Multimodal Reprogramming of the Tumor Microenvironment by MMR and Dual Checkpoint Blockade in Hepatocellular Carcinoma Models

Mulu Z Tesfay^1*Aleksandra Cios²Khandoker Usran Ferdous¹Randal S Shelton¹Bahaa Mustafa¹Camila C Simoes¹Murat Gokden¹Isabelle R. Miousse¹Kimberly J Krager¹Marjan Boerma¹Alicja Urbaniak¹Anuradha Kunthur¹Jeevani Obulareddy¹Joshua M Eichhorn¹Steven R Post¹Jean Christopher Chamcheu³Omeed Moaven⁴Yves Chiswili Chabu⁵Dan G Duda⁶Matteo Conti⁷Bruno Nardo⁸Rang Govindarajan¹Martin E Fernandez-Zapico⁹Lewis R Roberts⁹Mitesh Borad⁹Martin John Cannon¹Alexei Basnakian¹Bolni M Nagalo^2*

• ¹University of Arkansas for Medical Sciences, Little Rock, United States
• ²University of Maryland Baltimore, Baltimore, United States
• ³The University of Louisiana Monroe College of Pharmacy, Monroe, United States
• ⁴LSU Health New Orleans Stanley S Scott Cancer Center, New Orleans, United States
• ⁵University of Missouri, Columbia, United States
• ⁶Harvard University, Cambridge, United States
• ⁷Azienda Unita Sanitaria Locale di Imola, Imola, Italy
• ⁸Universita della Calabria, Arcavacata di Rende, Italy
• ⁹Mayo Clinic Minnesota, Rochester, United States

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide, thus, there is an urgent need to develop more effective therapeutic options for this dismal condition. Tumor-infiltrating lymphocytes (TILs) are associated with improved response to immune checkpoint blockade in HCC, but their low abundance in most cases limits their therapeutic efficacy. Here, we demonstrate, in mice, that low-dose intratumoral immunovirotherapy with the trivalent measles, mumps, and rubella vaccine (MMR) induces superior tumor-growth delay and extended host survival compared to individually administered vaccines for measles, mumps, or rubella viruses. Further, our results show that MMR therapy synergizes with PD-1 and CTLA-4 blockade to reprogram the tumor microenvironment, resulting in increased CD8+ TIL infiltration and reduced PD-1 expression on TILs, among other effects. These changes in the immunological landscape translated into greater survival and more durable tumor-specific and memory immune responses for hosts. Comprehensive toxicology analysis revealed no evidence of MMR-induced liver or kidney toxicity after intrahepatic administration. This work reinforces an unrecognized role of MMR plus ICB in reprogramming the immune landscape in HCC through multimodal immune activation, providing a strong rationale for further development of MMR-based therapies for HCC.