Abstract 1523: Cell Squeeze® delivery of antigen-encoding mRNA enables human PBMCs to drive antigen-specific CD8+ T cell responses for diverse clinical applications

Antigen-specific CD8+ T cell priming is critical for mounting an effective immune response against altered self or foreign antigens that are found in most tumors and virally infected cells. T cell priming events require antigen presenting cells (APCs) to process and load antigenic peptides onto MHC molecules. One recently established method for providing APCs with foreign antigen is through delivery of mRNA. An advantage of using antigen-encoding mRNA, compared to protein or peptides, is the inherent protein amplification of mRNA translation; a single mRNA can be translated multiple times. In addition, when antigen-encoding mRNA is delivered directly to the cell, the antigens will have the appropriate post-translational modifications, potentially increasing the immunogenicity. Whereas typical mRNA vaccine platforms require modified nucleotides and lipid nanoparticle encapsulation, Cell Squeeze® technology allows for delivery of uncomplexed and unmodified mRNA directly into the cytosol. Here, we use ex vivo microfluidic Cell Squeeze® technology to deliver uncomplexed and unmodified mRNA encoding for disease related antigens into human peripheral blood mononuclear cells (PBMCs) to generate PBMC APCs. We demonstrate that squeezing PBMCs with mRNA results in effective delivery and translation of mRNA in major cell subsets including T cells, B cells, NK cells, and monocytes. Delivery of antigen-encoding mRNA to human PBMCs enables them to function as APCs, capable of presenting antigenic peptides on MHC molecules for activation of antigen-specific T cells. Specifically, we demonstrate squeezing antigen-encoding mRNA (e.g. HPV16 E7) into human PBMCs results in PBMC APCs that can elicit robust in vitro activation of antigen-specific CD8+ T cells. This approach to APC engineering demonstrates Cell Squeeze® technology's potential to leverage the modularity, cost-effectiveness, and streamlined manufacturing of multiple mRNAs to create cellular vaccines for treating various tumor and infectious disease targets. Citation Format: Michael F. Maloney, Emrah Ilker Ozay, Christian Yee, Amy Merino, Paul R. Dunbar, Mubeen Mosaheb, Kelly Volk, Carolyne Smith, Katherine J. Seidl, Howard Bernstein, Scott M. Loughhead. Cell Squeeze® delivery of antigen-encoding mRNA enables human PBMCs to drive antigen-specific CD8+ T cell responses for diverse clinical applications [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 an