Abstract 3992: Leveraging Ultima Genomics ppmSeq WGS-cfDNA to accurately detect clonal evolution over sequential blood biopsies

Abstract Comprehensively modeling tumor evolution is important for cancer diagnosis, treatment, and minimal residual disease (MRD) monitoring. Liquid biopsies enable non-invasive sampling of tumor DNA during a patient’s cancer treatment. Accurate detection of low allele fraction somatic variants in circulating tumor DNA (ctDNA) has clinical importance for cancer detection and monitoring. Current ctDNA panel sequencing methods lack the breadth of variants captured in whole genome sequencing (WGS) and cannot fully track tumor clones over time. Moreover, deep and accurate cell-free DNA (cfDNA) sequencing is challenging due to constraints of low tumor fraction in the blood and the high cost of WGS sequencing. Ultima Genomics (UG) developed paired plus-minus sequencing (ppmSeq) technology, a whole-genome duplex sequencing approach that lowers observed error rates and allows for a high yield of duplex molecules. Using UG’s deep WGS ppmSeq, we developed our mutation calling algorithms to enable sensitive detection of somatic variants in cfDNA samples, allowing us to track clonal populations over time. To test whether we could identify shared clonal populations in patient sequences between other platforms and ppmSeq technology, we collected post-mortem tissue specimens from 12 patients of various cancer types (breast, cholangiocarcinoma, etc.) and 38 pre-mortem cfDNA samples from the same patients. First, these patients’ data were sequenced with existing Illumina sequencing technologies (WES) and then on the UG sequencing platform for both tissue and ppmSeq. We sequenced the ppmSeq data to an average depth of 135x (range 104x to 208x) and achieved an average duplex proportion of 38% (range 31% to 44%). For all patients’ tissue and cfDNA samples that were sequenced with both ppmSeq WGS (UG) and WES (Illumina), we reconstructed phylogenetic trees to determine the life history of the cancer using our PhylogicNDT suite of tools. Phylogenies reconstructed from ppmSeq WGS cfDNA identified many more mutations and richer trees than those reconstructed from WES of the same cfDNA samples. We compared the cfDNA ppmSeq phylogenies with those constructed from WGS of the same patient’s tissue samples and identified the same clonal populations, enabling us to match clones from the blood to the tissue and track the progression of individual clones. The identification of large clonal populations in ctDNA sequenced with Ultima Genomics’ ppmSeq approach is an improvement over methods with lower sensitivity and coverage; moreover, this approach can identify clonal populations that change over a given treatment course. This more accurate inference of cancer evolution information could enable us to better guide therapies and identify novel mechanisms of resistance that would have been missed with prior methods. Citation Format: Elizabeth E. Martin, Julian Hess, Carrie Cibulskis, Mendy Miller, Brian P. Danysh, Chip Stewart, Elena Helman, Ilya Soifer, Doga C. Gulhan, Dejan Juric, Doron Lipson, Gad Getz. Leveraging Ultima Genomics ppmSeq WGS-cfDNA to accurately detect clonal evolution over sequential blood biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3992.