Exploring JC Polyomavirus Sequences and Human Gene Expression in Brain Tissue of Patients With Progressive Multifocal Leukoencephalopathy.

Progressive multifocal leukoencephalopathy (PML) is a rare neurological condition associated with reactivation of dormant JC polyomavirus (JCPyV). In this study, we characterized gene expression and JCPyV rearrangements in PML brain tissue. Infection of white matter astrocytes and oligodendrocytes as well as occasional brain cortex neurons was shown. PML brain harbored exclusively rearranged JCPyV variants. Viral transcripts covered the whole genome on both strands. Strong differential expression of human genes associated with neuroinflammation, blood-brain barrier permeability, and neurodegenerative diseases was shown. Pathway analysis revealed wide immune activation in PML brain. The study provides novel insights into the pathogenesis of PML.

Nonclinical and clinical characterization of MAU868, a novel human-derived monoclonal neutralizing antibody targeting BK polyomavirus VP1.

Reactivation of BK polyomavirus (BKPyV) can cause significant kidney and bladder disease in immunocompromised patients. There are currently no effective, BKPyV-specific therapies. MAU868 is a novel, human immunoglobulin (Ig) G1 monoclonal antibody that binds the major capsid protein, VP1, of BKPyV with picomolar affinity, neutralizes infection by the 4 major BKPyV genotypes (EC50 ranging from 0.009-0.093 µg/mL; EC90 ranging from 0.102-4.160 µg/mL), and has comparable activity against variants with highly prevalent VP1 polymorphisms. No resistance-associated variants were identified in long-term selection studies, indicating a high in vitro barrier-to-resistance. The high-resolution crystal structure of MAU868 in complex with VP1 pentamer identified 3 key contact residues in VP1 (Y169, R170, and K172). A first-in-human study was conducted to assess the safety, tolerability, and pharmacokinetics of MAU868 following intravenous and subcutaneous administration to healthy adults in a randomized, placebo-controlled, double-blinded, single ascending dose design. MAU868 was safe and well-tolerated. All adverse events were grade 1 and resolved. The pharmacokinetics of MAU868 was typical of a human IgG, with dose-proportional systemic exposure and an elimination half-life ranging between 23 and 30 days. These results demonstrate the potential of MAU868 as a first-in-class therapeutic agent for the treatment or prevention of BKPyV disease.

Detection of JC and BK polyomaviruses in patients with colorectal cancer (CRC) by PCR.

BACKGROUND: Overall, 20-30% of all cancers are estimated to be linked to infectious agents. Polyomaviruses are oncogenic cause in rodent models, readily transform their cells, and cause chromosomal instability in animal and human cells in-vitro. Some reports have indicated the presence of JCPyV and BKPyV in some human tumors. The JCPyV and BKPyV genome encodes some transforming proteins such as LT-Ag. Thus, these viruses could cause or promote some neoplasia, such as lymphomas, pancreatic, prostate, and colorectal cancers. Colorectal cancer (CRC) is the third most common cancer in the world. Risk factors for developing CRC are associated with personal features or habits, such as age, lifestyle, and gut microbiota. MATERIALS AND METHODS: In this study, we examined the prevalence of JCPyV and BKPyV in the 23 fecal samples of CRC patients and 24 healthy samples (control group). Virus DNA was extracted by a Favorgen DNA extraction kit. The large T antigen of JCPyV and VP1 of BKPyV were investigated by optimized multiplex PCR. RESULTS: One of the samples was positive for the JCPyV (4.3%), while in the samples of healthy individuals, the JCPyV was negative. Also, positive results for BKPyV PCR were obtained for five cases (21.7%) in the samples of the CRC group and one case (4.1%) in healthy individuals. CONCLUSION: The result showed no direct correlation between tumorigenesis and polyomavirus infections in CRC development. However, the exact role of BKPyV and JCPyV is still controversial and needs further study with larger sample size.

Mapping of Human Polyomavirus in Renal Cell Carcinoma Tissues.

Worldwide, the incidence of renal cell carcinoma (RCC) is rising, accounting for approximately 2% of all cancer diagnoses and deaths. The etiology of RCC is still obscure. Here, we assessed the presence of HPyVs in paraffin-embedded tissue (FFPE) resected tissue from patients with RCC by using different molecular techniques. Fifty-five FFPE tissues from 11 RCC patients were included in this study. Consensus and HPyV-specific primers were used to screen for HPyVs. Both PCR approaches revealed that HPyV is frequently detected in the tissues of RCC kidney resections. A total of 78% (43/55) of the tissues tested were positive for at least one HPyV (i.e., MCPyV, HPyV6, HPyV7, BKPyV, JCPyV, or WUyV). Additionally, 25 tissues (45%) were positive for only one HPyV, 14 (25%) for two HPyVs, 3 (5%) for three HPyVs, and 1 one (1%) tissue specimen was positive for four HPyVs. Eleven (20%) RCC specimens were completely devoid of HPyV sequences. MCPyV was found in 24/55 RCC tissues, HPyV7 in 19, and HPyV6 in 8. The presence of MCPyV and HPyV6 was confirmed by specific FISH or RNA-ISH. In addition, we aimed to confirm HPyV gene expression by IHC. Our results strongly indicate that these HPyVs infect RCC and nontumor tissues, possibly indicating that kidney tissues serve as a reservoir for HPyV latency. Whether HPyVs possibly contribute to the etiopathogenesis of RCC remains to be elucidated.

JC Polyomavirus Modifies the Expression of Human microRNAs in Progressive Multifocal Leukoencephalopathy Brain.

Progressive multifocal leukoencephalopathy (PML) is a severe neurological condition caused by reactivation of JC polyomavirus (JCPyV) in immunosuppression. Asymptomatic JCPyV persists in peripheral tissues. Upon reactivation, neurotropic rearrangements may emerge, and the virus gains access to the brain. To assess the mechanisms of PML pathogenesis, brain tissue material from PML patients was collected for small RNA sequencing. Upregulation of 8 microRNAs (miRNAs) in PML brain was validated using quantitative microRNA polymerase chain reaction (PCR). Bioinformatics tools were utilized to identify major associations of the upregulated miRNAs: neuroinflammation and blood-brain barrier disruption. The results indicate involvement of human miRNA regulation in PML pathogenesis.

Detection of human neurotropic JCPyV DNA sequence in pediatric anaplastic xanthoastrocytoma.

Due to its peculiar histopathological findings, pleomorphic xanthoastrocytoma (PXA), a rare cerebral tumor of young adults with a slow growth and a good prognosis, resembles to the lytic phase of progressive multifocal leukoencephalopathy, a fatal neurodegenerative disease caused by JC polyomavirus (JCPyV). Therefore, the presence of JCPyV DNA was examined in an 11-year-old child with xanthoastrocytoma, WHO grade 3, by quantitative PCR (qPCR) and nested PCR (nPCR) using primers amplifying sequences encoding the N- and C-terminal region of large T antigen (LTAg), the non-coding control region (NCCR), and viral protein 1 (VP1) DNA. The expression of transcripts from LTAg and VP1 genes was also evaluated. In addition, viral microRNAs' (miRNAs) expression was investigated. Cellular p53 was also searched at both DNA and RNA level. qPCR revealed the presence of JCPyV DNA with a mean value of 6.0 × 104 gEq/mL. nPCR gave a positive result for the 5' region of the LTAg gene and the NCCR, whereas 3' end LTAg and VP1 DNA sequences were not amplifiable. Only LTAg transcripts of 5' end were found whereas VP1 gene transcript was undetectable. Although in most cases, either Mad-1 or Mad-4 NCCRs have been identified in association with JCPyV-positive human brain neoplasms, the archetype NCCR structure was observed in the patient's sample. Neither viral miRNA miR-J1-5p nor p53 DNA and RNA were detected. Although the expression of LTAg supports the possible role of JCPyV in PXA, further studies are warranted to better understand whether the genesis of xanthoastrocytoma could depend on the transformation capacity of LTAg by Rb sequestration.

Rapid-progressing progressive multifocal leukoencephalopathy in two patients newly diagnosed with HIV: case series and review of literature.

The JC Polyomavirus (JCPyV) is a virus of global distribution and is usually kept under control by the immune system. In patients with AIDS, a latent JCPyV infection can reactivate and develop into progressive multifocal leukoencephalopathy (PML). Around half of the patients with PML die within 2 years since the diagnosis, yet in rare cases, the disease advances significantly quicker and seems to be insusceptible to any medical actions. In our clinic, we observed two cases of such course in HIV-positive patients in the AIDS stage. On admission, both patients had mild neurological symptoms such as dizziness, vision disturbances, and muscle weakness. Both had extremely low CD4 lymphocyte count (7 cells/µL, 40 cells/µL) and high HIV-1 viral load (VL) (50,324 copies/ml, 78,334 copies/ml). PML was confirmed by PCR for JCPyV DNA in cerebrospinal fluid (CSF) coupled with clinical and radiological features. Despite receiving though antiretroviral (ARV) treatment paired with intra-venous (IV) steroids, the disease progressed rapidly with neurological manifestations exacerbating throughout the few weeks following the admission. Eventually, both patients developed respiratory failure and died within less than 3 months after the onset of the neurological symptoms. Even though such curse of the disease is not common, it should be a warning to all how deadly both PML and AIDS can be and remind doctors to offer testing even to asymptomatic patients.

Infection by human polyomaviruses JCPyV and BKPyV in blood donors of Argentina.

BACKGROUND AND OBJECTIVES: A spectrum of blood-borne infectious agents may be transmitted through transfusion of blood components from asymptomatic donors. Despite the persistence of polyomaviruses in blood cells, no studies have been conducted in Argentina to assess the risk of transfusion infection. MATERIALS AND METHODS: We investigated BKPyV and JCPyV in 720 blood donors, using polymerase chain reaction (PCR) for a region of T antigen common to both viruses. Positive T-antigen samples were subjected to two additional PCR assays targeting the VP1 region. Viral genotypes were characterized by phylogenetic analysis. RESULTS: Polyomaviruses were detected in 1.25% (9/720) of the blood samples selected; JCPyV was identified in 0.97% (7/720) and BKPyV in 0.28% (2/720) of them. Phylogenetic analysis showed that the JCPyV sequences clustered with 2A genotype and Ia of BKPyV. CONCLUSION: This study describes for the first time the prevalence of polyomavirus DNA in blood donors of Córdoba, Argentina. The polyomavirus DNAemia in healthy populations suggests that those viruses are present in blood components eligible for transfusion. Therefore, the epidemiological surveillance of polyomavirus in blood banks might be incorporated into haemovigilance programmes, to determine the infectious risk and implement newer interventions to ensure the safety of blood supplies, if required.

Prevalence of JC Polyomavirus in Patients with Neuroinvasive Disease of Unknown Etiology in Croatia.

Background and Objectives: John Cunningham polyomavirus (JCPyV) is a highly prevalent virus in the human population. The prevalence of JCPyV in patients with central nervous system disorders has not been examined extensively. The aim of this study was to analyze the prevalence of JCPyV DNA/antibodies in patients with neuroinvasive diseases (NID) of unknown etiology. Materials and Methods: The study included 132 patients with NID (febrile headache, meningitis, encephalitis) tested from January 2021 to December 2022. The control group consisted of 47 asymptomatic individuals. In patients with NID, serum and cerebrospinal fluid (CSF) samples were collected in the acute phase of the disease. CSF samples were tested for JCPyV DNA (PCR), while serum samples were tested for JCPyV IgG antibodies (ELISA). In controls, serum samples were tested for JCPyV IgG antibodies (ELISA). Results: JCPyV DNA was not detected in any of the CSF samples from patients with NID. JCPyV IgG antibodies were detected in 88.6% of patients and 74.5% of controls (p < 0.001). In the patients' group, a significant difference in the IgG prevalence was observed between males (94.6%) and females (81.0%). In addition, significant differences in the seropositivity between age groups were found. The lowest seroprevalence (28.6%) was in patients less than 20 years, followed by a sharp increase in the 20-29-year group (69.2%), after which the seroprevalence remained stable (90.0-94.1%) in patients up to 69 years. All patients older than 70 years were JCPyV IgG-seropositive. No significant difference in the seroprevalence was found in patients presenting with febrile headache (81.6%), meningitis (93.3%), or meningoencephalitis (91.3%). No difference in the seropositivity between genders was found in controls. Although the seropositivity steadily increased in older participants, these differences were not significant. Analyzing the JCPyV antibody levels in patients with NID, the median antibody titers differed significantly between groups, ranging from 248 AU/mL (younger age groups) to 400 AU/mL (older age groups). Conclusions: Higher seroprevalence in the patients' group highlights the need to further investigate the possible association of JCPyV and NID.

Traces of JC polyomavirus in papillary thyroid cancer: a comprehensive study in Iran.

BACKGROUND: JC polyomavirus (JCPyV) is known to induce solid tumors such as astrocytomas, glioblastomas, and neuroblastomas in experimental animals, and recent studies have shown that the virus may be correlated with carcinogenesis. This study aimed to evaluate the impact of JCPyV on the progression of papillary thyroid cancer (PTC). METHODS: A total of 1057 samples, including 645 paraffin-embedded PTC biopsy samples (PEBS) and 412 fresh biopsy samples (FBS), and 1057 adjacent non-cancerous samples were evaluated for the presence of JCPyV DNA and RNA. RESULTS: We observed that 10.8% (114/1057) samples, including 17.5% (72/412) FBS and 6.5% (42/645) PEBS were positive for the JCPyV DNA. Among the JCPyV-positive samples, the mean JCPyV copy number was lower in patients with PEBS (0.3 × 10-4 ± 0.1 × 10-4 copies/cell) compared to FBS (1.8 × 10-1 ± 0.4 × 10-1 copies/cell) and non-PTC normal samples (0.2 × 10-5 ± 0.01 × 10-5 copies/cell), with a statistically significant difference (P < 0.001). The LT-Ag RNA expression was lower in PEBS than in FBS, while no VP1 gene transcript expression was found. CONCLUSIONS: Although our results confirmed the presence of JCPyV in some Iranian patients with PTC, more research is needed to verify these results.

Antivirals against human polyomaviruses: Leaving no stone unturned.

Human polyomaviruses (HPyVs) encompass more than 10 species infecting 30%-90% of the human population without significant illness. Proven HPyV diseases with documented histopathology affect primarily immunocompromised hosts with manifestations in brain, skin and renourinary tract such as polyomavirus-associated nephropathy (PyVAN), polyomavirus-associated haemorrhagic cystitis (PyVHC), polyomavirus-associated urothelial cancer (PyVUC), progressive multifocal leukoencephalopathy (PML), Merkel cell carcinoma (MCC), Trichodysplasia spinulosa (TS) and pruritic hyperproliferative keratinopathy. Although virus-specific immune control is the eventual goal of therapy and lasting cure, antiviral treatments are urgently needed in order to reduce or prevent HPyV diseases and thereby bridging the time needed to establish virus-specific immunity. However, the small dsDNA genome of only 5 kb of the non-enveloped HPyVs only encodes 5-7 viral proteins. Thus, HPyV replication relies heavily on host cell factors, thereby limiting both, number and type of specific virus-encoded antiviral targets. Lack of cost-effective high-throughput screening systems and relevant small animal models complicates the preclinical development. Current clinical studies are limited by small case numbers, poorly efficacious compounds and absence of proper randomized trial design. Here, we review preclinical and clinical studies that evaluated small molecules with presumed antiviral activity against HPyVs and provide an outlook regarding potential new antiviral strategies.

JC Polyomavirus Infection Reveals Delayed Progression of the Infectious Cycle in Normal Human Astrocytes.

JC polyomavirus (JCPyV) infects 50 to 80% of the population and is the causative agent of a fatal demyelinating disease of the central nervous system (CNS). JCPyV presents initially as a persistent infection in the kidneys of healthy people, but during immunosuppression, the virus can reactivate and cause progressive multifocal leukoencephalopathy (PML). Within the CNS, JCPyV predominately targets two cell types, oligodendrocytes and astrocytes. Until recently, the role of astrocytes has been masked by the pathology in the myelin-producing oligodendrocytes, which are lytically destroyed by the virus. To better understand how astrocytes are impacted during JCPyV infection, the temporal regulation and infectious cycle of JCPyV were analyzed in primary normal human astrocytes (NHAs). Previous research to define the molecular mechanisms underlying JCPyV infection has mostly relied on the use of cell culture models, such as SVG-A cells (SVGAs), an immortalized, mixed population of glial cells transformed with simian virus 40 (SV40) T antigen. However, SVGAs present several limitations due to their immortalized characteristics, and NHAs represent an innovative approach to study JCPyV infection in vitro Using infectivity assays, quantitative PCR, and immunofluorescence assay approaches, we have further characterized JCPyV infectivity in NHAs. The JCPyV infectious cycle is significantly delayed in NHAs, and the expression of SV40 T antigen alters the cellular environment, which impacts viral infection in immortalized cells. This research establishes a foundation for the use of primary NHAs in future studies and will help unravel the role of astrocytes in PML pathogenesis.IMPORTANCE Animal models are crucial in advancing biomedical research and defining the pathogenesis of human disease. Unfortunately, not all diseases can be easily modeled in a nonhuman host or such models are cost prohibitive to generate, including models for the human-specific virus JC polyomavirus (JCPyV). JCPyV infects most of the population but can cause a rare, fatal disease, progressive multifocal leukoencephalopathy (PML). There have been considerable advancements in understanding the molecular mechanisms of JCPyV infection, but this has mostly been limited to immortalized cell culture models. In contrast, PML pathogenesis research has been greatly hindered because of the lack of an animal model. We have further characterized JCPyV infection in primary human astrocytes to better define the infectious process in a primary cell type. Albeit a cell culture model, primary astrocytes may better recapitulate human disease, are easier to maintain than other primary cells, and are less expensive than using an animal model.

BK Polyomavirus Hijacks Extracellular Vesicles for En Bloc Transmission.

Most people are asymptomatic carriers of the BK polyomavirus (BKPyV), but the mechanisms of persistence and immune evasion remain poorly understood. Furthermore, BKPyV is responsible for nephropathies in kidney transplant recipients. Unfortunately, the sole therapeutic option is to modulate immunosuppression, which increases the risk of transplant rejection. Using iodixanol density gradients, we observed that Vero and renal proximal tubular epithelial infected cells release two populations of infectious particles, one of which cosediments with extracellular vesicles (EVs). Electron microscopy confirmed that a single vesicle could traffic tens of viral particles. In contrast to naked virions, the EV-associated particles (eBKPyVs) were not able to agglutinate red blood cells and did not use cell surface sialylated glycans as an attachment factor, demonstrating that different entry pathways were involved for each type of infectious particle. However, we also observed that naked BKPyV and eBKPyV were equally sensitive to neutralization by the serum of a seropositive patient or commercially available polyvalent immunoglobulin preparations, which occurred at a postattachment step, after endocytosis. In conclusion, our work shows a new mechanism that likely plays a critical role during the primary infection and in the persistence, but also the reactivation, of BKPyV.IMPORTANCE Reactivation of BKPyV is responsible for nephropathies in kidney transplant recipients, which frequently lead to graft loss. The mechanisms of persistence and immune evasion used by this virus remain poorly understood, and a therapeutic option for transplant patients is still lacking. Here, we show that BKPyV can be released into EVs, enabling viral particles to infect cells using an alternative entry pathway. This provides a new view of BKPyV pathogenesis. Even though we did not find any decreased sensitivity to neutralizing antibodies when comparing EV-associated particles and naked virions, our study also raises important questions about developing prevention strategies based on the induction or administration of neutralizing antibodies. Deciphering this new release pathway could enable the identification of therapeutic targets to prevent BKPyV nephropathies. It could also lead to a better understanding of the pathophysiology of other polyomaviruses that are associated with human diseases.

The differential presence of human polyomaviruses, JCPyV and BKPyV, in prostate cancer and benign prostate hypertrophy tissues.

BACKGROUND: Studies have shown that human polyomavirus infection may be associated with various human cancers. We investigated the potential relationship between the prevalence of JCPyVor BKPyV and prostate cancer (PC) in patients from Taiwan. METHODS: Patients with PC and benign prostate hypertrophy (BPH; 76 and 30 patients, respectively) were recruited for this study. Paraffin-embedded tissues and clinical information of the patients were obtained. The tissue sections were used for viral DNA detection and immunohistochemistry analysis was performed for examining viral large T (LT) and VP1 proteins. Regression analysis was used to evaluate the relationship between the clinical characteristics of the patients and the risk of JCPyV/BKPyV infection. RESULTS: The prevalence of JCPyV/BKPyV DNA was different in PC and BPH tissues (27/76 [35.52%] and 2/30 [6.7%], respectively, p = 0.003)]. The LT and VP1 proteins were detected in 27 (35.52%) and 29 PC (38.2%) specimens, respectively, but neither protein was detected in BPH samples (p < 0.001). PC cells were more susceptible to JCPyV infection than BPH tissues [odds ratio (OR) 7.71, 95% CI: 1.71-34.09, p = 0.003). Patients with PC showing high levels of prostate-specific antigen and high Gleason scores were associated with a high risk of viral infection (ORs 1.1, 95% CI 1.000-1.003; p = 0.045 and ORs 6.18, 95% CI 1.26-30.33, p = 0.025, respectively). The expression of LT protein associated with the risk of PC increased 2923.39-fold (95% CI 51.19-166,963.62, p < 0.001). CONCLUSIONS: The findings indicate that JCPyV infection in PC cells may be associated with prostate cancer progression and prognosis.

High anti-JCPyV serum titers coincide with high CSF cell counts in RRMS patients.

BACKGROUND: Progressive multifocal leukoencephalopathy (PML) can in rare cases occur in natalizumab-treated patients with high serum anti-JCPyV antibodies, hypothetically due to excessive blockade of immune cell migration. OBJECTIVE: Immune cell recruitment to the central nervous system (CNS) was assessed in relapsing-remitting multiple sclerosis (RRMS) patients stratified by low versus high anti-JCPyV antibody titers as indicator for PML risk. METHODS: Cerebrospinal fluid (CSF) cell counts of 145 RRMS patients were quantified by flow cytometry. Generalized linear models were employed to assess influence of age, sex, disease duration, Expanded Disability Status Scale (EDSS), clinical/radiological activity, current steroid or natalizumab treatment, as well as anti-JCPyV serology on CSF cell subset counts. RESULTS: While clinical/radiological activity was associated with increased CD4, natural killer (NK), B and plasma cell counts, natalizumab therapy reduced all subpopulations except monocytes. With and without natalizumab therapy, patients with high anti-JCPyV serum titers presented with increased CSF T-cell counts compared to patients with low anti-JCPyV serum titers. In contrast, PML patients assessed before (n = 2) or at diagnosis (n = 5) presented with comparably low CD8 and B-cell counts, which increased after plasma exchange (n = 4). CONCLUSION: High anti-JCPyV indices, which could be indicative of increased viral activity, are associated with elevated immune cell recruitment to the CNS. Its excessive impairment in conjunction with viral activity could predispose for PML development.

Specific antibodies reacting to JC polyomavirus capsid protein mimotopes in sera from multiple sclerosis and other neurological diseases-affected patients.

Published data support the hypothesis that viruses could be trigger agents of multiple sclerosis onset. This link is based on evidence of early exposure to viral agents in patients affected by this neurologic disease. JC (JC polyomavirus [JCPyV]), BK (BKPyV), and simian virus 40 (SV40) neurotropic polyomavirus footprints have been detected in brain tissue specimens and samples from patients affected by different neurological diseases. In this investigation, serum samples from patients affected by multiple sclerosis and other inflammatory and noninflammatory neurologic diseases, as well as healthy subjects representing the control, were investigated for immunoglobulin G (IgG) antibodies against JCPyV. To this end, an immunologic approach was employed, which consists of employing indirect enzyme-linked immunosorbent assay testing with synthetic peptides mimicking viral capsid protein 1 antigens. A significantly lower prevalence of IgG antibodies against JCPyV VP1 epitopes, with a low titer, was detected in serum samples from patients with multiple sclerosis (MS) and other neurologic diseases than in healthy subjects. Our study indicates that the prevalence of JCPyV antibodies from patients with multiple sclerosis is 50% lower than in healthy subjects, suggesting specific immune impairments. These results indicate that patients affected by neurological diseases, including MS, respond poorly to JCPyV VP1 antigens, suggesting specific immunologic dysfunctions.

Investigation of simian virus 40 (SV40) and human JC, BK, MC, KI, and WU polyomaviruses in glioma.

The gliomagenesis remains not fully established and their etiological factors still remain obscure. Polyomaviruses were detected and involved in several human tumors. Their potential implication in gliomas has been not yet surveyed in Africa and Arab World. Herein, we investigated the prevalence of six polyomaviruses (SV40, JCPyV, BKPyV, MCPyV, KIPyV, and WUPyV) in 112 gliomas from Tunisian patients. The DNA sequences of polyomaviruses were examined by PCR assays. Viral infection was confirmed by DNA in situ hybridization (ISH) and/or immunohistochemistry (IHC). The relationships between polyomavirus infection and tumor features were evaluated. Specific SV40 Tag, viral regulatory, and VP1 regions were identified in 12 GBM (10.7%). DNA ISH targeting the whole SV40 genome and SV40 Tag IHC confirmed the PCR findings. Five gliomas yielded JCPyV positivity by PCR and DNA ISH (2.7%). However, no BKPyV, KIPyV, and WUPyV DNA sequences were identified in all samples. MCPyV DNA was identified in 30 gliomas (26.8%). For GBM samples, MCPyV was significantly related to patient age (p = 0.037), tumor recurrence (p = 0.024), and SV40 (p = 0.045) infection. No further significant association was identified with the remaining tumor features (p > 0.05) and patient survival (Log Rank, p > 0.05). Our study indicates the presence of SV40, JCPyV, and MCPyV DNA in Tunisian gliomas. Further investigations are required to more elucidate the potential involvement of polyomaviruses in these destructive malignancies.

Inhibition of JC polyomavirus infectivity by the retrograde transport inhibitor Retro-2.1.

JC polyomavirus (JCPyV) is a common human pathogen that results in a chronic asymptomatic infection in healthy adults. Under conditions of immunosuppression, JCPyV spreads to the central nervous system and can cause the fatal demyelinating disease progressive multifocal leukoencephalopathy (PML), a disease for which there are no vaccines or antiviral therapies. Retro-2 is a previously identified small molecule inhibitor that was originally shown to block retrograde transport of toxins such as ricin toxin from endosomes to the Golgi apparatus and endoplasmic reticulum (ER), and Retro-2.1 is a chemical analog of Retro-2 that has been shown to inhibit ricin intoxication of cells at low nanomolar concentrations. Retro-2 has previously been shown to prevent retrograde transport of JCPyV virions to the ER, but the effect of Retro-2.1 on JCPyV infectivity is unknown. Here it is shown that Retro-2.1 inhibits JCPyV with an EC50 of 3.9 µM. This molecule inhibits JCPyV infection at dosages that are not toxic to human tissue culture cells. Retro-2.1 was also tested against two other polyomaviruses, the human BK polyomavirus and simian virus 40, and was also shown to inhibit infection at similar concentrations. Viral uncoating studies demonstrate that Retro-2.1 inhibits BKPyV infectivity in a manner similar to Retro-2. These studies demonstrate that improved analogs of Retro-2 can inhibit infection at lower dosages than Retro-2 and further optimization of these compounds may lead to effective treatment options for those suffering from JCPyV infection and PML.

Human polyomavirus modulation of the host DNA damage response.

The human DNA damage response (DDR) is a complex signaling network constituting many factors responsible for the preservation of genomic integrity. Human polyomaviruses (HPyVs) are able to harness the DDR machinery during their infectious cycle by expressing an array of tumor (T) antigens. These molecular interactions between human polyomavirus T antigens and the DDR create conditions that promote viral replication at the expense of host genomic stability to cause disease as well as carcinogenesis in the cases of the Merkel cell polyomavirus and BK polyomavirus. This review focuses on the six HPyVs with disease association, emphasizing strain-dependent differences in their selective manipulation of the DDR. Appreciation of the HPyV-DDR interface at a molecular scale is conducive to the development of novel therapeutic approaches.

COS-7 cells are a cellular model to monitor polyomavirus JC miR-J1-5p expression.

Polyomavirus JC (JCPyV) is a ubiquitous human neurotropic virus that can cause progressive multifocal leukoencephalopathy (PML), sometimes as a consequence of drug treatment for disabling diseases, including Multiple Sclerosis. JCPyV expresses microRNAs (miRNAs), and in particular miR-J1-5p, but at now we have limited knowledge regarding this aspect. In the present study the expression of JCPyV miR-J1-5p was measured in infected COS-7, to verify if and when this miRNA is expressed in a cell model of JCPyV-MAD-4 strain infection. Results showed that miR-J1-5p expression was relatively constant inside the cells from 11 days to 35 days after infection (mean: 4.13 × 105 copies/µg), and became measurable in supernatants 18 days after infection (mean: 7.20 × 104 copies/µl). miR-J1-5p expression in supernatants peaked (3.76 × 105 copies/µl) 25 days after infection and started to decrease 32 days after infection (7.20 × 104 copies/µl). These data show that COS-7 cells, already used as model for JCPyV replication cycle, can be also utilized to study JCPyV miRNAs expression, potentially opening new research avenues for diseases in which current therapeutic approaches could result in severe adverse effects (e.g. Natalizumab-associated JCPyV reactivation in Multiple Sclerosis patients). In these situations monitoring of miR-J1-5p may shed light on the mechanisms of virus reactivation and may help the clarification of the mechanisms responsible for such severe side effects.