In vivo genetic circuit drives systemic PARP1 siRNA assembly into small extracellular vesicles for BRCA2-deficient breast cancer therapy.

BRCA2-mutant breast cancer is a major clinical challenge, with current poly ADP-ribose polymerase inhibitor (PARPi) therapies limited by systemic toxicity and acquired resistance. This study aimed to develop a RNA interference (RNAi) strategy to selectively inhibit PARP1 in breast tumors via a systemic, non-viral delivery platform. We designed a genetic circuit as a naked DNA plasmid, encoding a PARP1-specific siRNA embedded within a pre-miR-155 backbone under the control of a cytomegalovirus (CMV) promoter. Its therapeutic potential was assessed following intravenous injection in two orthotopic breast cancer models: an immunocompetent BRCA2-deficient model (E0771) and an immunodeficient BRCA2-mutant xenograft model (HCC1599). The plasmid was predominantly taken up by the liver, where it reprogrammed hepatocytes to produce and package the PARP1 siRNA into endogenous small extracellular vesicles (sEVs) for systemic circulation. These sEVs demonstrated intrinsic tumor-homing capabilities, leading to efficient delivery and significant PARP1 gene silencing within breast tumors. This resulted in potent inhibition of tumor growth, accompanied by increased apoptosis and reduced proliferation in both preclinical models. The platform showed enhanced tumor specificity and a reduction in off-target effects compared to conventional small-molecule PARPi. We present a novel synthetic biology approach that leverages hepatic sEV production for the systemic delivery of siRNA to breast cancer. This strategy effectively suppresses tumor growth in BRCA2-deficient mice models and offers a promising therapeutic alternative with potential to overcome the limitations of current PARP inhibitor treatments for breast cancer.