Chikungunya virus (CHIKV) is a re-emerging mosquito-borne Alphavirus that causes a clinical disease involving fever, myalgia, nausea and rash. The distinguishing feature of CHIKV infection is the severe debilitating poly-arthralgia that may persist for several months after viral clearance. Since its re-emergence in 2004, CHIKV has spread from the Indian Ocean region to new locations including metropolitan Europe, Japan, and even the United States. The risk of importing CHIKV to new areas of the world is increasing due to high levels of viremia in infected individuals as well as the recent adaptation of the virus to the mosquito species Aedes albopictus. CHIKV re-emergence is also associated with new clinical complications including severe morbidity and, for the first time, mortality. In this study, we characterized disease progression and host immune responses in adult and aged Rhesus macaques infected with either the recent CHIKV outbreak strain La Reunion (LR) or the West African strain 37997. Our results indicate that following intravenous infection and regardless of the virus used, Rhesus macaques become viremic between days 1–5 post infection. While adult animals are able to control viral infection, aged animals show persistent virus in the spleen. Virus-specific T cell responses in the aged animals were reduced compared to adult animals and the B cell responses were also delayed and reduced in aged animals. Interestingly, regardless of age, T cell and antibody responses were more robust in animals infected with LR compared to 37997 CHIKV strain. Taken together these data suggest that the reduced immune responses in the aged animals promotes long-term virus persistence in CHIKV-LR infected Rhesus monkeys.

A series of investigational oncolytic immunotherapies based on DNX-2401 (tasadenoturev) are under development for the treatment of advanced cancers. Viruses “armed” with cell surface or secreted immune modulatory molecules inserted in the DNX-2401 backbone were evaluated using human cell derived 3D microtumor/ T cell models to elucidate specific anti-tumor properties and determine differences in immune activation. Background: DNX-2401 is a conditionally replicative adenovirus serotype 5 containing: 1) a 24 bp deletion from the E1A gene (Rb-binding domain) to confer cancer cell selectivity, and 2) insertion of an integrin-binding motif (RGD-4C) in the fiber to enhance infectivity across a broad range of cancers. DNX-2401 selectively replicates in cancer cells, causes tumor destruction, and triggers the recruitment of lymphocytes, leading to anti-tumor response. Phase 1 and 2 studies of DNX-2401 are ongoing in patients with glioblastoma and diffuse intrinsic pontine glioma. The first immune modified virus in the series, DNX-2440, engineered to express OX40L, is being evaluated in a Phase 1 study in patients with glioblastoma. Results: Immune modulator coding sequences inserted in place of E3 in the DNX-2401 backbone expressed properly localized and biologically active transgenes. DNX-2440 infected tumor cells expressed OX40L localized to the cell surface capable of stimulating OX40-expressing T cells. Tumor cells infected with a second virus, which was engineered to express a cytokine, secreted high levels of active protein. Both armed viruses showed infection, replication, and oncolytic activity in a panel of cancer cell lines under standard culture conditions. 3D microtumors composed of melanoma or lung cancer cells and normal human dermal fibroblasts were utilized to assess immune activation following virus infection. Unstimlated or pre-stimulated human immune cells were co-cultured for 12 days with the 3D microtumors infected with DNX-2401 or each armed virus. Infection with DNX-2440 increased T cell-mediated killing of tumor cells that was dependent on OX40/OX40L engagement, as the effect was abrogated in the presence of an OX40L blocking antibody. The kinetics of oncolytic activity, immune cell activation, and tumor infiltration were dependent on the immune modulator expressed from the oncolytic virus, resulting in differences in tumor clearance. These results demonstrate how oncolytic viruses can be customized to have unique properties by inserting different immune modulators into the virus genome, thus providing a customized approach for treating different cancers. Citation Format: Laura K. Springgay, Amy M. Strauwald, Brett Ewald, Joan M. Robbins, Winnie M. Chan. Oncolytic adenoviruses expressing immune modulators enhance tumor cell killing in human cancer 3D microtumor models [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6713.

Interferon (IFN) production and the subsequent induction of IFNstimulated genes (ISGs) are highly effective innate strategies utilized by cells to protect against invading pathogens, including viruses. Critical components involved in this innate process are promyelocytic leukemia nuclear bodies (PML-NBs), which are subnuclear structures required for the development of a robust IFN response. As such, PML-NBs serve as an important hurdle for viruses to overcome to successfully establish an infection. Both Kaposi's sarcoma-associated herpesvirus (KSHV) and the closely related rhesus macaque rhadinovirus (RRV) are unique for encoding viral homologs of IFN regulatory factors (termed vIRFs) that can manipulate the host immune response by multiple mechanisms. All four KSHV vIRFs inhibit the induction of IFN, while vIRF1 and vIRF2 can inhibit ISG induction downstream of the IFN receptor. Less is known about the RRV vIRFs. RRV vIRF R6 can inhibit the induction of IFN by IRF3; however, it is not known whether any RRV vIRFs inhibit ISG induction following IFN receptor signaling. In our present study, we demonstrate that the RRV vIRF R12 aids viral replication in the presence of the type I IFN response. This is achieved in part through the disruption of PML-NBs and the inhibition of robust ISG transcription. IMPORTANCE KSHV and RRV encode a unique set of homologs of cellular IFN regulatory factors, termed vIRFs, which are hypothesized to help these viruses evade the innate immune response and establish infections in their respective hosts. Our work elucidates the role of one RRV vIRF, R12, and demonstrates that RRV can dampen the type I IFN response downstream of IFN signaling, which would be important for establishing a successful infection in vivo.