Abstract 4696: High-throughput single-cell targeted DNA sequencing using an updated TapestriTM Platform reveals rare clones and clonal evolution for multiple blood cancers

Abstract

Abstract Introduction: The challenge in precision medicine has been improving the understanding of cancer heterogeneity and clonal evolution, which has major implications in targeted therapy selection and disease monitoring. However, current bulk sequencing methods are unable to unambiguously identify rare pathogenic or drug-resistant cell populations and determine whether mutations co-occur within the same cell. Single-cell sequencing has the potential to provide unique insights on the cellular and genetic composition, drivers, and signatures of cancer at unparalleled sensitivity. Methods: Previously we have developed a high-throughput single-cell DNA analysis platform (TapestriTM) that leverages droplet microfluidics and a multiplex-PCR based targeted DNA sequencing approach, and demonstrated the generation of high-resolution maps of clonal architecture from acute myeloid leukemia (AML) tumors. Here we present an update to the Tapestri Platform which employs new biochemistry and features improved firmware, software, workflow, and data analysis solution resulting in higher throughput, better sensitivity, specificity and unprecedented flexibility. Results: From cell prep to sequencing-ready libraries, the workflow can be completed within 2 days, and new modifications have doubled the throughput to up to 20,000 genotyped cells per run from 10,000 shown previously. We have validated the performance of an AML (19 genes, 50 amplicons) and a CLL (chronic lymphocytic leukemia) (34 genes, 286 amplicons) panel. We also developed a robust web-based design portal for custom targets. The updated biochemistry enables easy addition of new gene and loci targets into existing panels for improved coverage and updated studies. Using longitudinal AML and CLL samples, we were able to detect rare subclones of <0.1% prevalence, identify mutation co-occurrence, and characterize clonal evolution due to disease progression and drug treatment. Conclusion: We demonstrate that single-cell DNA sequencing can reveal the heterogeneity of blood cancers and map the clonal architecture and clonal evolution with higher sensitivity than bulk NGS methods. This is critical in patient stratification and drug selection over the entire course of treatment. Besides the catalog AML and CLL panels, the flexibility of system allows for analyzing SNV and indel mutations of any custom cancer DNA targets. Additionally, the system provides capabilities for quality control of gene edited cells, further advancing research into cancer therapies. Citation Format: Nianzhen Li, Daniel Mendoza, Adam Sciambi, Mani Manivannan, Jacob Ho, Kaustubh Gokhale, Jacqueline Marin, Kathryn Thompson, Jamie Yates, Vasu Sharma, Steven Chow, Sombeet Sahu, Shu Wang, Dennis Eastburn, Keith Jones. High-throughput single-cell targeted DNA sequencing using an updated TapestriTM Platform reveals rare clones and clonal evolution for multiple blood cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4696.