Synthetic Lethal Interactions of APOBEC3A Deaminase with DNA Repair Inhibition in Acute Myeloid Leukemia
Mentor: Dr. Abby Green
A recently identified source of somatic mutation in cancer are the APOBEC3 (A3) cytosine deaminases. A3 enzymes mutate DNA cytosine bases, resulting in specific mutational patterns that are evident in cancer genome sequences. Our prior work identified single-stranded DNA (ssDNA) as a cellular substrate on which A3 enzymes act. We showed that the APOBEC3A (A3A) family member mutates replicating DNA and causes replication stress, leading to activation of the DNA replication checkpoint via ATR kinase signaling. Cancer cells with high levels of A3A expression are dependent on ATR signaling. We previously showed that A3A is highly expressed in a subset of pediatric and adult AML. Thus, we modeled A3A expression in AML and found that inhibition of ATR or Chk1 leads to cancer cell death.
We hypothesized that additional genome-protective responses would be activated by A3A deamination in cancer cells. We performed a functional genome-wide screen using a CRISPR-Cas9 library to determine gene knockouts that resulted in synthetic lethality with high A3A expression. Several gene sets associated with DNA damage responses were enriched in our screen. We hypothesize that A3A activity results in cancer cell dependence on specific DNA damage response factors that represent potential therapeutic vulnerabilities.
To confirm screen findings, we will use CRISPR-Cas9 gene editing to knock down individual candidate genes in a panel of AML cell lines with inducible A3A expression and will evaluate DNA damage (by phospho-H2AX staining), cytotoxicity (by viability assays), and apoptosis (by Annexin V staining). Validated synthetic lethal knockouts will be evaluated in primary AML cells with high levels of endogenous A3A. These studies will define therapeutic vulnerabilities in AML with high A3A expression.
