Abstract LB531: The addition of IFNα and 5-Aza-2'deoxycytidine treatments to a dendritic cell-targeting DNA vaccine enhance the expression of Th1-related genes and the infiltration of CD11c+ CD8+ T-cells
Authors: Gordy James T. , Comstock Kaitlyn , Sandhu Avinaash K. , Hui Yinan , Karanika Styliani , Sunshine Joel , Karakousis Petros C. , Markham Richard B. 
Published: June 2022

Background: DNA vaccines fusing the chemokine MIP-3α (CCL20), which binds to CCR6 on immature dendritic cells (DCs), to melanoma-associated antigens have enhanced anti-tumor immunity and efficacy compared to vaccines lacking the chemokine. To further enhance the therapeutic effects of the vaccine, our laboratory has added type-I interferon (IFNα) and 5-Aza-2’-deoxycitidine (5Aza) to the therapy. Here, we report that the efficacy seen in this combination results in deeper intratumoral penetration of CD8+ cells, enhanced expression of Th1 t-cell related genes, and increased influx of CD11c+ CD8+ T-cells.

Methods: Beginning on day five post-transplantation of B16F10 melanoma, the vaccine was administered intramuscularly (i.m.) by electroporation. CpG adjuvant was given two days later. 5Aza was given intraperitoneally at 1mg/kg and IFNα therapy intratumorally as noted. Tumor sizes over time were assessed. For immunohistochemistry (IHC), formalin-fixed tumors were processed, embedded, cut into slides, and stained for CD8 by on-site core facility. Stained slides were analyzed by a pathologist in a blinded fashion. Tumor lysate gene expression levels and tumor-infiltrating lymphocytes (TILs) were assessed by qRT-PCR and flow cytometry respectively.

Results: Previous work has shown that the combination of IFNα, 5Aza, and vaccine led to significantly reduced tumor burden and overall increases in mouse survival dependent upon all three components. We have since discovered by IHC that CD8+ cells are more abundant and penetrate further into the tumor with the combination treatment compared to vaccine or therapeutics alone. The tumor immune genetic profile shows broad increases of expression for genes responsible for T-cell activation, efficacy, and trafficking. Of note, the combination therapy doubled the proportion of CD8+ T cells in the tumor microenvironment expressing CD11c (p = 0.027), and the proportions of these cells correlated more strongly to tumor size than do the proportions of CD11c- CD8+ T-cells (r2=0.51; p&lt;0.0001 compared to r2=0.19; p = 0.02).

Conclusions: Efficient targeting of antigen to immature dendritic cells with a chemokine-fusion vaccine offers a potential alternative approach to classic and dendritic cell-based vaccines. Combining this approach with IFNα and 5Aza treatments significantly improved vaccine efficacy. This treatment creates a complex intratumoral network that facilitates T-cell trafficking, Th1-associated gene expression, and an increase of CD11c+CD8+ T-cells, which have recently been shown in the literature to have greater anti-tumor activity than CD11c- T-cells. Further analysis of the pathways engaged by our treatment protocol should provide insights into how to more efficiently activate this recently described, highly effective T cell population.

Citation Format: James T. Gordy, Kaitlyn Comstock, Avinaash K. Sandhu, Yinan Hui, Styliani Karanika, Joel Sunshine, Petros C. Karakousis, Richard B. Markham. The addition of IFNα and 5-Aza-2'deoxycytidine treatments to a dendritic cell-targeting DNA vaccine enhance the expression of Th1-related genes and the infiltration of CD11c+ CD8+ T-cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB531.