Technical Abstract As with many chemotherapeutic drugs, taxanes exhibit poor oral bioavailability; hence, they are administered intravenously (i.v.). This regimen results in significant toxicities, due to both the high drug dose and vehicle (Cremophore ELR) used. Attempts made to develop an oral chemotherapeutic, using liposomes and polymeric nanoparticles as carriers, have not made it to the clinic, due to their inherent limitations. In Phase I, 3P Biotechnologies proposed to develop a ?platform? technology for the oral delivery of chemo drugs and other therapeutics via bovine milk-derived nanoparticles known as exosomes. The Phase I specific aims were to (1) isolate and characterize milk exosomes; (2) prepare exosomal (Exo) formulations of the chemo drug, paclitaxel (PAC), and the plant therapeutic, withaferin A (WFA); and (3) show that Exo formulations enhance therapeutic responses while lacking toxicity. Phase I proved feasible based on the following findings: a) differential centrifugation of raw bovine milk provided exosomes; b) exosomes were taken up by cells in vitro and in vivo; c) Exo formulations of PAC (ExoPACTM), WFA (ExoWFATM), and other compounds were developed, which showed higher anti-proliferative, anti-inflammatory, and anti-cancer activities vs. the free drugs against lung cancer cells, both in vitro and in vivo; and d) milk exosomes lacked cross-species reactivity in wild-type mice. Additional data generated preliminary to Phase II showed that bovine colostrum powder produces exosomes with a yield and purity that are several-fold higher than can be obtained from fresh milk, resulted in significantly higher drug loading than milk, and that exosomes can be functionalized with folic acid for tumor targeting. In Phase II, a team of multidisciplinary researchers will advance this technology by pursuing the following specific aims: 1) Establish the reproducibility of colostrum exosome yield, maximize drug loading of the taxane PAC, identify and optimize loading of a tumor-targeting ligand, and test the resulting formulations for antiproliferative and anti-inflammatory activities against human lung cancer cells in vitro. 2) Determine the efficacy of Exo formulation of PAC (ExoPACTM) and functionalized-ExoPACTM for tumor targetability, minimizing off-target sites, using orthotopic xenograft, patient-derived tumor xenograft (PDX), and K-ras spontaneous tumors. 3) Determine toxicities of the ExoPACTM and functionalized-ExoPACTM formulations, analyze blood and tissue levels of PAC (PK/PD studies), and determine stability of the formulations. The innovation lies in the use of standardized colostrum-derived, functionalized exosomes for oral delivery for cancer therapeutics to circumvent bioavailability issues. The current taxane therapeutic market is dominated by a single product, Abraxane, an i.v. albumin-PAC product. A product that performs similar to or better than Abraxane, would potentially generate U.S. and worldwide markets of over $7 billion and $28 billion, respectively. If we are successful in achieving our objectives we will apply for Phase IIb, where we will establish large-scale production of ExoPACTM and will conduct Phase 1/2 clinical trials. Oncologists on the Advisory Board of 3P Biotechnologies will guide as we advance these drug formulations.