The Protein-Like Polymer (PLP)—A Tunable Proteomimetic Nanoplatform for the Development of Rationally Designed Cancer Vaccines with Therapeutic Efficacy in Multiple Tumor Models

<b>Background</b><br/>Utilizing tumor antigens for the development of patient-specific cancer vaccines has been a promising therapeutic strategy. However, significant challenges remain in delivering subunit vaccine components in a manner capable of eliciting antitumor immune responses. To overcome these, we developed rationally designed cancer vaccines using a novel nanoplatform called the Protein-Like Polymer, a reference to its globular structure reminiscent of native proteins, with unique characteristics allowing for sustained targeted delivery of tumor antigens in conjunction with immunomodulatory STING agonists.<br/><b>Methods</b><br/>PLPs containing peptide antigens were synthesized via ring-opening metathesis polymerization (ROMP) and characterized using NMR, HPLC, ESI, and SEC-MALS. A library of compounds with different sidechain linkage chemistries, degrees of polymerization (DP, number of side chains per polymer), and inclusion/exclusion of Oligo(ethylene glycol) (OEG) were made. Cell uptake and functional assays using payload-specific T Cells were conducted using polymers with or without fluorescent labels. Immunization in independent tumor models was done to show generalizability. Ability of PLPs to co-deliver adjuvants was tested by electrostatically coupling a small molecule STING agonist, 2’3’ cGAMP, forming stable nanostructures.<br/><b>Results</b><br/>Dense brush polymers wherein every side chain extending off a hydrophobic polymer backbone consists of peptide antigens were successfully generated with narrow polydispersity and predetermined DPs. Conjugating peptide antigens to the polymer backbone using a cleavable disulfide linkage, designed to reduce intracellularly in antigen presenting cells (APCs), resulted in increased endosomal localization, higher levels of induced T cell proliferation, cytokine production, and expression of activation markers in CTLs and APCs. Incorporating a diluent amount of OEG side chains reduced enzymatic degradation while increasing immunogenicity and uptake by APCs. Additionally, increasing the DP, and therefore the density of antigen side chains, further improved vaccine efficacy and resistance to proteolysis. Antigen-PLP conjugates enhanced dendritic cell activation and T-cell response only when paired with cells from their cognate system, with no response in immune cells not expressing receptors for the payload, further demonstrating antigen-specificity. Mice bearing established B16F10 melanomas treated with PLPs containing gp100 resulted in significant increases in survival time, reduced tumor burden, and corresponding changes in immune cell profiles. Impressively, mice treated with STING electrostatically complexed to antigen-PLPs showed significantly smaller tumors vs non-complexed combination treatment while allowing for subcutaneous administration of 2’3’ cGAMP, which alone requires intratumoral injection due to rapid diffusion. Studies on MC38, TC-1, and LLC1-OVA tumors, paired with adpgk, HPV-E7, and OVA-bearing PLPs respectively, as well as pools of neoantigens multiplexed onto a single PLP are ongoing.<br/><b>Conclusion</b><br/>This work validates the ability of PLPs to overcome major limitations in cancer vaccine development, enabling sustained delivery of antigens and adjuvants. The modularity of the platform allows for complex nano-architectures including systems capable of delivering challenging compounds, ie small molecule STING agonists, subcutaneously through stable electrostatic coupling. The ability to multiplex multiple different antigenic sequences as mosaic structures further highlights the PLP platform’s potential to revolutionize cancer vaccinology.