A targeted CRISPR screen reveals new genes that are involved in the resistance of acute myeloid leukemia cells to Cytosine arabinoside (ARA-C).

Acute myeloid leukemia (AML) is often associated with poor outcome, particularly in older patients that become refractory to initial chemotherapy. In a previous study, we identified a copy number neutral-loss of heterozygosity (CNN-LOH) region on chromosome 1p that was more common in old patients comparing to young patients. CNN-LOH usually occurs in areas containing molecular aberrations, resulting in loss of the WT allele and duplication of the mutant allele. Therefore, we hypothesized that the minimal common region of chr-1p CNN-LOH shared among AML patients might harbor genetic aberrations that lead to chemo-resistance. In this study we screened for genes within that region that confer resistance to cytosine arabinoside (ARA-C) chemotherapy upon loss of function. For that purpose we designed a CRISPR-sgRNA library that targeted 420 coding genes in the 1p CNN-LOH region. OCI-AML2 cell line with stable Cas9 expression were transduced by the pooled library, selected with ARA-C at the IC₉₀ and IC₉₉ and subjected to NGS and decoding of resistant clones. Validation of top resistant clones revealed that knockout of 2 genes, CASP9 and SLC45A1, enabled cell growth under ARA-C treatment. Further studies will elaborate on the mechanism by which silencing of each of these genes confer chemo-resistance. These results may pave the way to the development new drugs that in combination with ARA-C will improve the efficacy of the standard AML therapy.