Abstract 5683: Single-cell RNA-seq reveals heterogeneity, clonal evolution and strategies to overcome ibrutinib-venetoclax dual resistance in mantle cell lymphoma

Introduction The clinical, pathological, genetic, and transcriptomic characteristics of mantle cell lymphoma (MCL) are extremely heterogeneous, and MCL patients frequently develop therapeutic resistance after frontline therapies. In this study, we performed longitudinal transcriptomic analysis on patient specimens at the single-cell level, aiming to understand the dynamic and complex cellular and molecular changes underlying therapeutic resistance and to identify potential targets to overcome dual resistance to ibrutinib and venetoclax. Methods Single-cell RNA-seq was performed on sequential MCL patient specimens. Integrative computational approaches were employed to characterize the cellular and molecular basis of therapeutic resistance and clonal evolution. Bulk RNA sequencing, deep WES, and 17-color flow analysis of multiple patient cohorts were used to validate the novel findings. An orthotopic PDX model derived from one of the ibrutinib-venetoclax non-responders was used for further analysis and to investigate the in vivo efficacy of therapeutic agents against multiple actionable targets. Results To dissect the inter- and intra-tumor heterogeneity underlying the therapeutic resistance, we performed sequential scRNA-seq analysis of 21 specimens (18,794 cells in total) collected from 3 ibrutinib responders and 2 ibrutinib-venetoclax non-responders along the course of ibrutinib and/or venetoclax treatments. The analysis revealed a high degree of inter- and intra-tumor heterogeneity with distinct cellular and transcriptomic profiling within and across ibrutinib-responders and ibrutinib-venetoclax non-responders. The observed transcriptomic evolution of cancer hallmarks associated with disease progression and therapeutic resistance were faithfully passed onto the orthotopic PDX model derived from one of the ibrutinib-venetoclax non-responders. We tracked the clinical ibrutinib-induced lymphocytosis at a single-cell level in ibrutinib responders and disease progression-associated clonal evolution in non-responders, which was validated by deep WES analysis. 17q gain in non-responders was further validated in a larger patient cohort. Our analysis revealed the reprogramming of tumor microenvironment and tumor immune evasion, which was further validated by 17-color flow analysis in a large cohort (n = 70). Inhibition of multiple actionable targets showed effective anti-MCL activity in the PDX model to overcome the ibrutinib-venetoclax dual resistance. Conclusions This study demonstrates the potential of longitudinal single-cell transcriptomic analysis to reveal the molecular mechanisms underlying tumor heterogeneity, clonal evolution, disease progression, and therapeutic resistance, and to identify potential novel targets to circumvent therapeutic resistance in MCL and other diseases. Citation Format: Changying Jiang, Shaojun Zhang, Yuanxin Wang, Rongjia Zhang, Yang Liu, Joseph McIntosh, Guangchun Han, Ruiping Wang, David Santos, Maria Badillo, Angela Leeming, Zhihong Chen, Kimberly Hartig, John Bigcal, Hun Lee, Raphael Steiner, Jorge Romaguera, Preetesh Jain, Krystle Nomie, Andy Futreal, Linghua Wang, Michael Wang. Single-cell RNA-seq reveals heterogeneity, clonal evolution and strategies to overcome ibrutinib-venetoclax dual resistance in mantle cell lymphoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5683.