Abstract PO-132: CaTCH - A barcode-guided CRISPRa-inducible reporter to isolate clones from heterogeneous populations

The emergence of resistant cell clones to targeted therapies poses a significant issue in the treatment of metastatic melanoma. While these founding clones are often extremely rare in a starting population, their isolation and characterization holds unique potential for understanding disease processes, uncovering novel biomarkers and developing therapeutic concepts. The functional characterization of such founder clones and comprehensive comparisons to their post-selection counterparts requires live cells. To achieve this, we developed a novel lineage tracing tool termed CaTCH (CRISPRa tracing of clones in heterogeneous cell populations). CaTCH combines precise mapping of the lineage history of millions of cells with the ability to isolate any given clone alive from a complex population based on genetic barcodes. CaTCH thereby enables the retrospective isolation and analysis of founding clones from heterogeneous cell populations prior to evolutionary selection. In first applications, we use CaTCH to provide insights into the development of resistance to targeted cancer therapies. We demonstrate that CaTCH can be used to trace and isolate a single pre-existing therapy-resistant clone from a complex cancer cell population in vitro. Furthermore, we validate the utility of CaTCH for applications in vivo by investigating the origins of resistance to clinically relevant RAF/MEK inhibition in an immunocompetent melanoma mouse model. Here we find that most clones have the capacity to acquire resistance to combined RAF/MEK inhibitor therapy, indicating that resistance to this clinically relevant regimen is a universally achievable state in this model. We envision that CaTCH will address fundamental questions in basic and translational research (e.g., how cell identity states and trajectories are determined in therapy resistance, metastasis formation, tissue development and somatic cell re-programming), potentially revealing new vulnerabilities that can serve as targets for therapies. Citation Format: Christian Umkehrer, Felix Holstein, Laura Formenti, Julian Jude, Kimon Froussios, Tobias Neumann, Shona M. Cronin, Lisa Haas, Jesse Lipp, Thomas R. Burkard, Michaela Fellner, Thomas Wiesner, Johannes Zuber, Anna C. Obenauf. CaTCH - A barcode-guided CRISPRa-inducible reporter to isolate clones from heterogeneous populations [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-132.