As biochemical and functional studies of membrane protein remain a challenge, there is growing interest in the application of nanotechnology to solve the difficulties of developing membrane protein therapeutics. Exosome, composed of lipid bilayer enclosed nanosized extracellular vesicles, is a successful platform for providing a native membrane composition. This study reports an enzymatic exosome, which harbors native PH20 hyaluronidase (Exo-PH20), which is able to penetrate deeply into tumor foci via hyaluronan degradation, allowing tumor growth inhibition and increased T cell infiltration into the tumor. This exosome-based strategy is developed to overcome the immunosuppressive and anticancer therapy-resistant tumor microenvironment, which is characterized by an overly accumulated extracellular matrix. Notably, this engineered exosome with the native glycosylphosphatidylinositol-anchored form of hyaluronidase has a higher enzymatic activity than a truncated form of the recombinant protein. In addition, the exosome-mediated codelivery of PH20 hyaluronidase and a chemotherapeutic (doxorubicin) efficiently inhibits tumor growth. This exosome is designed to degrade hyaluronan, thereby augmenting nanoparticle penetration and drug diffusion. The results thus show that this is a promising exosome-based platform that harbors not only a membrane-associated enzyme with high activity but also therapeutic payloads.

BackgroundStatins preferentially promote tumor-specific apoptosis by depleting isoprenoid such as farnesyl pyrophosphate and geranylgeranyl pyrophosphate. However, statins have not yet been approved for clinical cancer treatment due, in part, to poor understanding of molecular determinants on statin sensitivity. Here, we investigated the potential of statins to elicit enhanced immunogenicity of KRAS-mutant (KRASmut) tumors.MethodsThe immunogenicity of treated cancer cells was determined by western blot, flow cytometry and confocal microscopy. The immunotherapeutic efficacy of mono or combination therapy using statin was assessed in KRASmut tumor models, including syngeneic colorectal cancer and genetically engineered lung and pancreatic tumors. Using NanoString analysis, we analyzed how statin influenced the gene signatures associated with the antigen presentation of dendritic cells in vivo and evaluated whether statin could induce CD8+ T-cell immunity. Multiplex immunohistochemistry was performed to better understand the complicated tumor-immune microenvironment.ResultsStatin-mediated inhibition of KRAS prenylation provoked severe endoplasmic reticulum (ER) stress by attenuating the anti-ER stress effect of KRAS mutation, thereby resulting in the immunogenic cell death (ICD) of KRASmut cancer cells. Moreover, statin-mediated ICD enhanced the cross-priming ability of dendritic cells, thereby provoking CD8+ T-cell immune responses against KRASmut tumors. Combination therapy using statin and oxaliplatin, an ICD inducer, significantly enhanced the immunogenicity of KRASmut tumors and promoted tumor-specific immunity in syngeneic and genetically engineered KRASmut tumor models. Along with immune-checkpoint inhibitors, the abovementioned combination therapy overcame resistance to PD-1 blockade therapies, improving the survival rate of KRASmut tumor models.ConclusionsOur findings suggest that KRAS mutation could be a molecular target for statins to elicit potent tumor-specific immunity.