Combinatorial leaky probiotic for anticancer immunopotentiation and tumor eradication

Combination therapies present a compelling therapeutic regimen against the immunosuppressive and heterogeneous microenvironment of solid tumors. However, incorporating separate therapeutic modalities in regimen designs can be encumbered by complex logistical, manufacturing, and pharmacokinetic considerations. Herein, we demonstrate a single-vector combinational anticancer therapy using an lpp gene knockout leaky probiotic for simultaneous secretion of immunotherapeutic and oncolytic effector molecules. Through fusion protein design and vector optimization, a Nissle1917 (EcN) bacteria vector is engineered to secrete Neoleukin-2/15 (Neo-2/15) cytokine-functionalized anti-PDL1 nanobody (aPDL1-Neo2/15) and anti-mesothelin-functionalized hemolysin E (HlyE-aMSLN). The multifunctional leaky probiotic enables synchronous immune activation and tumor-targeted cytolytic activity for effective tumor suppression, elevation of tumor immune cell infiltration, and establishment of anticancer immunological memory. lpp gene knockout is further shown to improve probiotic tolerability and intravenous applicability, offering a therapeutically viable approach for combination regimen development. [Display omitted] •Δlpp gene knockout enhances leakiness and tolerability of bacterial vectors•Engineered leaky probiotic secretes immunomodulatory and oncolytic chimeric proteins•Combinatorial leaky probiotic exerts tumor suppression and induces adaptive immunity•Combinatorial leaky probiotic is amenable to intravenous anticancer treatment Liu et al. establish a multifunctional leaky probiotic that co-expresses an immunotherapeutic and an oncolytic chimeric protein for anticancer immunopotentiation and tumor eradication. The loss-of-function strategy renders the bacterial vector leaky for therapeutic protein release. Additionally, the strategy also improves the bacterial vector’s safety profile for intravenous anticancer therapies.