Targeting tumor-selective membrane-associated PCNA enables selective antibody-drug conjugate therapy.

Membrane-associated proliferating cell nuclear antigen (mPCNA) is an attractive tumor-selective antigen for antibody-drug conjugate (ADC) development. We generated a chimeric IgG1 ADC, RC14-MMAE, to exploit three key properties of mPCNA that support its use as an efficient delivery platform. First, flow-cytometric and imaging analyses show abundant mPCNA surface expression on multiple tumor cell lines and in JIMT1 xenografts, providing accessible epitopes for binding. Second, comparative profiling against non-malignant counterparts and normal mouse tissues reveals minimal or undetectable mPCNA at the plasma membrane of healthy cells, indicating a sharp tumor-normal differential expected to limit off-tumor targeting and improve the therapeutic window. Third, live-cell internalization assays demonstrate that RC14 rapidly accumulates in LAMP1-positive lysosomes after engaging mPCNA, confirming efficient trafficking along the endolysosomal pathway required for protease-mediated cleavage of the valine-citrulline linker and release of monomethyl auristatin E (MMAE). Consistent with this target profile, RC14-MMAE exhibits potent antigen-dependent cytotoxicity in mPCNA-positive tumor models in NSG mice, favorable pharmacokinetics, sustained payload retention in serum, robust in vivo antitumor activity with limited overt toxicity, and a significantly increased fraction of F4/80⁺ tumor-infiltrating myeloid cells within dissociated tumors. These findings establish mPCNA as a functionally validated, tumor-selective internalizing target and support RC14-MMAE as a first-in-class mPCNA-directed ADC with broad translational potential.