HSV-1 Virus Research Articles

Chelerythrine inhibits HSV-1 infection by suppressing virus binding to the cells

ObjectiveHerpes simplex virus type 1 (HSV-1) is a worldwide pathogen and mainly puts immunocompromised patients at higher risk of various complications. This research aims to demonstrate the antiviral activity of chelerythrine (CHE), an alkaloid compound derived from the traditional Chinese medicine Mcleaya cordata, towards the HSV-1 F strain. MethodsBioassay-guided fractionation was carried out in order to isolate the antiviral compound CHE. The plaque reduction assay was used to evaluate the antiviral activity of CHE. The combination of quantitative PCR (qPCR) and western blot was used to elucidate whether CHE interferes with viral DNA synthesis, followed by time-of-addition assay to examine which steps of HSV-1 infection CHE affected. The immunofluorescent assay and transmission electron microscopy assay were performed to further analyze the intracellular localization and the ultrastructural features of HSV-1 virions with the treatment of CHE. The binding affinity in CHE to targeted viral glycoprotein was investigated by surface plasmon resonance (SPR) assay and docking simulation. ResultsCHE displayed a significant antiviral effect against HSV-1 at 0.312 to 2.5 µM with a viral titer reduction of 29.38% to 61.02%. CHE had an EC50 value of 1.78 µM against HSV-1, and the selectivity index (SI) for CHE was 4.07. Additionally, CHE inhibits ICP0 expression in the immediate early stage of infection, which further affected the early stage of infection, and subsequent viral replication. Results of the SPR assay showed that CHE had strong-binding affinity to gB with a KD value of 7.80 × 107 M, while gD with a value of 1.18 × 106 M. TEM revealed that CHE can clearly inhibit the virion release. In silico experiments further confirmed that CHE had low binding free energy in the activity sites of gB and gD. ConclusionThis study revealed that CHE directly targeted viral gB or gD as a potent glycoprotein inhibitor, which suggested that CHE could be a virus entry inhibitor for HSV-1 treatment.

Statistical optimization and characterization of fucose-rich polysaccharides extracted from pumpkin (Cucurbita maxima) along with antioxidant and antiviral activities

Biologically active phytochemicals from pumpkin reveal versatile medical applications, though little is known about their antiviral activity. The fucose-rich polysaccharide extraction conditions were optimized through Box-Behnken design and purified by column chromatography. The purified fucose-rich polysaccharide was characterized through SEM, FT-IR, 1H NMR, XRD, TGA, and GS-MS. The analysis results revealed an irregular and porous surface of the purified polysaccharide with high fucose, rhamnose, galactose, and glucose contents. The tested fucose-rich polysaccharides revealed significant antioxidant and anti-inflammatory activity at very low concentrations. The purified fucose-rich polysaccharides exerted a broad-spectrum antiviral activity against both DNA and RNA viruses, accompanied by high safety toward normal cells, where the maximum safe doses (EC100) were estimated to be about 3–3.9 mg/mL for both Vero and PBMC cell lines. Treatment of HCV, ADV7, HSV1, and HIV viruses with the purified polysaccharides showed a potent dose-dependent inhibitory activity with IC50 values of 95.475, 20.96, 5.213, and 461.75 μg/mL, respectively. This activity was hypothesized to be through inhibiting the viral entry in HCV infection and inhibiting the reverse transcriptase activity in HIV. The current study firstly reported the antioxidant, anti-inflammatory, and antiviral activities of Cucurbita maxima fucose-rich polysaccharide against several viral infections.

Ethanol extract from Artemisia argyi leaves inhibits HSV-1 infection by destroying the viral envelope

Herpes simplex virus type 1 (HSV-1) is a widely disseminated virus that establishes latency in the brain and causes occasional but fatal herpes simplex encephalitis. Currently, acyclovir (ACV) is the main clinical drug used in the treatment of HSV-1 infection, and the failure of therapy in immunocompromised patients caused by ACV-resistant HSV-1 strains necessitates the requirement to develop novel anti-HSV-1 drugs. Artemisia argyi, a Traditional Chinese Medicine, has been historically used to treat inflammation, bacterial infection, and cancer. In this study, we demonstrated the antiviral effect and mechanism of ethanol extract of A. argyi leaves (hereafter referred to as ‘AEE’). We showed that AEE at 10 μg/ml exhibits potent antiviral effects on both normal and ACV-resistant HSV-1 strains. AEE also inhibited the infection of HSV-2, rotavirus, and influenza virus. Transmission electron microscopy revealed that AEE destroys the membrane integrity of HSV-1 viral particles, resulting in impaired viral attachment and penetration. Furthermore, mass spectrometry assay identified 12 major components of AEE, among which two new flavones, deoxysappanone B 7,3ʹ-dimethyl ether, and 3,7-dihydroxy-3′,4ʹ-dimethoxyflavone, exhibited the highest binding affinity to HSV-1 glycoprotein gB at the surface site critical for gB–gH–gL interaction and gB-mediated membrane fusion, suggesting their involvement in inactivating virions. Therefore, A. argyi is an important source of antiviral drugs, and the AEE may be a potential novel antiviral agent against HSV-1 infection.

Primary Herpetic Gingivostomatitis.

An 82-year-old female presented with painful oral lesions associated with fever and dysphagia since 3 days. Examination revealed multiple grouped tender white to yellow plaques and few discrete vesicles on an erythematous base over the tongue and palate [Figure 1] with tender cervical lymphadenopathy. Tzanck smear from the vesicles showed multinucleate epidermal giant cells [Figure 2]. A diagnosis of primary herpetic gingivostomatitis was made and the lesions resolved following treatment with oral valacyclovir 1 g twice daily for 7 days [Figure 3]. Herpetic gingivostomatitis is typically caused by Herpes simplex virus type-1 (HSV-1), but cases caused by HSV-2 have also been reported. HSV-1 spreads by direct contact, via droplets, or by sexual contact. Primary herpetic gingivostomatitis usually presents with yellow to white plaques and vesicles which rupture and coalesce to form ulcers over the buccal mucosa, tongue, palate, gingiva, and the lips. Severe cases can have associated fever, malaise, myalgia, and local lymphadenopathy. Following the primary episode, the virus being neurotropic may become latent in the peripheral nerves and ganglions, with subsequent reactivations causing recurrent infections in around one-thirds of patients. Recurrent herpes simplex infections can present as localized vesicles commonly on the vermillion border of the lips, perioral skin, or the hard palate.[1] The diagnosis of herpes gingivostomatitis is primarily clinical. The differential diagnoses include aphthous stomatitis, oral candidiasis, herpangina, Behcet disease, erythema multiforme, Steven–Johnson syndrome, hand, foot and mouth disease and immunobullous disorders.[2] Tzanck smear from vesicles demonstrating viral cytopathic changes can support the diagnosis; however, it cannot distinguish between HSV-1, HSV-2, or varicella zoster virus. Crusted or ulcerative lesions are diagnosed by more accurate tests like direct fluorescent antibody test, polymerase chain reaction, and viral cultures. Serological tests are useful but not generally indicated for acute infections; also many people exposed to HSV may demonstrate asymptomatic seroconversion.[12]Figure 1: Multiple grouped tender white to yellow plaques and discrete vesicles on an erythematous base over the tongue and palateFigure 2: Tzanck smear showing multinucleate epidermal giant cellFigure 3: Complete resolution post treatmentMost cases of herpes gingivostomatitis are self-resolving, requiring only supportive measures. Severe cases may require systemic antiviral therapy – oral Acyclovir 15 mg/kg five times a day; or valacyclovir, 1g twice daily; or famciclovir, 500 mg twice daily; for 5 to 7 days. In case of frequent recurrences, prophylactic suppressive acyclovir (400 mg twice daily), valacyclovir (500 mg orally once daily), or famciclovir (250 mg twice daily) may be considered.[23] Declaration of patient consent The authors certify that they have obtained all appropriate patient consent forms. In the form, the patient(s) has/have given his/her/their consent for his/her/their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed. Financial support and sponsorship Nil. Conflicts of interest There are no conflicts of interest.

Identification of potent and selective inhibitors of PKR via virtual screening and traditional design

Protein Kinase RNA-activated (PKR) inhibition is thought to be relevant for immunology due to the potential to reduce macrophage and dendritic cell responses to bacteria and its signaling downstream of TNFα. PKR is also associated with neuroscience indications such as Alzheimer’s disease due to its activation by the double stranded DNA (dsDNA) virus HSV1, a virus suggested to be important in the development of AD. Studies exploring the mechanistic role of PKR with existing tool molecules such as the tricyclic oxindole C16 are clouded by the poor selectivity profile of this ATP-competitive, Type I kinase inhibitor. Type II kinase leads such as the benzothiophene or pyrazolopyrimidine scaffolds from literature are equally poor in their selectivity profiles. As such, it became necessary to identify more potent and selective chemical matter to better understand PKR biology. A dual approach was taken. The first step of the strategy included virtual screening of the AbbVie compound collection. A combination of pharmacophore-based and GPU shape-based screening was pursued to identify selective chemical matter from promiscuous leads. The second step of the strategy followed traditional compound design. This step initiated from a literature lead with PKR cross reactivity. Combined, the two parallel efforts led to identification of more selective leads for investigation of PKR biology.

Macrophage Delivered HSV1716 Is Active against Triple Negative Breast Cancer

Oncolytic viruses (OV) promote anti-tumour responses through the initiation of immunogenic cancer cell death which activates the host’s systemic anti-tumour immunity. We have previously shown that intravenously administered HSV1716 is an effective treatment for mammary cancer. However, intravenous administration of a virus has the potential to result in neutralization and sequestration of the virus which may reduce efficacy. Here, we show that the oncolytic virus HSV1716 can be administered within a cellular carrier (macrophages). PyMT and 4T1 murine mammary cancer cell lines were implanted into immuno-competent murine models (orthotopic primary, early metastatic and brain metastasis models). HSV1716 or macrophages armed with HSV1716 (M-HSV1716) were administered intravenously, and tumour size was quantified using caliper measurement or bioluminescence imaging. Administration of M-HSV1716 led to tumour shrinkage and increased the survival of animals. Furthermore, these results were achieved with a 100-fold lower viral load, which has the potential for decreased toxicity. Our results demonstrate that M-HSV1716 is associated with activity against murine mammary cancers and provides an alternative platform for the systemic delivery of OV.

Anti herpes simplex virus-2 activity of the extracts of Sericostoma pauciflorum stocks ex wight and Lepidagathis trinervis Nees

To determine the anti-HSV 2 properties and mechanism of action of two important medicinal plants Stocks ex Wight (Boraginaceae) and Nees (Acanthaceae) To identify natural products with anti-HSV-2 activity, the aqueous and 50% ethanolic extracts were prepared from whole plants of and . Cytotoxicity assay using Vero cells as target revealed CC values of these extracts ranging from 301.8 to 436.8 µg ml Extracts from both the plants showed dose dependent inhibition of HSV-2 infection using Plaque Reduction Assay. Among the tested extracts, 50% ethanolic extract of showed lowest IC value of 6.86 µg ml. To study the possible mode of antiviral action, all the extracts showed 50 to 97% reduction in the formation of plaques in the attachment assay. Extracts prepared from also inhibited the HSV-2 penetration in a time and dose dependent manner. In post-attachment assay, both the extracts from as well as 50% ethanolic extract from showed significant inhibition in HSV-2 infection.These studies document anti-HSV-2 activity of the two commonly used medicinal plants that will have implications in the development of an alternate remedy for HSV-2 infection. Significant and impact of the study: This work provides information on the anti HSV-2 properties of Indian medicinal plants that showed their significant inhibition of HSV-2 virus in system.

In vitro Virucidal Activity of Silver Nanoparticles against H1N1 Influenza A Virus and Herpes Simplex Virus-1

Silver nanoparticles (AgNPs) have provided a broad spectrum of antiviral activity against some types of enveloped and non-enveloped viruses. Silver nanoparticles are currently the most widely commercialized nanomaterials. In recent times, viral infections are emerging as one of the most common diseases with high mortality in the human population. The present study investigates the antiviral efficacy of the silver nanoparticles synthesized by the green method against HSV-1 and H1N1 influenza viruses. Characterization studies revealed that the silver nanoparticles showed an average particle size of 47 nm with a surface plasmon resonance peak at 426 nm. Silver nanoparticles exhibited 50% cytotoxicity in Vero cells at 197 µg/mL concentration, and 50% Inhibitory concentration (IC50) against HSV-1 was observed at 19.6 μg/mL. Silver nanoparticles demonstrated > 1 log reduction in H1N1 influenza A virus at 17 µg/mL. These findings indicate that silver nanoparticles possess excellent antiviral activity, which can be suitably used in various formulations to eradicate the spread of viral infections.

Chitosan-poly(ethylene oxide) nanofibrous mat as a vaginal platform for tenofovir disoproxyl fumarate – The effect of vaginal pH on drug carrier performance

In the present work, a solution blow spun nanofibrous mat comprised of chitosan (CS) and poly(ethylene oxide) (PEO) was obtained as vaginal platform for tenofovir disoproxil fumarate (TDF) to prevent sexually transmitted infections. Apart from physicochemical and mechanical analysis, the specific steps involved studies on nanofibrous mat mucoadhesive and swelling characteristics upon pH fluctuations over the physiological range. Physicochemical analysis showed uniform drug distribution within the CS/PEO mat volume and pointed toward physical interactions between the drug and polymers. TDF-loaded CS/PEO nanofibrous mat was shown potentially safe when evaluated by the MTT metabolic activity and JC-1 assays in human vaginal epithelial cells VK2-E6/E7. In vitro antiviral studies indicated inhibition efficacy of TDF-CS/PEO nanofibrous mat toward HSV-2 virus and proved the SBS process does not change the microbicidal activity of drug molecule. Fluctuations in the physiological vaginal pH range of 3.8 to 5.0 substantially affected mucoadhesive and swelling behavior of chitosan which in turn impacted drug dissolution rate from polymer carrier. The rate of permeation and accumulation of TDF in vaginal tissue differed in response to vaginal pH. Faster drug permeation assessed at pH 5.0 suggests that an increase in vaginal pH could improve TDF bioavailability at earlier time points.

Preliminary seroprevalence study of neurotropic virus antibodies in Nodding syndrome.

Nodding syndrome (NS) is a mostly East African pediatric epileptiform encephalopathy of unknown etiology that shares some clinical features with measles-associated subacute sclerosing panencephalitis (SSPE) and progressive rubella panencephalitis. Two independent studies in northern Uganda identified an association between NS and prior measles infection, while an earlier study in South Sudan found an inverse association. We report preliminary serologic analyses of antibodies to measles (MV), rubella (RV), HSV-1, and CMV viruses in northern Ugandan children with NS and Household (HC) and Community (CC) Controls. Only MV-positive titers were significantly different (3-fold and > 2-fold) in NS relative to HC and HC + CC, respectively. While these results are consistent with greater prior measles infection in Ugandan persons with NS, further studies are needed to determine whether Measles virus (MV) plays any role in the etiology and pathogenesis of NS. Resolving this issue will be invaluable for the thousands of children at risk for this devastating yet often neglected condition.

Vaccinium bracteatum Thunb Extract Inhibits HSV-1 Infection by Regulating ER Stress and Apoptosis.

Herpes simplex Type 1 (HSV-1) is a neurotropic virus that infects the peripheral and central nervous system. Usually, after primary infection in epithelial cells, HSV-1 migrates retrograde to the peripheral nervous system (PNS), where it establishes a latent infection. HSV-1 can remain latent in the nervous system, and its reactivation in the brain can rarely cause acute HSV-1 encephalitis, often a life-threatening condition, or asymptomatic reactivations that could lead to neuronal damage and ultimately neurodegenerative disorders. Acyclovir and related nucleoside analogs have been used as therapeutic agents for HSV-1 infection, but resistance to the drug can arise, and the protective effect of HSV-1 on brain cells is limited. Therefore, there is an urgent need for research into safe and effective new antiviral agents that can protect brain cells from the damage that is caused by HSV-1 infection. Vaccinium bracteatum Thunb. (VBT) is widely distributed in Korea and China, and has pharmacological actions such as anti-inflammatory, antioxidant, and antidiabetic activity. Studies on the antiviral effect of VBT on HSV-1 infection have not been reported so far. Therefore, we sought to determine the HSV-1 antiviral effect and molecular mechanism of VBT at the cellular level. We confirmed that VBT repressed the VP16 and IE genes in both Vero and SK-N-SH cells. We also found that the generation of HSV-1 virions was inhibited by VBT treatment. VBT inhibited the activities of the HSV-1-induced endoplasmic reticulum (ER) stressors PERK, ATF4, and CHOP. We confirmed that VBT inhibited the activity of apoptosis factors by regulating the expression of death receptor (DR) after HSV-1 infection. As HSV-1 is closely associated with brain diseases, the study of the antiviral drug effects and mechanism of VBT is meaningful. Further studies using animal models of infection will also be performed to determine the potential of VBT as an antiviral agent.

Inhibitory mechanism of ternary fusion protein MSIK on HPV virus.

We investigated whether the ternary fusion protein (MBP-SERPINA3-IFN-κ, MSIK) can inhibit the proliferation of human papillomavirus (HPV) and the possible mechanism of this inhibitory effect. First, the purchased MSIK protein was prepared into MSIK protein solutions of different concentrations, and then epithelial cells and 300 mice were involved in the research. The HSV-1 virus was used as the infection pathogen to explore the mechanism of MSIK reinhibition of HPV virus. The virus content in each sample was detected by PCR technology. Epithelial cells treated with different concentrations of MSIK had inhibitory effects on the invasion and replication of HSV-1 virus. MSIK protein had a positive effect on wound healing in mice at the same time, and it had an inhibitory effect on the invasion of HSV-1 virus, and the higher concentration of MSIK, the better the effect. MSIK can quickly help wound healing and inhibit the replication of HSV-1 virus. It can also effectively prevent the influx of HPV virus and replication has a positive effect on the prevention and treatment of HPV virus.

Transcription-independent regulation of STING activation and innate immune responses by IRF8 in monocytes

Sensing of cytosolic DNA of microbial or cellular/mitochondrial origin by cGAS initiates innate immune responses via the adaptor protein STING. It remains unresolved how the activity of STING is balanced between a productive innate immune response and induction of autoimmunity. Here we show that interferon regulatory factor 8 (IRF8) is essential for efficient activation of STING-mediated innate immune responses in monocytes. This function of IRF8 is independent of its transcriptional role in monocyte differentiation. In uninfected cells, IRF8 remains inactive via sequestration of its IRF-associated domain by its N- and C-terminal tails, which reduces its association with STING. Upon triggering the DNA sensing pathway, IRF8 is phosphorylated at Serine 151 to allow its association with STING via the IRF-associated domain. This is essential for STING polymerization and TBK1-mediated STING and IRF3 phosphorylation. Consistently, IRF8-deficiency impairs host defense against the DNA virus HSV-1, and blocks DNA damage-induced cellular senescence. Bone marrow-derived mononuclear cells which have an autoimmune phenotype due to deficiency of Trex1, respond to IRF-8 deletion with reduced pro-inflammatory cytokine production. Peripheral blood mononuclear cells from systemic lupus erythematosus patients are characterized by elevated phosphorylation of IRF8 at the same Serine residue we find to be important in STING activation, and in these cells STING is hyper-active. Taken together, the transcription-independent function of IRF8 we describe here appears to mediate STING activation and represents an important regulatory step in the cGAS/STING innate immune pathway in monocytes.

Phytochemical Discrimination, Biological Activity and Molecular Docking of Water-Soluble Inhibitors from Saussurea costus Herb against Main Protease of SARS-CoV-2.

Siddha medicine is one of the oldest medical systems in the world and is believed to have originated more than 10,000 years ago and is prevalent across ancient Tamil land. It is undeniable that inhibitor preferences rise with increasing solubility in water due to the considerations pertaining to the bioavailability and the ease of which unabsorbed residues can be disposed of. In this study, we showed the phytochemical discrimination of Saussurea costus extracted with water at room temperature as a green extraction procedure. A total of 48 compounds were identified using gas chromatography-mass spectrometry (GC-MS). The fatty acids had a high phytochemical abundance at 73.8%, followed by tannins at 8.2%, carbohydrates at 6.9%, terpenoids at 4.3%, carboxylic acids at 2.5%, hydrocarbons at 2.4%, phenolic compounds at 0.2%, and sterols at 1.5%. Of these compounds, 22 were docked on the active side and on the catalytic dyad of His41 and Cys145 of the main protease of SARS-CoV-2 (Mpro). Eight active inhibitors were carbohydrates, five were fatty acids, three were terpenoids, two were carboxylic acids, one was a tannin, one was a phenolic compound, and one was a sterol. The best inhibitors were 4,8,13-Cyclotetradecatriene-1,3-diol, 1,5,9-trimethyl-12-(1-methylethyl), Andrographolide, and delta.4-Androstene-3.beta.,17.beta.-diol, with a binding affinity that ranged from −6.1 kcal/mol to −6.5 kcal/mol. The inhibitory effect of Saussurea costus of SARS-CoV-2 entry into the cell was studied using a pseudovirus with Spike proteins from the D614G variant and the VOC variants Gamma and Delta. Based on the viral cycle of SARS-CoV-2, our results suggest that the Saussurea costus aqueous extract has no virucidal effect and inhibits the virus in the events after cell entry. Furthermore, the biological activity of the aqueous extract was investigated against HSV-1 virus and two bacterial strains, namely Staphylococcus aureus ATCC BAA 1026 and Escherichia coli ATCC 9637. According to this study, an enormous number of water-soluble inhibitors were identified from Saussurea costus against the Mpro, and this is unprecedented as far as we know.

Chiral sulfonamides with various N-heterocyclic and aromatic units – Synthesis and antiviral activity evaluation

Chiral sulfonamides with aromatic fragments are important chemical building blocks found widely in many natural products, catalysts, and molecules of biological importance. In this report, we describe the efficient synthesis of a series of chiral sulfonamides which, in addition to the aromatic part (phenyl, biphenyl, and dansyl units), possess N-heterocyclic systems. The described compounds were obtained by nucleophilic substitution of chiral N-heterocyclic amines and commercially available aromatic sulfonyl chlorides under mild conditions. All derivatives were examined in antiviral assay against AdV5, HSV-1, HPIV-3, HCMV, and EMCV viruses.

A Baseline Cellular Antiviral State Is Maintained by cGAS and Its Most Frequent Naturally Occurring Variant rs610913.

Upon recognition of aberrantly located DNA, the innate immune sensor cyclic GMP-AMP synthase (cGAS) activates stimulator of IFN genes (STING)/IFN regulatory factor (IRF)3-driven antiviral responses. In this study, we characterized the ability of a specific variant of the human cGAS-encoding gene MB21D1, rs610913, to alter cGAS-mediated DNA sensing and viral infection. rs610913 is a frequent G>T polymorphism resulting in a P261H exchange in the cGAS protein. Data from the International Collaboration for the Genomics of HIV suggested that rs610913 nominally associates with HIV-1 acquisition in vivo. Molecular modeling of cGAS(P261H) hinted toward the possibility for an additional binding site for a potential cellular cofactor in cGAS dimers. However, cGAS(wild-type [WT]) or cGAS(P261H)-reconstituted THP-1 cGAS knockout cells shared steady-state expression of IFN-stimulated genes, as opposed to cells expressing the enzymatically inactive cGAS(G212A/S213A). Accordingly, cGAS(WT) and cGAS(P261H) cells were less susceptible to lentiviral transduction and infection with HIV-1, HSV-1, and Chikungunya virus as compared with cGAS knockout or cGAS(G212A/S213A) cells. Upon DNA challenge, innate immune activation appeared to be mildly reduced upon expression of cGAS(P261H) compared with cGAS(WT). Finally, DNA challenge of PBMCs from donors homozygously expressing rs610913 provoked a trend toward a slightly reduced type I IFN response as compared with PBMCs from GG donors. Taken together, the steady-state activity of cGAS maintains a baseline antiviral state rendering cells more refractory to IFN-stimulated gene-sensitive viral infections. rs610913 failed to grossly differ phenotypically from the WT gene, suggesting that cGAS(P261H) and WT cGAS share a similar ability to sense viral infections in vivo.

Viral Etiology in Acute Encephalitis Syndrome in North Indian Children

Background and Objective: Acute Encephalitis Syndrome (AES) leads to significant mortality and morbidity in children. This study was undertaken to identify the viral causes of AES and its correlation with clinicodemographic profile. Methods: This cross-sectional study included 100 children aged 1 to 12 years presenting with fever of <14 days duration and a change in mental status (including symptoms such as confusion, disorientation, coma, or inability to talk) and/or new onset of seizures (excluding simple febrile seizures) [1]. Cerebrospinal Fluid (CSF) was collected aseptically by lumbar puncture for biochemical and cytological analysis, and for bacteriological culture. Multiplex PCR for the viruses HSV1, HSV2, Varicella zoster virus, Enterovirus, Mumps and Parecho virus was carried out. Serology for Japanese Encephalitis (JE) was done in blood and CSF by ELISA method. Dengue NS1 antigen and IgM ELISA was carried out in both blood and CSF samples. Serum IgM antibody for scrub typhus was done by ELISA method. Results: Out of 100 children viral cause was detected in 21 patients (Japanese encephalitis -16, Dengue – 4, Varicella zoster virus, Enterovirus and Mumps virus – 1 each). Serum IgM ELISA for scrub typhus was positive in 6 patients. Multiple viral etiology was seen in 2 patients. Mortality was significantly more in females (p=0.021), GCS <8 (p <0.001). Mortality was significantly more with higher CSF neutrophils and higher proteins (p<0.05). Interpretation and Conclusion: This study concludes that viral causes specially JE contributed in AES in North India. Hence viral pathogens including JE needs to be considered as one of major causative agents while treating patients of AES and further emphasis has to be given for JE vaccination in this region.

Abstract LB020: PTEN-L expressing HSV induces glioma stem cell differentiation and sensitizes glioblastoma to radiation in mice

Abstract GBM, a WHO classified Grade IV glioma, is one of the most lethal and heterogeous primary brain tumors with inevitable recurrence, limiting the median survival time less than 21 months. Current standard-of-care treatment including surgical resection followed by chemo- and radio-therapy remains palliative because of therapy resistance, majorly conferred by GBM stem cells (GSCs), leading to tumor recurrence. With no current effective treatments, novel approaches to overcome GSCs-mediated resistance to chemotherapy and irradiation are urgently required in order to achieve long-term success in GBM therapy. Oncolytic viral (OV) therapy represents a novel and promising biological therapy for solid tumor that preferentially targets tumor cells for lytic destruction, sparing the healthy cells and in the process activating host anti-tumor immune response. Among all OVs, oncolytic Herpes Simplex Virus (oHSV) is substantially ahead in the clinic, with an oHSV T-VEC approved by the FDA for metastatic melanoma treatment. Recently, G47∆, another HSV1 virus, has been granted conditional approval for the treatment of GBM in Japan. Further, several other oHSVs including G207 and HSV1716 are currently being tested for safety and efficacy against GBM. This has fueled great expectations towards OVs as a promising alternative to conventional therapies. Our group has previously shown that PTENα expression by an oHSV (HSV-P10) resulted in further improved long-term survivors in intracranial tumor-bearing mice compared to HSVQ treatment. Here we aim to dissect the molecular mechanisms associated with improved therapeutic efficacy of HSV-P10 against GBM, and if HSV-P10 can overcome GBM cell radioresistance. The RNA sequencing and GSEA analyses of primary human GBM cells infected with control HSVQ or HSV-P10 reveals that while HSVQ virus infection leads to an increase in genes regulating IL6-STAT3 pathway, pivotal in maintaining stemness properties, HSV-P10 infection causes a reduction in the genes regulating this pathway. As a consequence, HSV-P10 reduces CD133+/CD44+ stem cell fraction, induces DNA damage and sensitizes the GBM cells to irradiation. Our findings reveal a novel mechanism induced by HSV-P10 in combination with irradiation whereby HSV-P10 modulates IL6-STAT3 signaling downregulating Sox2, a core transcription factor in the maintenance of GSCs, with a simultaneous decrease in Nestin expression and enhanced GFAP expression promoting GSC differentiation. HSV-P10 infection in combination with irradiation reduces GSC tumor sphere formation in vitro and sensitizes GBMs to radiotherapy in an intracranial mouse xenograft model. Collectively, our findings provide a potential avenue to overcome GSC-mediated therapy resistance to improve the therapeutic efficacy for GBM patients. Citation Format: Upasana Sahu, Matthew P. Mullarkey, Bangxing Hong, Balveen Kaur. PTEN-L expressing HSV induces glioma stem cell differentiation and sensitizes glioblastoma to radiation in mice [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 LB020.

Abstract 3570: Prospects for personalized cancer vaccines: The oncolytic and immunotherapeutic HSV-1(VC2) virus expressing an ovalbumin (OVA) antigen elicits robust anti-OVA immune responses

Abstract Cancers result from unique cellular transformation events resulting in the expression of cancer antigens that vary widely among different types of cancers. Ideally, a personalized cancer treatment approach needs to target specific cancer antigen. Efforts to induce specific immune responses against tumor associated antigens (TAAs) include the expression of TAAs through viral vectors capable of expressing multiple antigens. We have previously shown that the HSV-1(VC2) vaccine vector can induce potent antitumor immune responses utilizing the B16F10-derived syngeneic melanoma mouse model. To investigate whether heterologous antigens expressed by this vaccine vector could elicit potent immune responses, we generated the recombinant virus VC2-OVA that expresses the ovalbumin antigen (OVA) fused in-frame at the amino terminus of the viral VP26 protein. Western blot analysis of infected cells in cell culture showed efficient expression of the OVA-VP26 protein. The anti-tumor efficacy of the VC2-OVA virus was tested in the B16cOVA and E.G7-OVA mouse models that constitutively express the OVA antigen to test whether this vaccine can either prevent the formation of new tumors (prophylactic modality) or reduce tumor metastasis (therapeutic modality). In addition, we evaluated the differences between intradermal, subcutaneous, and intramuscular routes of vaccination with VC2-OVA. Treatment with VC2-OVA resulted in significant reduction of tumor formation and metastasis in both the prophylactic and therapeutic modalities. Importantly, the anti-tumor efficacy was dependent on the specific mouse tumor model. Further, we found that subcutaneous and intradermal was superior to intramuscular vaccination. The VC2 virus is available as a bacterial artificial chromosome allowing for the rapid isolation of recombinant VC2-derived virus expressing multiple tumor antigens. Therefore, these results demonstrate that the VC2 vaccine virus can be used to produce personalized cancer vaccines targeting specific TAAs. Citation Format: Ifeanyi K. Uche, Brent Stanfield, Jared Rudd, Konstantin Kousoulas, Paul J. Rider. Prospects for personalized cancer vaccines: The oncolytic and immunotherapeutic HSV-1(VC2) virus expressing an ovalbumin (OVA) antigen elicits robust anti-OVA immune responses [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 3570.

Inconsistencies in Modeling the Efficacy of the Oncolytic Virus HSV1716 Reveal Potential Predictive Biomarkers for Tolerability.

Treatment with HSV1716 via intralesional administration has proven successful for melanoma patients with the hope that oncolytic virotherapy would become another weapon in the systemic anticancer therapy (SACT) arsenal. In addition to challenges surrounding the systemic delivery of oncolytic viruses (OVs), problems associated with its in vivo modeling have resulted in low predictive power, contributing to the observed disappointing clinical efficacy. As OV’s efficacy is elicited through interaction with the immune system, syngeneic orthotopic mouse models offer the opportunity to study these with high reproducibility and at a lower cost; however, inbred animals display specific immune characteristics which may confound results. The systemic delivery of HSV1716 was, therefore, assessed in multiple murine models of breast cancer. Tolerability to the virus was strain-dependent with C57/Bl6, the most tolerant and Balb/c experiencing lethal side effects, when delivered intravenously. Maximum tolerated doses were not enough to demonstrate efficacy against tumor growth rates or survival of Balb/c and FVB mouse models; therefore; the most susceptible strain (Balb/c mice) was treated with immunomodulators prior to virus administration in an attempt to reduce side effects. These studies demonstrate the number of variables to consider when modeling the efficacy of OVs and the complexities involved in their interpretation for translational purposes. By reporting these observations, we have potentially revealed a role for T-cell helper polarization in viral tolerability. Importantly, these findings were translated to human studies, whereby a Th1 cytokine profile was expressed in pleural effusions of patients that responded to HSV1716 treatment for malignant pleural mesothelioma with minimal side effects, warranting further investigation as a biomarker for predictive response.