Graduate Research Assistant Roswell Park Comp. Cancer Ctr. Buffalo, New York, United States
Disclosure(s):
Gavin Twoey, BS: No financial relationships to disclose
Introduction/Rationale: The use of antigen (Ag) presenting dendritic cells (DC) strategy can prompt effective and personalized anti-tumor immune responses with little immune-mediated toxicity. However, current formulations have resulted in clinical objective responses amongst only a small subset of patients. Current strategies rely on cytokine-based maturation cocktails that result in heterogenous maturation of DCs as measured by costimulatory molecule expression. We propose a novel hybrid vector (HV) that utilizes a bacterial core encased in a polymeric shell to deliver plasmid DNA (pDNA)-encoded Ag, resulting in simultaneous delivery of maturation stimuli and target Ag to be durably expressed within DC. We hypothesize that this method will improve DC maturation and resultant Ag-specific T cell expansion, thereby improving antitumor responses.
Methods: To evaluate the effect of the HV, human monocytes isolated from whole blood were differentiated for 7 days with GM-CSF and IL-4 into monocyte-derived DC and subsequently co-cultured with HV constructs or a maturation cytokine cocktail of IL-1β, TNFα, IL-6, and PGE2. Following stimulation, DC were directly analyzed via flow cytometry or co-cultured with T cells before analysis.
Results: HV-conditioned DC showed a 1.9-fold increase in CD80 expression and a 2.7-fold increase in CD83 expression after 48 hours compared to standard DC differentiation protocols. Using plasmid-encoded fluorescent reporters, expression of pDNA-encoded protein was detected via imaging flow cytometry and fluorescent microscopy only in HV-conditioned DC, suggesting successful pDNA translocation and expression within DC. DC conditioned with the HV encoding MART-1 Ag promoted a 4-fold increase in the expansion of MART-1-specific T cells when co-cultured compared to a plasmid control.
Conclusion: The HV provides a novel and highly modifiable platform for both enhancing DC maturation and Ag presentation while also offering mechanistic insight into both DC biology and resultant immune responses.
