The Effect of Vector Silencing during Picornavirus Vaccination against Experimental Melanoma and Glioma

Abstract

Virus vector-based vaccination against tumor-specific antigens remains a promising therapeutic approach to overcome the immune suppressive tumor microenvironment. However, the extent that the desired CD8 T cell response against the targeted tumor antigen is impacted by the CD8 T cell response against the virus vector is unclear. To address this question, we used picornavirus vaccination with Theiler’s murine encephalomyelitis virus (TMEV) as our vector against tumor-expressed ovalbumin (OVA257-264) antigen in both the B16-OVA murine melanoma and GL261-quad cassette murine glioma models. Prior to vaccination, we employed vector silencing to inhibit the CD8 T cell response against the immunodominant TMEV antigen, VP2121-130. We then monitored the resulting effect on the CD8 T cell response against the targeted tumor-specific antigen, ovalbumin. We demonstrate that employing vector silencing in the context of B16-OVA melanoma does not reduce tumor burden or improve survival, while TMEV-OVA vaccination without vector silencing controls tumor burden. Meanwhile, employing vector silencing during picornavirus vaccination against the GL261-quad cassette glioma resulted in a lower frequency of tumor antigen-specific CD8 T cells. The results of this study are relevant to antigen-specific immunotherapy, in that the virus vector-specific CD8 T cell response is not competing with tumor antigen-specific CD8 T cells. Furthermore, vector silencing may have the adverse consequence of reducing the tumor antigen-specific CD8 T cell response, as demonstrated by our findings in the GL261-quad cassette model.

Non-equivalent antigen presenting capabilities of dendritic cells and macrophages in generating brain-infiltrating CD8 + T cell responses

“…We next assessed the capacity for CD8 + T-cell priming during acute brain infection with recombinant TMEV encoding the model K b -restricted OVA (SIINFEKL) peptide 29 . Intracranial infection with TMEV-OVA generates a robust CD8 + T-cell response to OVA peptide presented in the context of the K b MHC I molecule 7 days post infection 5 , 29 . We intracranially infected CD11c-cre K b cKO, CMV-cre K b cKO, LysM-cre K b cKO, and cre-negative littermate controls with TMEV-OVA and isolated brain infiltrating lymphocytes 7 days post infection (Fig.…”

“…GL261 gliomas can be inoculated in immune-competent C57BL/6 mice, allowing for the study of adaptive immunotherapy 5 – 7 , 31 , 32 . A modified GL261 glioma line, termed GL261-quad cassette, expresses OVA peptide and enables analysis of K b :OVA-specific CD8 + T-cell responses 5 – 7 . We observed little effect of K b deletion on the naturally generated immune response to GL261-quad gliomas 21 days post tumor inoculation.…”

“…The GL261-quad cassette cell line was provided by the lab of John R. Ohlfest, PhD from the Masonic Cancer Center (University of Minnesota, Minneapolis, MN). The GL261-quad cassette cell line is engineered to express four model antigens: the K b -restricted OVA 257-264 , as well as OVA 323-339 , human GP100 25-33 , and the alloantigen I-E a 52-68 5 – 7 . The GL261-quad cassette cell line also expresses luciferase, which serves as a surrogate for tumor size via bioluminescence imaging.…”

“…The GL261-quad cassette cell line also expresses luciferase, which serves as a surrogate for tumor size via bioluminescence imaging. 6×10 4 GL261-quad cassette cells were implanted by stereotactic injection into the right striatum of each animal 5 – 7 . Prior to implantation, 8- to 12-week-old animals were anesthetized with 20 mg/kg ketamine and 5 mg/kg xylazine to minimize discomfort during the procedure.…”

“…GL261-quad cassette-bearing animals were assessed for tumor burden using bioluminescence imaging 5 – 7 . Animals received 150 mg/kg d -luciferin in PBS in a total volume of 200 µl intraperitoneally (i.p.)…”