Engineered MAP30ER: A plant toxin-derived platform for EGFR-Targeted delivery of protein and chemotherapeutic...

The nonspecific toxicity of traditional chemotherapeutic agents and the intracellular delivery barriers of protein-based drugs have limited their clinical applications. Efficient targeted delivery properties of toxin proteins themselves provide important insights into the design of novel drug delivery systems. Inspired by the natural targeting properties of the plant-derived type I ribosome-inactivating protein (RIPs) MAP30, we engineered a detoxified carrier, MAP30ER, through site-directed mutagenesis of key enzymatic residues (E158A/R161A). This variant retained its cell-binding capability while exhibiting significantly reduced cytotoxicity. ELISA and molecular docking identified EGFR as the primary functional receptor of MAP30, and flow cytometry confirmed a 3-fold higher binding affinity of MAP30 to EGFR-high tumor cells (A431) compared to EGFR-low cells (HeLa). To address endosomal entrapment, we screened triterpenoid saponins and found that Saikosaponin A (SSA) enhanced the endosomal escape efficiency of MAP30ER by inducing vesicle membrane disruption. When applied to deliver Apoptin (a apoptosis-inducing protein), the MAP30ER-SSA system elevated tumor cell killing efficacy from 65 % to 90 % upon SSA co-treatment, and Western blot confirmed that it induced apoptosis through activation of caspase-3/9 and cleavage of PARP1. Furthermore, we extended antibody-drug conjugate (ADC) technology to the MAP30ER platform, constructing a MAP30ER-MMAE conjugate that achieved targeted delivery to A431 cells with negligible toxicity to normal cells. This study establishes a novel paradigm for engineering plant-derived RIPs and provides a multifunctional delivery platform with targeting specificity, safety, and versatility, offering innovative solutions to overcome drug delivery challenges in cancer therapy.