<i>SHLD2</i> loss is a synthetic vulnerability to Polθ inhibition combined with radiotherapy.

Abstract

Inhibition of DNA polymerase theta (Polθ), an essential enzyme for repairing DNA double-strand breaks (DSBs) via microhomology-mediated end joining (MMEJ), has proven to be an exquisitely effective monotherapy in HR-deficient tumor models. In addition, Polθ inhibition (Polθi) can induce tumor-selective radiosensitization, but unlike its monotherapy use, no clinically actionable biomarkers have yet been identified to predict this effect. Here, we profiled 54 cancer cell lines and found that Polθi induces substantial radiosensitization in most models, although with marked variability not explained by indicators of Polθ activity. To pinpoint molecular determinants of radiosensitization by Polθi, we performed a CRISPR knockout screen which revealed loss of the TP53BP1/Shieldin pathway component <i>SHLD2</i> (<i>FAM35A</i>) as a vulnerability to Polθi combined with RT. We found that <i>SHLD2</i> is deleted in a subset of human prostate cancers, frequently alongside <i>PTEN</i> loss, an adverse prognostic factor. We demonstrated that <i>SHLD2</i> loss not only increases sensitivity to RT alone, as reported previously, but also enhances the radiosensitizing effect of Polθi, independently of <i>PTEN</i> status and without requiring HR deficiency. Moreover, our findings support a model in which <i>SHLD2</i> deficiency increases Polθ dependence following RT, with Polθ activity limiting DSB accumulation and chromosomal instability, via a compensatory mechanism independent of canonical MRE11/CtIP-mediated DNA end resection. In summary, we found that <i>SHLD2</i> loss is a collateral vulnerability that can be exploited through combined treatment with Polθi and RT.