Abstract 3206: Synthetic lethal interactions with APOBEC3A identified by functional genomic screening

Abstract

Abstract The APOBEC3 (A3) family of cytosine deaminase enzymes is integral to innate defenses by editing viral genomes to limit infectivity. However, several members of the A3 enzyme family (A3A-A3H) have also been implicated in mutational signatures observed in human cancers. The A3 mutational signature has emerged as the most prevalent across all cancers, implicating A3 enzymes as powerful mutators of the human genome. While few prospective studies have addressed the potential for A3 enzymes to induce specific mutational patterns in human genomes, the possibility of exploiting the mutagenic activity of A3 for cancer therapy is beginning to be explored. We recently showed that A3A is highly expressed in human acute myeloid leukemia (AML), and that deamination activates the replication checkpoint via ATR kinase signaling. In an AML model, we showed that inhibition of ATR or downstream Chk1 resulted in accumulated DNA damage by A3A and ultimately death of leukemia cells. In search of additional cellular pathways that may be manipulated to take advantage of A3 activity for therapeutic effect, we performed a functional genomics screen using a genome-wide CRISPR knock-out approach in a model of human leukemia with inducible A3A expression. We introduced Cas9 and the Brunello guide RNA lentivirus library into the human AML cell line, THP1, which we engineered to express doxycycline-inducible A3A. The Brunello library consists of 4 guide RNAs targeting each human gene. Transduction was performed to enable one lentivirus integration per cell, thus resulting in a single gene disruption per cell. Cells were treated with doxycycline and genomic DNA was harvested from treated and untreated cells every week. Guide RNAs were amplified by PCR from genomic DNA and identified by next-generation sequencing. We applied the MaGECK algorithm to identify the quantity of each guide RNA remaining in treated samples and compared to untreated samples. In THP1 cells induced to express A3A, knockout of genes in cell cycle and DNA repair pathways resulted in synthetic lethality. Specifically, expression of A3A synergized with genes essential for cell cycle checkpoints and DNA damage responses to cause leukemia cell death. Clinical-grade inhibitors are available for protein products of many of the genes identified; thus these represent feasible targets for therapeutic intervention in AML that has elevated A3A activity. In addition to leukemia, A3A and the closely related enzyme A3B have been implicated in several malignant processes including breast, bladder, and ovarian cancers. We show that functional genomic screening is a powerful tool for identification of synthetic lethal interactions, which can provide therapeutic opportunities for additional cancers in which A3 enzymes are active. Citation Format: Abby M. Green, Katharina E. Hayer, Julia H. Szeto, Ophir Shalem, Matthew D. Weitzman. Synthetic lethal interactions with APOBEC3A identified by functional genomic screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3206.