Safe and effective “designer” adenoviruses target tumors after systemic administration.

The expression of cytokines and chemokines in response to adenovirus infection is tightly regulated by the innate immune system. Cytokine-mediated toxicity and cytokine storm are known clinical phenomena observed following naturally disseminated adenovirus infection in immunocompromised hosts as well as when extremely high doses of adenovirus vectors are injected intravenously. This dose-dependent, cytokine-mediated toxicity compromises the safety of adenovirus-based vectors and represents a critical problem, limiting their utility for gene therapy applications and the therapy of disseminated cancer, where intravenous injection of adenovirus vectors may provide therapeutic benefits. The mechanisms triggering severe cytokine response are not sufficiently understood, prompting efforts to further investigate this phenomenon, especially in clinically relevant settings. In this review, we summarize the current knowledge on cytokine and chemokine activation in response to adenovirus- and adenovirus-based vectors and discuss the underlying mechanisms that may trigger acute cytokine storm syndrome. First, we review profiles of cytokines and chemokines that are activated in response to adenovirus infection initiated via different routes. Second, we discuss the molecular mechanisms that lead to cytokine and chemokine transcriptional activation. We further highlight how immune cell types in different organs contribute to synthesis and systemic release of cytokines and chemokines in response to adenovirus sensing. Finally, we review host factors that can limit cytokine and chemokine expression and discuss currently available and potential future interventional approaches that allow for the mitigation of the severity of the cytokine storm syndrome. Effective cytokine-targeted interventional approaches may improve the safety of systemic adenovirus delivery and thus broaden the potential clinical utility of adenovirus-based therapeutic vectors.

The increased cancer incidence and associated mortality call for the development of novel treatment modalities to address needs of cancer patients, which do not respond to existing therapies. Oncolytic viruses emerge as a new modality in cancer therapy due to their natural ability to infect and kill cancer cells, and to stimulate patient's own immune system to eliminate cancer cells throughout the body. For this study, we used novel engineered oncolytic adenovirus, Ad5/35-3M (AVID-317), which demonstrated greatly-improved safety profile after systemic virus administration due to a set of targeted mutations in the adenovirus capsid (Atasheva et al, Science Translational Medicine, 2020, eabc6659). Systemic administration of AVID-317 to NCr/Nude mice with disseminated lung adenocarcinoma slowed tumor progression and significantly extended survival, compared to buffer-treated control tumor-bearing mice. In AVID-317 treated group, about 25% of mice demonstrated complete rejection of disseminated tumors and the rest of mice showed significantly extended survival. To understand the mechanism of AVID-317-mediated tumor suppression and to design more effective combination therapies, we analyzed disseminated tumors from the lungs in NCr/Nude mice using single cell RNA-seq and Mass Cytometry (Cy-TOF) after virotherapy. Our scRNA-seq analysis showed that in a subset of mice responding poorly to AVID-317 therapy, tumor-associated CSF1R-expressing myeloid cells expressed higher levels of MHC-II, Ctsb, and Arg1. In addition, CD3e+ T cells expressed functional exhaustion marker, PD-1, exclusively in mice that responded poorly to virotherapy. Mass Cytometry analysis with 37 metal isotope-labeled antibodies confirmed elevated expression of activation markers on myeloid cells after virotherapy. Next, we analyzed the efficacy of a combination therapy where in addition to AVID-317 administration, tumor-bearing mice were treated with anti PD-1 antibody to block immune-suppressive signaling in T cells, anti-CSF1R antibody to block CSF1R signaling, and anti-Asialo GM1 antibody to deplete anti-viral NK cells. Our analyses showed that combination therapy regimen significantly improved therapeutic outcomes in mice with disseminated lung cancer, by extending median survival to 95 days from 55-56 days observed in mice treated with either virus alone or combination drugs without the virus. Taken together, our study demonstrated feasibility of using AVID-317 oncolytic adenovirus for systemic therapy of disseminated lung cancer. Our study also showed that a combination of virotherapy with a defined select of immunotherapeutic drugs allows for the significant improvement of therapeutic efficacy in pre-clinical model of disseminated lung cancer. Citation Format: Jia Yao, Svetlana Atasheva, Dmitry Shayakhmetov. Novel oncolytic adenovirus for systemic treatment of disseminated lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3366.

The majority of lung cancer patients do not respond to current immune-oncology drugs due to low PD1 and PD-L1 expression on immune and cancer cells. Oncolytic viruses are being actively explored as an alternate modality for cancer therapy due to their natural ability to kill infected cells through viral replication. In this study, we analyzed the anti-tumor efficacy and safety profile of AVID-317 oncolytic virus in pre-clinical mouse models of disseminated lung cancer. AVID-317 is a novel oncolytic adenovirus possessing a set of mutations in the virus capsid, which ablate virus interactions with cellular b3 integrins on macrophages and hepatic cells, resulting in virus de-targeting from the liver after intravenous administration. In vitro studies demonstrated that 70% of tested human non-small cell lung cancer (NSCLC)-derived cell lines (N=16) are sensitive to AVID-317 infection, and 85% of AVID-317-sensitive NSCLC lines express no or low PD-L1. Intravenous injection of control parental human adenovirus type 5 (HAdv5) at a dose of 1e11 v.p. per mouse into wild-type mice resulted in release of pro-inflammatory cytokines IL-1b, IL-6, IL-16, and G-CSF into the bloodstream at 6 hours post virus administration. Furthermore, this dose of HAdv5 was lethal due to severe hepatotoxicity within 48 h post virus injection. In contrast, intravenous administration of AVID-317 in the range of doses up to 1e12 v. p. per mouse resulted in no detectible IL-6 and significantly reduced IL-1b, IL-16, G-CSF amounts in the blood. Importantly, AVID-317 administration did not result in mortality. Next, we grafted PD-L1-low human adenocarcinoma A549-Luc-C8 cells into lungs of NCr nude mice to establish an orthotopic model of disseminated lung cancer. When tumor burden reached between 2xe6 to 8e6 RLU, mice were enrolled in four cohorts and treated with AVID-317 at doses of 5e10, 1e11, or 3e11 v.p. per mouse or administered with saline only (control group; N=>10 in each cohort). Mice were euthanized upon detection of 20% weight loss compared to baseline. The median survival in saline-treated group was 57 days post treatment. However, median survival has not be reached for AVID-317-treated groups for the duration of the study (80 days). Taken together, our study demonstrates feasibility of using AVID-317 oncolytic adenovirus for systemic therapy of disseminated lung cancer and its greatly improved safety profile observed upon intravenous virus delivery. Citation Format: Svetlana Atasheva, Jia Yao, Cedrick Young, Nelson C. Di Paolo, Henry Wyche, Dmitry M. Shayakhmetov. Preclinical assessment of efficacy and safety of novel oncolytic adenovirus for therapy of disseminated lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4781.