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Background Ovarian cancer has a five-year relative survival rate of 50.9%, making it the most lethal gynecological cancer. Treatment typically involves surgery, chemotherapy, or both. However, 80% of ovarian cancer diagnoses are made in advanced stages (Stage III and IV), where the five-year prognosis is further diminished to 20-40% and chemotherapy resistance has increased. The tumor immune microenvironment (TIME) in ovarian cancer is highly tumor suppressive due to the primary tumor and the malignant ascites that accumulates with disease progression. Existing immunotherapies are not effective in 85-90% all ovarian cancer patients, and, consequently, there is a need to develop an improved immunotherapy platform for ovarian cancer that can overcome the immunosuppressive environment. Here we describe a gene therapy using poly(beta-amino ester) (PBAE) nanoparticles (NPs) to deliver T-cell co-activation signals 4-1BBL and IL-12 into ovarian cancer cells. These signals, collectively with the intrinsic MHC:antigen signal in tumor cells, reprogram tumor cells into tumor-associated antigen presenting cells (tAPCs), leading to a strong anti-tumor response that can be paired with checkpoint inhibitors.Methods We delivered (PBAE) nanoparticles containing 4-1BBL and IL-12 DNA into ovarian cancer cells to reprogram them into tumor-associated antigen-presenting cells (tAPCs). First, we validated tumor reprogramming in vitro in three different murine ovarian cancer cell lines (ID8, ID8 VEGF, and OV2944) by co-culturing with splenocytes isolated from C57BL/6 mice. We then measured overall survival and immune cell infiltration in vivo after administration of four groups: 1) GFP NP, 2) GFP NP + systemically delivered anti-PD1, 3) 4-1BBL/IL-12 NPs, and 4) 4-1BBL/IL-12 NPs + systemically delivered anti-PD1.Results In all three ovarian cancer cell lines, an increase in IFNγ (interferon-gamma) response was observed after co-delivery of 4-1BBL and IL-12 compared to delivery of GFP nanoparticles. In ID8 and OV2944 cell lines, the 4-1BBL/IL-12 NP elicited significantly higher IFNγ secretion than all other NP groups (p<0.0001). In the ID8 VEGF cell line, which was engineered to display a more aggressive phenotype, the 4-1BBL/IL12 NP also elicited significantly higher IFNγ secretion compared to GFP NPs (p<0.0001) and IL-12 NPs (p<0.001). In vivo, both groups delivering 4-1BBL/IL-12 NPs demonstrated significantly improved survival compared to the GFP NP group (p<0.001). Furthermore, in the combination group with 4-1BBL/IL-12 NPs + anti-PD1, there were two long term survivors, where all other groups had zero long term survivors (figure 1).Conclusions The 4-1BBL/IL-12 nanoparticle reprogramming approach represents a personalized immunotherapy platform for ovarian cancer without requiring a priori antigen knowledge.Abstract 1030 Figure 1Ovarian cancer reprogramming