Oncolysis of malignant human melanoma tumors by Coxsackieviruses A13, A15 and A18

Many RNA viruses are displaying great promise in the field of oncolytic virotherapy. Previously, we reported that the picornavirus Coxsackievirus A21 (CVA21) possessed potent oncolytic activity against cultured malignant melanoma cells and melanoma xenografts in mice. In the present study, we demonstrate that three additional Group A Coxsackieviruses; Coxsackievirus A13 (CVA13), Coxsackievirus A15 (CVA15) and Coxsackievirus A18 (CVA18), also have similar oncolytic activity against malignant melanoma. Each of the viruses grew quickly to high titers in cancer cells expressing ICAM-1 and intratumoral injection of preformed subcutaneous SK-Mel-28 xenografts in mice with CVA13, CVA15 and CVA18 resulted in significant tumor volume reduction

Adaptive mutations in the genomes of enterovirus 71 strains following infection of mouse cells expressing human P-selectin glycoprotein ligand-1

We recently identified human P-selectin glycoprotein ligand-1 (PSGL-1) as a functional enterovirus 71 (EV71) receptor and demonstrated PSGL-1-dependent replication for some EV71 strains in human leukocytes. Here, we report that four out of five PSGL-1-binding strains of EV71 replicated poorly in mouse L929 cells stably expressing human PSGL-1 (L-PSGL-1 cells). Therefore, we compared the replication kinetics and entire genomic sequence of five original EV71 strains and the corresponding EV71 variants (EV71-LPS), which were propagated once in L-PSGL-1 cells. Direct sequence comparison of the entire genome of the original EV71 strains and EV71-LPS variants identified several possible adaptive mutations during the course of replication in L-PSGL-1 cells, including a putative determinant of the adaptive phenotype in L-PSGL-1 cells at VP2-149. The results suggest that an adaptive mutation, along with a PSGL-1-binding phenotype, may facilitate efficient PSGL-1-dependent replication of the EV71 strains in L-PSGL-1 cells

A gene signature for post-infectious chronic fatigue syndrome

Background: At present, there are no clinically reliable disease markers for chronic fatigue syndrome. DNA chip microarray technology provides a method for examining the differential expression of mRNA from a large number of genes. Our hypothesis was that a gene expression signature, generated by microarray assays, could help identify genes which are dysregulated in patients with post-infectious CFS and so help identify biomarkers for the condition. Methods: Human genome-wide Affymetrix GeneChip arrays (39,000 transcripts derived from 33,000 gene sequences) were used to compare the levels of gene expression in the peripheral blood mononuclear cells of male patients with post-infectious chronic fatigue (n = 8) and male healthy control subjects (n = 7). Results: Patients and healthy subjects differed significantly in the level of expression of 366 genes. Analysis of the differentially expressed genes indicated functional implications in immune modulation, oxidative stress and apoptosis. Prototype biomarkers were identified on the basis of differential levels of gene expression and possible biological significance Conclusion: Differential expression of key genes identified in this study offer an insight into the possible mechanism of chronic fatigue following infection. The representative biomarkers identified in this research appear promising as potential biomarkers for diagnosis and treatment

Oncolytic Viruses for Cancer Therapy: Overcoming the Obstacles

Targeted therapy of cancer using oncolytic viruses has generated much interest over the past few years in the light of the limited efficacy and side effects of standard cancer therapeutics for advanced disease. In 2006, the world witnessed the first government-approved oncolytic virus for the treatment of head and neck cancer. It has been known for many years that viruses have the ability to replicate in and lyse cancer cells. Although encouraging results have been demonstrated in vitro and in animal models, most oncolytic viruses have failed to impress in the clinical setting. The explanation is multifactorial, determined by the complex interactions between the tumor and its microenvironment, the virus, and the host immune response. This review focuses on discussion of the obstacles that oncolytic virotherapy faces and recent advances made to overcome them, with particular reference to adenoviruses

Plasmacytoid dendritic cells orchestrate innate and adaptive anti-tumor immunity induced by oncolytic coxsackievirus A21

Background: The oncolytic virus, coxsackievirus A21 (CVA21), has shown promise as a single agent in several clinical trials and is now being tested in combination with immune checkpoint blockade. Combination therapies offer the best chance of disease control; however, the design of successful combination strategies requires a deeper understanding of the mechanisms underpinning CVA21 efficacy, in particular, the role of CVA21 anti-tumor immunity. Therefore, this study aimed to examine the ability of CVA21 to induce human anti-tumor immunity, and identify the cellular mechanism responsible. Methods: This study utilized peripheral blood mononuclear cells from i) healthy donors, ii) Acute Myeloid Leukemia (AML) patients, and iii) patients taking part in the STORM clinical trial, who received intravenous CVA21; patients receiving intravenous CVA21 were consented separately in accordance with local institutional ethics review and approval. Collectively, these blood samples were used to characterize the development of innate and adaptive anti-tumor immune responses following CVA21 treatment. Results: An Initial characterization of peripheral blood mononuclear cells, collected from cancer patients following intravenous infusion of CVA21, confirmed that CVA21 activated immune effector cells in patients. Next, using hematological disease models which were sensitive (Multiple Myeloma; MM) or resistant (AML) to CVA21-direct oncolysis, we demonstrated that CVA21 stimulated potent anti-tumor immune responses, including: 1) cytokine-mediated bystander killing; 2) enhanced natural killer cell-mediated cellular cytotoxicity; and 3) priming of tumor-specific cytotoxic T lymphocytes, with specificity towards known tumor-associated antigens. Importantly, immune-mediated killing of both MM and AML, despite AML cells being resistant to CVA21-direct oncolysis, was observed. Upon further examination of the cellular mechanisms responsible for CVA21-induced anti-tumor immunity we have identified the importance of type I IFN for NK cell activation, and demonstrated that both ICAM-1 and plasmacytoid dendritic cells were key mediators of this response. Conclusion: This work supports the development of CVA21 as an immunotherapeutic agent for the treatment of both AML and MM. Additionally, the data presented provides an important insight into the mechanisms of CVA21-mediated immunotherapy to aid the development of clinical biomarkers to predict response and rationalize future drug combinations

The Actin Cytoskeleton as a Barrier to Virus Infection of Polarized Epithelial Cells

Many diverse viruses target a polarized epithelial monolayer during host invasion. The polarized epithelium is adept at restricting the movement of solutes, ions, macromolecules, and pathogens across the mucosa. This regulation can be attributed to the presence of a junctional complex between adjacent cells and to an intricate network of actin filaments that provides support to the subapical membrane and stabilizes intercellular junctions. It is therefore not surprising that many viruses have evolved highly varied strategies to dissolve or modulate the cortical actin meshwork to promote infection of polarized cells. In this review, we will discuss the cell biological properties of the actin cytoskeleton in polarized epithelial cells and review the known mechanisms utilized by viral pathogens to manipulate this system in order to facilitate their infection

Echovirus 7 Entry into Polarized Intestinal Epithelial Cells Requires Clathrin and Rab7

Enteroviruses invade the host by crossing the intestinal mucosa, which is lined by polarized epithelium. A number of enteroviruses, including echoviruses (EV) and group B coxsackieviruses (CVB), initiate infection by attaching to decay-accelerating factor (DAF), a molecule that is highly expressed on the apical surface of polarized epithelial cells. We previously observed that entry of DAF-binding CVB3 into polarized intestinal epithelial cells occurs by an unusual endocytic mechanism that requires caveolin but does not involve clathrin or dynamin. Here we examined the entry of a DAF-binding echovirus, EV7. We found that drugs, small interfering RNAs (siRNAs), and dominant negative mutants that target factors required for clathrin-mediated endocytosis, including clathrin and dynamin, inhibited both EV7 infection and internalization of virions from the cell surface. Once virus had entered the cell, it colocalized with markers of early endosomes (EEA1) and then late endosomes (LAMP-2). Inhibition of endosomal maturation—with siRNAs or dominant negative mutants targeting Rab5 and Rab7—inhibited infection and prevented release of viral RNA into the cell. These results indicate that EV7 is internalized by clathrin-mediated endocytosis and then moves to early and late endosomes before releasing its RNA. Trafficking through endosomes is known to be important for viruses that depend on low pH or endosomal cathepsin proteases to complete the entry process. However, we found that EV7 infection required neither low pH nor cathepsins

Validity of the Common Cold Questionnaire (CCQ) in Asthma Exacerbations

Background: The common cold questionnaire (CCQ) is used to discriminate those with and without a viral infection. Its usefulness in people with acute asthma is unknown. Our aim was to asess the ability of the CCQ to detect viral infection and to monitor recovery during a viral induced asthma exacerbation and confirmed by virological testing. Methodology/Principal Findings: We studied subjects (≥7 yrs) admitted to hospital with acute asthma and diagnosed as positive (n=63), or negative to viral infection (n=27) according to molecular and virological testing from respiratory samples. CCQ asthma history and asthma control questionaire were completed and repeated 4-6 weeks later. Sensitivity specificity, and response to change of the CCQ were assessed by receiver operator curve (ROC) analysis and effect size calculation respectively. The CCQ did not discriminate between viral and non-viral infection for subjects with asthma (sensitivity = 76.2%; specificity = 29.6%). ROC analysis could not differentiate between positive or negative virus in subjects with asthma. The CCQ had a large responce to change following recovery (effect size = 1.01). 39% of subjects recovering from viral exacerbation remained positive to virological testing at follow-up despite improvement in clinical symptoms. The CCQ reflected clinical improvement in these subjects, thus providing additional information to complement virological testing. Conclusions/Significance: The CCQ is a useful instrument for monitorong response to viral infection in people with asthma. Reliable differentiation between viral and non-viral asthma exacerbations was not achieved with the CCQ and requires specific virological testing. When combined with virological testing, the CCQ should be a useful outcome measure for evaluating therapies in viral-induced asthma