SARS-CoV-2 RBD protein enhances the oncolytic activity of the vesicular stomatitis virus.

Despite recent advances in the research on oncolytic viruses (OVs), a better understanding of how to enhance their replication is key to improving their therapeutic index. Understanding viral replication is important to improve treatment outcomes based on enhanced viral spreading within the tumor milieu. The VSV-Δ51 oncolytic virus has been widely used as an anticancer agent with a high selectivity profile. In this study, we examined the role of the SARS-CoV-2 spike protein receptor-binding domain (RBD) in enhancing VSV-Δ51 viral production and oncolytic activity. To test this hypothesis, we first generated a novel VSV-Δ51 mutant that encoded the SARS-COV-2 RBD and compared viral spreading and viral yield between VSV-Δ51-RBD and VSV-Δ51 in vitro. Using the viral plaque assay, we demonstrated that the presence of the SARS-CoV-2 RBD in the VSV-Δ51 genome is associated with a significantly larger viral plaque surface area and significantly higher virus titers. Subsequently, using an ATP release-based assay, we demonstrated that the SARS-CoV-2 RBD could enhance VSV-Δ51 oncolytic activity in vitro. This observation was further supported using the B16F10 tumor model. These findings highlighted a novel use of the SARS-CoV-2 RBD as an anticancer agent.

Lung cancer and oncolytic virotherapy--enemy's enemy.

Lung cancer is one of the malignant tumors that seriously threaten human health worldwide, while the covid-19 virus has become people's nightmare after the coronavirus pandemic. There are too many similarities between cancer cells and viruses, one of the most significant is that both of them are our enemies. The strategy to take the advantage of the virus to beat cancer cells is called Oncolytic virotherapy. When immunotherapy represented by immune checkpoint inhibitors has made remarkable breakthroughs in the clinical practice of lung cancer, the induction of antitumor immunity from immune cells gradually becomes a rapidly developing and promising strategy of cancer therapy. Oncolytic virotherapy is based on the same mechanisms that selectively kill tumor cells and induce systemic anti-tumor immunity, but still has a long way to go before it becomes a standard treatment for lung cancer. This article provides a comprehensive review of the latest progress in oncolytic virotherapy for lung cancer, including the specific mechanism of oncolytic virus therapy and the main types of oncolytic viruses, and the combination of oncolytic virotherapy and existing standard treatments. It aims to provide new insights and ideas on oncolytic virotherapy for lung cancer.

A novel virotherapy encoding human interleukin-7 improves ex vivo T lymphocyte functions in immunosuppressed patients with septic shock and critically ill COVID-19.

A majority of patients with sepsis surviving the first days in intensive care units (ICU) enter a state of immunosuppression contributing to their worsening. A novel virotherapy based on the non-propagative Modified Virus Ankara (MVA) expressing the human interleukin-7 (hIL-7) cytokine fused to an Fc fragment, MVA-hIL-7-Fc, was developed and shown to enhance innate and adaptive immunity and confer survival advantages in murine sepsis models. Here, we assessed the capacity of hIL-7-Fc produced by the MVA-hIL-7-Fc to improve ex vivo T lymphocyte functions from ICU patients with sepsis. Primary hepatocytes were transduced with the MVA-hIL-7-Fc or an empty MVA, and cell supernatants containing the secreted hIL-7-Fc were harvested for in vitro and ex vivo studies. Whole blood from ICU patients [septic shock = 15, coronavirus disease 2019 (COVID-19) = 30] and healthy donors (n = 36) was collected. STAT5 phosphorylation, cytokine production, and cell proliferation were assessed upon T cell receptor (TCR) stimulation in presence of MVA-hIL-7-Fc-infected cell supernatants. Cells infected by MVA-hIL-7-Fc produced a dimeric, glycosylated, and biologically active hIL-7-Fc. Cell supernatants containing the expressed hIL-7-Fc triggered the IL-7 pathway in T lymphocytes as evidenced by the increased STAT5 phosphorylation in CD3+ cells from patients and healthy donors. The secreted hIL-7-Fc improved Interferon-γ (IFN-γ) and/or Tumor necrosis factor-α (TNF-α) productions and CD4+ and CD8+ T lymphocyte proliferation after TCR stimulation in patients with bacterial and viral sepsis. This study demonstrates the capacity of the novel MVA-hIL-7-Fc-based virotherapy to restore ex vivo T cells immune functions in ICU patients with sepsis and COVID-19, further supporting its clinical development.

Phage-Derived Oncolytic Viruses with 3C from Seneca Valley Virus for Targeted Therapy of Cervical Cancer

Cancer gene therapy based on various gene delivery vectors has some potential but also has obvious disadvantages. In this study, a new M13 phage-based oncolytic virus is constructed that carried the RGD peptides to target tumor cells and the 3C gene of Seneca Valley virus (SVV) preceded by a eukaryotic initial transcriptional region (ITR) to transcribe an oncolytic protein to kill tumor cells. Recombinant virus particles of 1200 nm in length are obtained in large quantities by transfecting the recombinant M13 phage plasmid into the host BL2738 and are investigated in vitro in tumor cells and in vivo in tumor-bearing mice to evaluate their antitumor effect. The experiments using Hela cells confirm that the engineered M13 phage can target and enter Hela cells, and express the SVV 3C protein, resulting in apoptosis of target cells by upregulating the expression of caspase 3. Furthermore, the results of experiments in vivo also show that the recombinant phage significantly inhibits the enhanced tumor volume in nude mice compared to the control groups. The M13 phage may be engineered to fuse with a variety of oncolytic proteins to inhibit the growth of tumor cells in the future, providing a promising phage-based targeted oncolytic reagent.

Redirection of anti-virus immunity towards anti-tumor immunity using nanoparticle-induced immune tolerance for oncolytic adenovirus therapy

Recent clinical observations that some coronavirus infections induced complete remissions in lymphoma patients emphasized again the potential of cancer virotherapy. Infection of cancer cells with oncolytic viruses reshapes the tumor microenvironment by activating anti-viral and anti-tumor immunity. A phase 1 clinical trial using oncolytic adenovirus Delta-24-RGD (DNX-2401) to treat recurrent malignant gliomas demonstrated activation of CD8+ T-cells and significant clinical benefits for a subset of patients. However, both anti-virus and anti-tumor immune responses are contingent on the activation of respective clones of CD8+ T-cells, which compete for clonal expansion. Thus, overexpansion of T-cells against viral antigens reduces the frequency of subdominant clones against tumor antigens. We hypothesized that inducing immune tolerance for viral antigens will decrease anti-viral immunity and in turn derepress anti-tumor immunity, resulting in enhanced efficacy of cancer virotherapy. In this work, we used nanoparticles encapsulating adenoviral antigens E1A, E1B and hexon that distributed to liver resident macrophages (P<0.0001) and induced peripheral immune tolerance. Functional experiments to restimulate immune cells with viral or tumor antigens showed that injection of nanoparticles induced virus-specific immune tolerance and redirected the focus of the immune response towards tumor peptides as measured by interferon-gamma secretion (P<0.0001). Co-culture experiments also showed increased activation of immune cells against fixed tumor cells after nanoparticle treatment (P<0.0001). Reduction of virus-specific T-cells and concurrent expansion of tumor-specific T-cell clones were further confirmed with E1A or OVA tetramers (P<0.05). Flow cytometry analysis suggested increased anti-tumor responses were due to differences in T-cell clones and not due to other immune populations including natural killer cells or myeloid-derived suppressor cells (P=0.3). Importantly, virotherapy in combination with nanoparticle-induced immune tolerance towards viral antigens in tumor-bearing mice increased the overall survival and doubled the percentage of long-term survivors compared to virus treatment alone. Our data should propel the development of a future clinical trial aiming to maximize the potential of anti-tumor immunity during cancer virotherapies.

Reshaping the tumor microenvironment with oncolytic viruses, positive regulation of the immune synapse, and blockade of the immunosuppressive oncometabolic circuitry.

BACKGROUND: Oncolytic viruses are considered part of immunotherapy and have shown promise in preclinical experiments and clinical trials. Results from these studies have suggested that tumor microenvironment remodeling is required to achieve an effective response in solid tumors. Here, we assess the extent to which targeting specific mechanisms underlying the immunosuppressive tumor microenvironment optimizes viroimmunotherapy. METHODS: We used RNA-seq analyses to analyze the transcriptome, and validated the results using Q-PCR, flow cytometry, and immunofluorescence. Viral activity was analyzed by replication assays and viral titration. Kyn and Trp metabolite levels were quantified using liquid chromatography-mass spectrometry. Aryl hydrocarbon receptor (AhR) activation was analyzed by examination of promoter activity. Therapeutic efficacy was assessed by tumor histopathology and survival in syngeneic murine models of gliomas, including Indoleamine 2,3-dioxygenase (IDO)-/- mice. Flow cytometry was used for immunophenotyping and quantification of cell populations. Immune activation was examined in co-cultures of immune and cancer cells. T-cell depletion was used to identify the role played by specific cell populations. Rechallenge experiments were performed to identify the development of anti-tumor memory. RESULTS: Bulk RNA-seq analyses showed the activation of the immunosuppressive IDO-kynurenine-AhR circuitry in response to Delta-24-RGDOX infection of tumors. To overcome the effect of this pivotal pathway, we combined Delta-24-RGDOX with clinically relevant IDO inhibitors. The combination therapy increased the frequency of CD8+ T cells and decreased the rate of myeloid-derived suppressor cell and immunosupressive Treg tumor populations in animal models of solid tumors. Functional studies demonstrated that IDO-blockade-dependent activation of immune cells against tumor antigens could be reversed by the oncometabolite kynurenine. The concurrent targeting of the effectors and suppressors of the tumor immune landscape significantly prolonged the survival in animal models of orthotopic gliomas. CONCLUSIONS: Our data identified for the first time the in vivo role of IDO-dependent immunosuppressive pathways in the resistance of solid tumors to oncolytic adenoviruses. Specifically, the IDO-Kyn-AhR activity was responsible for the resurface of local immunosuppression and resistance to therapy, which was ablated through IDO inhibition. Our data indicate that combined molecular and immune therapy may improve outcomes in human gliomas and other cancers treated with virotherapy.

Remission of liquid tumors and SARS-CoV-2 infection: A literature review.

The coronavirus disease 2019 (COVID-19) pandemic has produced a new global challenge for patients with cancer. The disease and the immunosuppression induced by cancer therapies have generated a perfect storm of conditions to increase the severity of the symptoms and worsen the prognosis. However, a few clinical reports showcased the power of viruses to induce remission in some patients suffering from liquid tumors. Here, we reviewed six cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that resulted in cancer remission, simultaneously highlighting the strengths and the unique challenges of oncolytic virotherapy. Virotherapy has become a special case of cancer immunotherapy. This paradigm-shifting concept suggests that oncolytic viruses are not only promising agents to combat particularly immunologically suppressed, immunotherapy-resistant tumors but also that the trigger of local inflammation, such as SARS-CoV-2 infection of the respiratory pathways, may trigger an abscopal effect sufficient to induce the remission of systemic cancer.

Understanding Immune Responses to Viruses-Do Underlying Th1/Th2 Cell Biases Predict Outcome?

Emerging and re-emerging viral diseases have increased in number and geographical extent during the last decades. Examples include the current COVID-19 pandemic and the recent epidemics of the Chikungunya, Ebola, and Zika viruses. Immune responses to viruses have been well-characterised within the innate and adaptive immunity pathways with the outcome following viral infection predominantly attributed to properties of the virus and circumstances of the infection. Perhaps the belief that the immune system is often considered as a reactive component of host defence, springing into action when a threat is detected, has contributed to a poorer understanding of the inherent differences in an individual's immune system in the absence of any pathology. In this review, we focus on how these host factors (age, ethnicity, underlying pathologies) may skew the T helper cell response, thereby influencing the outcome following viral infection but also whether we can use these inherent biases to predict patients at risk of a deviant response and apply strategies to avoid or overcome them.

A novel virotherapy encoding human interleukin 7 enhances ex vivo lymphocyte functions in immunosuppressed septic shock and critically ill COVID-19 patients

Introduction: After viral or bacterial sepsis, most intensive care unit (ICU) patients enter a state of profound immunosuppression contributing to patients' worsening. Transgene has developed an immunotherapy based on a viral vector encoding human interleukin-7 (hIL-7) to restore both innate and adaptive immune responses. Here, we assessed the capacity of hIL-7 to improve ex vivo T lymphocyte function from septic shock and COVID-19 patients. Methods: Primary human hepatocytes were transduced with MVAhIL- 7-Fc, a recombinant Modified Vaccinia virus Ankara (MVA) encoding the hIL-7 fused to the human IgG2 Fc fragment, or with empty MVA as control. Cell culture supernatants were harvested for further assays. T cells were collected from ICU patients (septic shock = 11, COVID- 19 = 29) and healthy donors (n = 21). STAT5 phosphorylation, cytokine production (ELISpot and intracellular staining) and cell proliferation were assessed upon TCR stimulation with supernatants containing or not hIL-7 produced after MVA transduction or with the counterpart recombinant hIL-7 (rhIL-7). Results: Patients with viral and bacterial sepsis display T lymphocyte alterations compared to healthy donors with a decreased production of cytokines and a decreased proliferation capacity. Supernatant containing hIL-7 induces STAT5 phosphorylation in CD3 lymphocytes of all patients. With 90% of responders, hIL-7 boosts cytokines production (single and double IFN-TNF) and T lymphocytes proliferation capacity at the same level as rhIL-7 in both cohorts whereas empty MVA has no effect. Conclusions: This study indicates that hIL-7-Fc produced after MVA transduction initiates IL-7 signaling through the phosphorylation of STAT5 and restores ex vivo human lymphocyte functions in cells from septic patients with acquired immunosuppression. This proof-of-concept study, along with experimental results in animal models, supports the clinical development of the MVA-hIL-7-Fc in sepsis immunosuppressed patients.

TP53 Synthetic Lethal Screen in Organoid Avatars to Discover Novel Therapeutic Targets for Colon Cancer

This application addresses the FY18 PRCRP topic area of Colorectal Cancer. Military relevant risk factors such as ionizing radiation often produce cancers with mutations in TP53. This proposal seeks to address a gap in colorectal cancer treatment that has profound impact on the health and well-being of military service Veterans, their beneficiaries, and extends to the American public as a whole. TP53 mutations are common in colon cancers (50 ), and most often lead to resistance to chemotherapy. A systematic screen for synthetic lethal interactions between gene knockout, TP53 mutation, and chemotherapy such as Cisplatin will produce novel targets useful for treating a large fraction of colon cancers.Hypothesis: Three-way synthetic

Combinatorial Treatment for Reach-and Grasp Recovery in Chronic Contusive Cervical SCI

Our main goals in this year were to (1) complete our experiments and prepare the data for a submission to a peer-reviewed journal, (2) move our lab and equipment from Miami, FL, to the Shirley Ryan Ability Lab, where the PI, Dr. Oudega had accepted a new position as per January 2020, (3) transfer the award administratively to the Shirley Ryan AbilityLab, (4) set up the lab and animal-related facilities, (5) write new IACUC protocols for the local committee at Northwestern University. We have accomplished all of these goals successfully in this last year. The lab and equipment were moved by the end of January. Lab set-up was started in February and then at first interrupted by COVID-19. Recently, the set-up was completed. Most time was spend to these moving and set-up efforts. Other time was spend to write animal protocols, and a manuscript for publication in a peer-reviewed journal. The manuscript is in its last phase of revisions befop0re submission.

MOLECULAR AND EPIDEMIOLOGICAL STUDY ON COVID-19 OVERALL AND IRAQ ESPECIALLY AL-ANBAR PROVINCE AS A MODEL

COVID-19 is the virus that engulfed the globe in the year 2020 that caused many deaths and disrupted countries' economies. In this report, we reviewed all aspects to explain the subjects, where we performed a systematic and thorough report of the Covid-19 outbreak, its medical conditions, its sources, diagnosis and prevention mechanisms and available methods of treatment. This also discussed the epidemiological situation in Iraq and the distribution of the province of Anbar as a sample for the report. Overall, from all of this analysis we conclude that epidemiology, virology, COVID-19 clinical is still enigmatic and that (viral) epidemics occasionally kill humanity, so we need a broader understanding of viruses.

Autophagy: The Potential Link between SARS-CoV-2 and Cancer.

COVID-19 infection survivors suffer from a constellation of symptoms referred to as post-acute COVID-19 syndrome. However, in the wake of recent evidence highlighting the long-term persistence of SARS-CoV-2 antigens in tissues and emerging information regarding the interaction between SARS-CoV-2 proteins and various components of the host cell macroautophagy/autophagy machinery, the unforeseen long-term consequences of this infection, such as increased risk of malignancies, should be explored. Although SARS-CoV-2 is not considered an oncogenic virus, the possibility of increased risk of cancer among COVID-19 survivors cannot be ruled out. Herein, we provide an overview of the possible mechanisms leading to cancer development, particularly obesity-related cancers (e.g., colorectal cancer), resulting from defects in autophagy and the blockade of the autophagic flux, and also immune escape in COVID-19 survivors. We also highlight the potential long-term implications of COVID-19 infection in the prognosis of patients with cancer and their response to different cancer treatments. Finally, we consider future directions for further investigations on this matter.

Adenovirus Type 6: Subtle Structural Distinctions from Adenovirus Type 5 Result in Essential Differences in Properties and Perspectives for Gene Therapy.

Adenovirus vectors are the most frequently used agents for gene therapy, including oncolytic therapy and vaccine development. It's hard to overestimate the value of adenoviruses during the COVID-19 pandemic as to date four out of four approved viral vector-based SARS-CoV-2 vaccines are developed on adenovirus platform. The vast majority of adenoviral vectors are based on the most studied human adenovirus type 5 (HAdV-C5), however, its immunogenicity often hampers the clinical translation of HAdV-C5 vectors. The search of less seroprevalent adenovirus types led to another species C adenovirus, Adenovirus type 6 (HAdV-C6). HAdV-C6 possesses high oncolytic efficacy against multiple cancer types and remarkable ability to induce the immune response towards carrying antigens. Being genetically very close to HAdV-C5, HAdV-C6 differs from HAdV-C5 in structure of the most abundant capsid protein, hexon. This leads to the ability of HAdV-C6 to evade the uptake by Kupffer cells as well as to distinct opsonization by immunoglobulins and other blood proteins, influencing the overall biodistribution of HAdV-C6 after systemic administration. This review describes the structural features of HAdV-C6, its interaction with liver cells and blood factors, summarizes the previous experiences using HAdV-C6, and provides the rationale behind the use of HAdV-C6 for vaccine and anticancer drugs developments.