DEEPGENTM—A Novel Variant Calling Assay for Low Frequency Variants

Abstract

Detection of genetic variants in clinically relevant genomic hot-spot regions has become a promising application of next-generation sequencing technology in precision oncology. Effective personalized diagnostics requires the detection of variants with often very low frequencies. This can be achieved by targeted, short-read sequencing that provides high sequencing depths. However, rare genetic variants can contain crucial information for early cancer detection and subsequent treatment success, an inevitable level of background noise usually limits the accuracy of low frequency variant calling assays. To address this challenge, we developed DEEPGENTM, a variant calling assay intended for the detection of low frequency variants within liquid biopsy samples. We processed reference samples with validated mutations of known frequencies (0%–0.5%) to determine DEEPGENTM’s performance and minimal input requirements. Our findings confirm DEEPGENTM’s effectiveness in discriminating between signal and noise down to 0.09% variant allele frequency and an LOD(90) at 0.18%. A superior sensitivity was also confirmed by orthogonal comparison to a commercially available liquid biopsy-based assay for cancer detection.

Blood-Based Multi-Cancer Detection Using a Novel Variant Calling Assay (DEEPGENTM): Early Clinical Results

“…This finding was observed for a "full model"-including all assessed cancers as well as organ-specific models with varying ranges of performance for the different cancers. As such, it seems plausible to assume that the described liquid biopsy and data analytics innovations not only transpire to an excellent technical performance [23], but also into a relevant clinical capability. This finding is in line with similar projects in the field of liquid biopsy using different technological approaches [7,21,22,26].…”

“…The platform addresses the common challenges of previous generation liquid biopsy platforms by combining yield-optimized cfDNA storage and extraction protocols, a broad panel assay, advanced error reduction chemistry, and ultradeep next-generation sequencing (NGS) with a customized bioinformatics pipeline, highly scalable cloud software and innovative AI techniques to identify low-frequency variants accurately and consistently in the blood. Initial validation studies demonstrated that DEEPGEN TM has high detection capabilities, capturing over 3000 mutations at VAFs down to >0.09%, and thus out-performs other publicly available platforms on a technical level [23]. Additionally, the mutations undergo a proprietary machine learning protocol that considers mutations and their VAFs to derive a prediction if cancer might be the underlying cause of the observation.…”

Opportunities for Early Cancer Detection: The Rise of ctDNA Methylation-Based Pan-Cancer Screening Technologies

“…The authors have used a bioinformatics to identify a broad panel of cancer-specific genomic variants that are present at very low frequency in blood. The assay targets 3062 of these mutant fragments (including single nucleotides polymorphisms, multi nucleotide polymorphism, and short insertion or deletion) using proprietary primers for advanced PCR and next-generation sequencing analysis, resulting in high capture efficiency at VAF down to >0.09% [ 62 ]. DEEPGEN™ has been used in an initial comprehensive prospective study for the detection of seven different types of cancers (bladder, prostate, lung, liver, pancreatic, colorectal, and breast) using a cohort of 260 cancer samples across stages I-IV (27% stage I, 21% stage II, 29% stage III, 10% stage IV and 13% undetermined) and 415 healthy controls.…”

Shifting the Cancer Screening Paradigm: The Rising Potential of Blood-Based Multi-Cancer Early Detection Tests

“…Interestingly, a study evaluating patients’ perceptions about stool-based multi-cancer detection reported that 98% of participants would use such a test, preferring it over conventional colorectal cancer screening, and highlighted its pan-cancer feature as the most relevant [ 25 ]. Subsequently, Quantgene Inc. developed DEEPGEN TM , a blood test based on next-generation sequencing (NGS) that detects low-frequency genetic abnormalities at a variant allele frequency of 0.09% [ 26 , 27 ]. When applied to the detection of seven cancer types, this assay displayed 43% sensitivity at 99% specificity with an area under ROC curve (AUC) of 0.90.…”