Zhendong Cao, Sixiang Yu, Jacqueline Peng, Declan R Barrett, Yuqiao Liu, Jonathan H. Sussman, Changya Chen, Anusha Thadi, Li Liu, Fatemeh Alikarami, Jason Xu, Martin P. Carroll, Kai Tan, Kathrin M. Bernt, Junwei Shi. CRISPR-based functional genomics for dissecting therapeutic dependency in primary acute myeloid leukemia samples. Mol Cell. 2026 Feb 26:S1097-2765(26)00098-5. doi: 10.1016/j.molcel.2026.02.003. Epub ahead of print. PMID: 41759529.

Abstract

Cancer functional genomics enables high-throughput target discovery and mechanistic investigation, yet its application has remained largely confined to mouse models and established human cancer cell lines. Direct functional interrogation of heterogeneous primary tumors offers a powerful opportunity to evaluate therapeutic targets and uncover cancer dependencies or resistance mechanisms. Here, we developed an optimized CRISPR-based platform for functional genomics in patient-derived xenograft and primary acute myeloid leukemia (AML) samples harboring diverse pathogenic mutations. Integrated in vitro and in vivo CRISPR-Cas9 knockout and CRISPR interference (CRISPRi) dropout screens validated known AML-biased targets and identified cis-regulatory elements essential for leukemic growth. Coupling pooled CRISPR perturbations with single-cell RNA sequencing (Perturb-seq) further resolved the perturbation-induced alterations in regulatory networks, cell cycle states, and cellular hierarchies in primary AML samples. Together, these studies establish a general and robust framework for leveraging CRISPR-based functional genomics to directly dissect cancer dependencies and cellular heterogeneity in primary AML patient samples.