNA, SPOT-MAS is highly sensitive and specific, even allowing identification of the tissue of origin for a primary tumour. Researchers applied SPOT-MAS to blood samples from 738 treatment-naïve patients with non-metastatic breast, colorectal, gastric, lung or liver tumours, and 1,550 healthy controls. The five cancer types were detected with average 72.4% sensitivity and 97.0% specificity, with 62.3% and 73.9% sensitivity at stages I and II respectively. Moreover, the tissue-of-origin accuracy score was 0.7 in independent validation cohorts using graph convolutional neural network models.
The highest sensitivity and specificity detection rates were found in liver cancer patients, at 89.6% and 91.1% respectively, in line with previous reports of high levels of ctDNA shedding by liver tumours. Conversely, the detection of breast cancer, particularly at early stage, is notoriously challenging due to low levels of ctDNA shedding and high heterogeneity of breast cancer subtypes. Using SPOT-MAS, breast cancer had the lowest detection rates out of the five cancer types at 58.3%. However, this cancer type had the highest level of tumour-of-origin accuracy at 0.8, meaning SPOT-MAS may be a useful tool to identify a potential tumour location when a generalised cancer diagnosis is suspected.
Diagnosis before symptoms
Development of liquid biopsy screening tests which can accurately detect and localise cancer at an early stage could revolutionise cancer diagnosis and improve patient outcomes. The SPOT-MAS assay demonstrates comparable performance to other ctDNA based assays, but the shallow sequencing required significantly reduces costs. This may mean that the use of SPOT-MAS could be a feasible option for widespread, multi-cancer screening of at-risk populations in future. Further testing on other cancer types and on larger patient cohorts, alongside consideration of variables such as patient age, is currently ongoing. Preliminary data from interim reports is encouraging, even demonstrating the ability of SPOT-MAS to detect cancer in patients who exhibited no symptoms at the time of testing.
