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Priming of a Novel Subset of CD28⁺ Rapidly Expanding High-Avidity Effector Memory CTL by Post Maturation Electroporation-CD40L Dendritic Cells Is IL-12 Dependent¹

Research Department, Argos Therapeutics, Inc, Durham, NC 27704

Dendritic cell (DC)-based immunotherapeutics must induce robust CTL capable of killing tumor or virally infected cells in vivo. In this study, we show that RNA electroporated post maturation and coelectroporated with CD40L mRNA (post maturation electroporation (PME)-CD40L DC) generate high-avidity CTL in vitro that lyse naturally processed and presented tumor Ag. Unlike cytokine mixture-matured DC which induce predominantly nonproliferative effector memory CD45RA⁺ CTL, PME-CD40L DC prime a novel subset of Ag-specific CTL that can be expanded to large numbers upon sequential DC stimulation in vitro. We have defined these cells as rapidly expanding high-avidity (REHA) CTL based on: 1) the maintenance of CD28 expression, 2) production of high levels of IFN- and IL-2 in response to Ag, and 3) the demonstration of high-avidity TCR that exhibit strong cytolytic activity toward limiting amounts of native Ag. We demonstrate that induction of REHA CTL is dependent at least in part on the production of IL-12. Interestingly, neutralization of IL-12 did not effect cytolytic activity of REHA CTL when Ag is not limiting, but did result in lower TCR avidity of Ag-reactive CTL. These results suggest that PME-CD40L DC are uniquely capable of delivering the complex array of signals needed to generate stable CD28⁺ REHA CTL, which if generated in vivo may have significant clinical benefit for the treatment of infectious disease and cancer.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ This study was supported by research funding from Argos Therapeutics and Kirin Pharma, Ltd (to M.A.D., D.M.C., H.K., A.H.G., J.M.H., I.Y.T., C.A.N., and D.G.H.).

² Address correspondence and reprint requests to Dr. Mark A. DeBenedette, Argos Therapeutics, Inc., 4233 Technology Drive, Durham, NC 27704. E-mail address: mdebenedette{at}argostherapeutics.com

³ Abbreviations used in this paper: DC, dendritic cell; REHA, rapidly expanding high avidity; PME, post maturation electroporation; MART-1, melanoma Ag recognition by T cell 1; PSA, prostate-specific Ag; APL, altered peptide ligand; ECD, PE-Texas Red-X.