1134 A novel mRNA therapeutic vaccine for polyomavirus associated merkel cell carcinoma

Field: Medicine (Health Sciences)

Background

Merkel Cell Carcinoma (MCC) is a rare cutaneous neoplasm of neuroendocrine origin associated with integration of a truncated form of the Merkel Cell Polyomavirus Large T Antigen (LTA) in ~80% of U.S. cases.1 LTA is both antigenic and requisite for continued proliferation of MCC, making it an attractive target for a therapeutic vaccine strategy.2

Methods

Expression of truncated LTA was induced in a B16-F10 melanoma cell line to create an in vivo murine MCC model. Modified mRNA coding for LTA was produced using in vitro transcription and was packaged in lipid nanoparticles for delivery by intramuscular injection. LTA expressing tumors were placed subcutaneously in C57BL/6 mice and treated with LTA vaccine or placebo with or without additional anti-PD1 antibody. Tumor growth and survival were measured and characterization of tumor immune infiltration by flow cytometry was performed. PBMCs and matched tumor cells from MCC patients were used to establish a model of in vitro vaccination. Monocyte derived dendritic cells (moDCs) were generated from the PBMC pool and were transfected with LTA mRNA. The PBMC pool was pulsed with antigen loaded or placebo moDCs every 7 days, and immunophenotypes, cytokine release, and specific tumor cell killing were assessed.

Results

LTA mRNA treatment resulted in tumor growth suppression compared to placebo by day 21 (mean volume 199.8mm3 vs. 830.0mm3 P=.007 figure 1A) and increased median survival (35.5 vs. 25.5 days P=.004 figure 1A). Combination of vaccine with anti-PD1 outperformed monotherapy with growth suppression on day 21 (mean volume 103.2mm3 vs. 398.7mm3 P=.037 figure 1B). Treated murine tumors exhibited increased immune infiltration (mean CD45+ 11.56% vs. 7.81% P=.037 figure 1C). Among infiltrating lymphocytes, there was an increased proportion of CD3+ cells (mean 69.74% vs. 57.36% P=.0013 figure 1D), and CD8+ cells were more cytotoxic (MFI GZMB 26785 vs. 13607 P=.0026 figure 1E). In vitro vaccination of patient PBMCs resulted in expansion of CD8+ cells by day 35 (42.2% vs. 22.2% P=.0013 figure 2B), and an increase in activation and memory markers (PD1+ 13.9% vs. 3.91% P=.01; CD45RO+ 77.5% vs. 39.6% P=.009 figure 2C). Exposure of treated cells to antigen loaded DCs resulted in increased IFNg release (422.9pg/mL vs. 96.16pg/mL P<.0001 figure 2D) and increased specific tumor killing (Mean fluorescent count 12 vs. 9.5 P=.038 figure 2E).

Conclusions

Treatment with a novel mRNA vaccine targeting LTA increases infiltration of cytotoxic immune populations, resulting in significant growth suppression and increased median survival. In vitro vaccination increases cytotoxic CD8+ populations, IFNg release, and specific killing of patient derived tumor cells.

References

Paulson KG, Park SY, Vandeven NA, Lachance K, Thomas H, Chapuis AG, Harms KL, Thompson JA, Bhatia S, Stang A, Nghiem P. Merkel cell carcinoma: Current US incidence and projected increases based on changing demographics. J Am Acad Dermatol. 2018 Mar;78(3):457–463. Houben R, Adam C, Baeurle A, Hesbacher S, Grimm J, Angermeyer S, Henzel K, Hauser S, Elling R, Bröcker EB, Gaubatz S, Becker JC, Schrama D. An intact retinoblastoma protein-binding site in Merkel cell polyomavirus large T antigen is required for promoting growth of Merkel cell carcinoma cells. Int J Cancer. 2012 Feb 15;130(4):847–56.

Ethics Approval

Tumors were collected with the approval of the Yale University Institutional Review Board, IRB # 0609001869, approval date: 7/19/2022, and expiration Date: 7/18/2023. Participants gave informed consent before taking part. Animal studies were conducted with the approval of the Yale University Institutional Animal Care and Use Committee, protocol #2022–20307