APOBEC3 activity and DNA polymerase-ε deficiency are associated with distinct IDH1 R132 hotspot mutations.

IDH1 R132 mutations are among the most frequent hotspot mutations in cancer, but their mutational origins have remained unclear. Here, we provide evidence that IDH1 R132C, the predominant IDH1 mutation in cholangiocarcinoma, acute myeloid leukemia, and melanoma, likely arises through APOBEC3-mediated mutagenesis, specifically by APOBEC3A or APOBEC3B. IDH1 R132C is a TpC>TpT substitution on the lagging-strand DNA template within a hairpin-forming sequence context, consistent with APOBEC3 susceptibility. In vitro assays showed that APOBEC3A and APOBEC3B can deaminate the relevant cytosine, while APOBEC3A and APOBEC3B expression patterns in tumor types with recurrent IDH1 R132C were consistent with their potential involvement in generating this mutation. IDH1 R132G, a TpC>TpG substitution at the same site, may similarly result from APOBEC3A or APOBEC3B activity. By contrast, IDH1 R132H, the predominant IDH1 mutation in lower grade glioma and glioblastoma, is a CpG>TpG substitution at a methylated cytosine on the leading-strand DNA template, a pattern more consistent with DNA polymerase epsilon replication error. Concordantly, tumor types enriched for IDH1 R132H showed relatively low POLE expression. Together, these in vitro and bioinformatic analyses provide insight into the distinct mutational mechanisms that likely underlie recurrent IDH1 hotspot mutations in cancer.