25: Combination Immunotherapy with Oncolytic Virus Induces Immunogenic Cell Death and T-cell Proliferation in Colon Cancer

Introduction: We discovered that forced expression of TNFSF14 (LIGHT) in preclinical models of advanced colon cancer results in tumor regression and increased T-cell activation, proliferation, and trafficking. Adenoviral vectors have been constructed to deliver LIGHT into the tumor microenvironment, however, the optimal virus to perform this function is unclear. Whether a replication competent oncovirus or replication deficient virus can both deliver LIGHT and induce immunogenic cell death (ICD) in colon cancer remains to be determined.


Methods: Human oncolytic adenoviruses containing LIGHT (Ad-LIGHT^onco) and not containing LIGHT (Ad-Null^onco) were constructed with a tumor specific promoter allowing for selective replication competence. Replication-deficient (RD) adenoviruses containing LIGHT (Ad-LIGHT^RD) and not containing LIGHT (Ad-Null^RD) were generated for comparison. CT26 murine colon cancer cells expressing the human coxsackie and adenovirus receptor (hCAR) were utilized. The hallmarks of ICD, calreticulin translocation and HMGB1 secretion, were quantified by flow cytometric assessment and ELISA, respectively.


Results: Colon cancer cells treated with tumor-selective, replication competent Ad-Null^onco and Ad-LIGHT^onco demonstrated greater apoptosis, necrosis, and necroptosis measured by Annexin V and propidium iodide staining, compared to RD treated and control cells. Calreticulin translocation was significantly increased at 48 hours in oncoviral treated cancer cells compared to RD virus treated and control cells (Ad-LIGHT^onco: 11.9%, Ad-LIGHT^RD: 1.4%, control: 0.5%; p< 0.005). Significant increases in secreted HMGB1 levels were identified following infection with Ad-Null^onco and Ad-LIGHT^onco (Figure). In a coculture of syngeneic CD3+ T-cells and cancer cells there was ~3-fold increase in T-cell proliferation, measured by ³H-thymidine incorporation, in LIGHT-expressing oncolytic virus treated cells compared to controls (Figure).


Conclusions: Oncolytic virus treatment of colon cancer cells induced significant immunogenic cells death not seen with standard adenoviral treatment. Further, viral-delivered LIGHT protein induced impressive T-cell proliferation. Thus, a tumor-selective, replication competent oncolytic virus can deliver new immunotherapies into the tumor microenvironment while simultaneously inducing ICD that may augment the antitumor immune response.