PROJECT ABSTRACT Mucinous colorectal and appendiceal cancers (MCAC) are unique histologic subtypes that frequently metastasize to the peritoneal cavity (known as mucinous carcinoma peritonei [MCP]). MCP is frequently unresectable, responds poorly to standard intravenous chemotherapy, and often recurs after “curative” surgery with intraperitoneal (IP) chemotherapy, resulting in poor oncologic outcomes. Intraperitoneal chemotherapy for MCP faces two major challenges. First, MCP is characterized by abundant extracellular mucus that forms a protective barrier around cancer cells, hindering IP chemotherapeutic drug delivery. We have previously demonstrated robust mucolysis, in patient-derived in vitro and in vivo models of MCP, using mucolytic drugs (e.g. bromelain [BRO], N-acetylcysteine [NAC] and trypsin [TRYP]). We also found that the baseline net negative charge of mucus was significantly increased after mucolysis (ζ-potential in our studies: undigested mucus -1.93 mV; digested mucus -17.2 mV). Second, commonly administered IP drugs for MCP (e.g. doxorubicin [DOX] and mitomycin C [MITO]) are rapidly absorbed across the peritoneal membrane, resulting in short IP retention time, low intra-tumoral (IT) penetration, and systemic toxicity. Therapeutic nanoparticle formulations have longer IP retention and IT penetration than free drugs because of enhanced permeability and retention effect and provide protection from early degradation and pre-absorption. The aim of this proposal is to leverage nanotechnology and the significant negative charge of mucus following mucolysis to enhance IP retention, IT penetration, and delivery of positively charged anticancer drugs in MCP. To this end, we have synthesized mucus modulating multipurpose TRYP nanoparticles (MTN) comprised of three components; (a) a core of negatively charged TRYP clusters, consisting of 4 arms of polyethylene glycol (PEG) and TRYP, for enzymatic mucolysis and drug delivery; (b) nanoparticle-conjugated NAC, for mucus disruption and mucoadhesion; and (c) nanoparticle-loaded and positively-charged mitochondria-targeted anticancer drugs (mitocans), specifically triphenyl phosphonium (TPP)- doxorubicin (TPP-DOX) and TPP-mitomycin C (TPP-MITO), for anti-cancer effect. We hypothesize that our MTN will disrupt the structural integrity of mucus, enhance IP/IT retention and penetration of loaded drugs, and deliver positively charged TPP-DOX or TPP-MITO across a progressively higher negative charge-gradient from the nanoparticle surface to digested mucus to mitochondria (ζ-potential: digested mucus -17.2 mV; cell membranes -30 to -60 mV; mitochondrial membranes -160 mV). Our research proposal provides a novel therapeutic strategy to overcome the cytoprotective mucus barrier and improve drug delivery in MCP. It is expected that the proposed MTN will provide a pharmacokinetic and pharmacodynamic advantage over non- nanocarrier formulations of the drugs. Notably, the proposed MTN are synthesized from bioc...