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Background Merkel cell carcinoma (MCC) is an aggressive skin cancer that evades immune detection by downregulating HLA class-I surface expression through impaired antigen processing machinery (APM) including the proteasome subunits LMP2/7 and peptide transporters TAP1/2. Here, we synthesize and screen poly(beta-amino ester) (PBAE) polymer nanoparticles (NPs) to efficiently transfect MCC and deliver genes that restore APM and drive HLA-I/II expression.Methods PBAE NPs delivering eGFP plasmid were formulated from a library of diverse polymer structures and administered to two human virus-negative MCC cell lines, MCC13 and UISO. Delivery efficacy was evaluated by transfection efficiency (percent GFP+ cells and fluorescence intensity via flow cytometry) and viability. The best-performing PBAE NP, generated from the linear 4 polymer, was subsequently loaded with plasmids encoding APM components (TAP1, TAP2, LMP2, LMP7), the class-I transactivator NLRC5, or the class-II transactivator CIITA. Incubation with 25 ng/mL IFNγ provided a positive control. The impact on APM mRNA levels and HLA-ABC/DR surface expression was assessed by qPCR and flow cytometry, respectively.Results The optimized PBAE NP preserved high cellular viability in both MCC13 and UISO (81% and 63%, respectively) ( figure 1A) and efficient transfection (56% and 74% GFP+, respectively, >40-fold increase in fluorescence intensity in each cell line) (figure 1B,C).Delivery of NLRC5 via NPs significantly increased class I antigen presentation, increasing HLA-ABC expression by 10.8-fold in MCC13 and 14.7-fold in UISO versus untreated cells (figure 2A, B). Delivery of CIITA via NPs significantly enhanced both class I and class II antigen presentation, increasing HLA-ABC expression by 14.6-fold in MCC13 and 12.0-fold in UISO and increasing HLA-DR 26.3-fold in MCC13 and 36.0-fold in UISO versus untreated (figure 2A-D). HLA-ABC increases in both treatments were comparable to IFNy; however, neither NLRC5 NPs nor IFN-γ treated cells exceed a 1.5-fold increase in HLA-DR presentation.PBAE NPs delivering individual APM genes demonstrated target gene expression increasing from >60- to >9,000-fold across both lines (figure 2E,F). Interestingly, off-target upregulation was also observed with delivery of certain APM genes. IFN-γ broadly upregulated class I APM genes in both lines but had minimal effect on CIITA expression.Conclusions Optimized PBAE NPs restore key APM components and markedly enhance both HLA class I and class II surface expression in MCC, providing a strategy to re-engage CD8 + and CD4+ immune responses. Future studies will investigate this approach in virus-positive MCC to enhance presentation of viral antigens and assess antigen-specific T cell responses.Abstract 1011 Figure 1Identification of optimal PBAE nanoparticle formulation for transfection of human MCC cell lines. (A) Cell viability of MCC13 and UISO cells at 48 hours following 2-hour treatment with indicated PBAE NPs. (B) Transfection efficiency measured as the percentage of GFP+ cells. (C) GFP geometric mean fluorescence intensity normalized to cells onlyAbstract 1011 Figure 2HLA surface expression and APM gene expression following PBAE NP delivery in MCC cell lines. (A-B) HLA-ABC expression intensity and UT-normalized fold change in MCC13 and UISO after PBAE NP or IFNγ treatment. (C-D) HLA-DR expression intensity and UT-normalized fold change. (E-F) qPCR of APM genes in MCC13 (E) and UISO (F) post-treatment