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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., (© 2020 Wiley Periodicals, Inc.)

JC polyomavirus (JCPyV) was identified in 1971 in the brain tissue of a patient (J.C.) affected by the progressive multifocal leukoencephalopathy (PML). JCPyV encodes for the oncoproteins large T antigen (Tag) and small t-antigen (tag). These oncoproteins are responsible of the cell transformation and tumorigenesis in experimental animals. JCPyV is ubiquitous in human populations. After the primary infection, which is usually asymptomatic, JCPyV remains lifelong in the host in a latent phase. Its reactivation may occur in heathy subjects and immunocompromised patients. Upon reactivation, JCPyV could reach (i) the CNS inducing the PML, (ii) the kidney of transplant patients causing the organ rejection. Association between JCPyV, which is a small DNA tumor virus, and gliomas and colorectal carcinomas has been published. In the present investigation, we report on a new indirect ELISA with two specific synthetic peptides mimicking JCPyV VP1 immunogenic epitopes to detect specific serum IgG antibodies against JCPyV. Serum samples of healthy subjects (n = 355) ranging 2-100 years old, were analyzed by this new indirect ELISA. The linear peptides VP1 K and VP1 N resemble the natural JCPyV VP1 capsidic epitopes constituting a docking site for serum antibodies. Data from this innovative immunologic assay indicate that the overall prevalence of JCPyV-VP1 antibodies in healthy subjects is at 39%. The innovative indirect ELISA with JCPyV VP1 mimotopes seems to be a useful method to detect specific IgG antibodies against this virus, without cross-reactivity with the closely related SV40 and BKPyV polyomaviruses., (© 2018 Wiley Periodicals, Inc.)

Anti-JCV antibody status is used for PML-risk-stratification in MS patients before and during Natalizumab therapy. JCV antibodies can be detected in around 60% of MS patients, however, only a small proportion actually develop PML. As anti-viral antibodies tend to occur unspecifically, the aim of this study was to correlate JCV antibody status and index with other common anti-viral antibodies. A total of 123 samples of MS-patients were tested for anti-JCV antibodies by JCV-Stratify-ELISA at Unilabs, Denmark. The same samples were analyzed for measles, rubella, varicella zoster, EBV, and CMV IgG and IgM antibodies by ELISA, or chemiluminescence-microparticle immunoassay. For all antibody-titers correlations were calculated and group comparisons of JCV-positive and -negative patients were performed. Fifty-three patients (43.1%) were JCV negative and 70 (56.9%) positive. CMV-IgM antibodies were detected in six patients. Otherwise no IgM antibodies were detected. IgG antibodies against measles, rubella, varicella zoster, and EBV were detected in ≥97% of patients and 47 samples (38.2%) tested positive for CMV-IgG. There was no significant correlation between any of the antibody titers including JCV index, however, a significantly higher prevalence (P = 0.003) of CMV-IgG in JCV positive compared to JCV negative patients, whereas no difference was detected for measles, rubella, varicella zoster, and EBV IgG. In conclusion, the JCV antibody response in MS patients seems to be largely independent of any other anti-viral immunity. The only coincidence was found with CMV IgG antibodies which might point towards some immunological cross-reactivity in anti-viral immune response or other mechanisms leading to combined viral infections such as shared transmission. J. Med. Virol. 89:3-9, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Objective: Many JC virus antibody-positive relapsing-remitting multiple sclerosis (RRMS) patients who are stable on natalizumab switch to other therapies to avoid progressive multifocal leukoencephalopathy., Methods: We compared outcomes for all RRMS patients switching from natalizumab due to JC virus antibody positivity at 3 Swedish multiple sclerosis centers with different preferential use of rituximab and fingolimod (Stockholm, n = 156, fingolimod 51%; Gothenburg, n = 64, fingolimod 88%; Umeå, n = 36, fingolimod 19%), yielding a total cohort of N = 256 (fingolimod 55%)., Results: Within 1.5 years of cessation of natalizumab, 1.8% (rituximab) and 17.6% (fingolimod) of patients experienced a clinical relapse (hazard ratio for rituximab = 0.10, 95% confidence interval [CI] = 0.02-0.43). The hazard ratio (favoring rituximab) for adverse events (5.3% vs 21.1%) and treatment discontinuation (1.8% vs 28.2%) were 0.25 (95% CI = 0.10-0.59) and 0.07 (95% CI = 0.02-0.30), respectively. Furthermore, contrast-enhancing lesions were found in 1.4% (rituximab) versus 24.2% (fingolimod) of magnetic resonance imaging examinations (odds ratio = 0.05, 95% CI = 0.00-0.22). Differences remained when adjusting for possible confounders (age, sex, disability status, time on natalizumab, washout time, follow-up time, and study center)., Interpretation: Our findings suggest an improved effectiveness and tolerability of rituximab compared with fingolimod in stable RRMS patients who switch from natalizumab due to JC virus antibody positivity. Although residual confounding factors cannot be ruled out, the shared reason for switching from natalizumab and the preferential use of either rituximab or fingolimod in 2 of the centers mitigates these concerns. Ann Neurol 2016;79:950-958., (© 2016 American Neurological Association.)

Many neurological diseases of the central nervous system (CNS) are underpinned by malfunctions of the immune system, including disorders involving opportunistic infections. Progressive multifocal leukoencephalopathy (PML) is a lethal CNS demyelinating disease caused by the human neurotropic polyomavirus JC (JCV) and is found almost exclusively in individuals with immune disruption, including patients with human immunodeficiency virus/acquired immunodeficiency syndrome, patients receiving therapeutic immunomodulatory monoclonal antibodies to treat conditions such as multiple sclerosis, and transplant recipients. Thus, the public health significance of this disease is high, because of the number of individuals constituting the at-risk population. The incidence of PML is very low, whereas seroprevalence for the virus is high, suggesting infection by the virus is very common, and so it is thought that the virus is restrained but it persists in an asymptomatic state that can only occasionally be disrupted to lead to viral reactivation and PML. When JCV actively replicates in oligodendrocytes and astrocytes of the CNS, it produces cytolysis, leading to formation of demyelinated lesions with devastating consequences. Defining the molecular nature of persistence and events leading to reactivation of the virus to cause PML has proved to be elusive. In this review, we examine the current state of knowledge of the JCV life cycle and mechanisms of pathogenesis. We will discuss the normal course of the JCV life cycle including transmission, primary infection, viremia, and establishment of asymptomatic persistence as well as pathogenic events including migration of the virus to the brain, reactivation from persistence, viral infection, and replication in the glial cells of the CNS and escape from immunosurveillance., (© 2015 American Neurological Association.)

Objective: Progressive multifocal leukoencephalopathy (PML) is a serious side effect associated with natalizumab treatment in multiple sclerosis (MS). PML risk increases in individuals seropositive for anti-John Cunningham virus (JC) antibodies, with prolonged duration of natalizumab treatment, and with prior exposure to immunosuppressants. We explored whether the presence of lipid-specific immunoglobulin M oligoclonal bands in cerebrospinal fluid (CSF; IgM bands), a recognized marker of highly inflammatory MS, may identify individuals better able to counteract the potential immunosuppressive effect of natalizumab and hence be associated with a reduced risk of developing PML., Methods: We studied 24 MS patients who developed PML and another 343 who did not suffer this opportunistic infection during natalizumab treatment. Patients were recruited at 25 university hospitals. IgM bands were studied by isoelectric focusing and immunodetection. CSF lymphocyte counts were explored in 151 MS patients recruited at Ramon y Cajal Hospital in Madrid, Spain., Results: IgM bands were independently associated with decreased PML risk (odds ratio [OR] = 45.9, 95% confidence interval [CI] = 5.9-339.3, p < 0.0001) in patients treated with natalizumab. They were also associated with significantly higher CSF CD4, CD8, and B-cell numbers. Patients positive for IgM bands and anti-JC antibodies had similar levels of reduced PML risk to those who were anti-JC negative (OR = 1.55, 95% CI = 0.09-25.2, p = 1.0). Higher risk was observed in patients positive for anti-JC antibodies and negative for IgM bands (19% of the total cohort, OR = 59.71, 95% CI = 13.6-262.2)., Interpretation: The presence of IgM bands reflects a process that may diminish the risk of PML by counteracting the excess of immunosuppression that may occur during natalizumab therapy., (© 2015 American Neurological Association.)

JC and BK polyomaviruses (JCV and BKV) infect humans and can cause severe illnesses in immunocompromised patients. Merkel cell polyomavirus (MCPyV) can be found in skin carcinomas. In this study, we assessed the occurrence of serum antibodies against MCPyV, BKV, and JCV polyomaviruses in a healthy population of the Czech Republic. Serum samples from 991 healthy individuals (age range: 6-64 years) were examined by enzyme-linked immunoassay (ELISA) using virus-like particles (VLPs) based on the major VP1 capsid proteins of these viruses. Overall, serum antibodies against MCPyV, JCV, and BKV were found in 63%, 57%, and 69%, respectively, of this population. For all three viruses, these rates were associated with age; the occurrence of antibodies against MCPyV and JCV was highest for those older than 59 years, while the occurrence of antibodies against BKV was highest in those aged 10-19 years and 20-29 years. This is the first large study to determine the seroprevalence rates for BKV, JCV, and MCPyV polyomaviruses in the general Czech Republic population., (© 2013 Wiley Periodicals, Inc.)

An infectious etiology for bladder cancer has long been suspected. Merkel cell virus (MCV), BKV and JCV polyomaviruses are possible causative agents but data remain scarce. Therefore, we evaluated the seroresponse to these three polyomaviruses in association with bladder cancer risk. 1,135 incident bladder cancer subjects from five Spanish regions and 982 hospital controls matched by sex, age and region were included. 99% of cases were urothelial-cell carcinomas. Antibody response against MCV, BKV and JCV was measured by enzyme immunoassay using Virus-Like-Particles. Our results show a similar seroprevalence in cases and controls: 64/60% for BKV, 83/82% for MCV and 87/83% for JCV. However, among seropositive subjects, higher median seroreactivities were observed in cases compared to controls for BKV (0.84 vs. 0.70, p-value = 0.009) and MCV (1.81 vs. 0.65, p-value < 0.001). Increased bladder cancer risk was observed for BKV (OR = 1.4, 95%CI 1.04-1.8) and for MCV (OR = 1.5, 95%CI 1.2-1.9), when comparing highest to lowest seroreactivity tertiles. The associations of BKV and MCV with bladder cancer were independent of each other and neither smoking status nor disease stage and grade modified them. Furthermore, no association was observed between seroresponse to JCV and bladder cancer. Therefore, we conclude that BKV and MCV polyomavirus infection could be related to an increased bladder cancer risk., (Copyright © 2013 UICC.)

The human polyomaviruses BKV and JCV cause mostly subclinical infections in childhood. Systemical immunosuppression after organ transplantation can lead to reactivation of persistent polyomavirus infections which may cause rejection of the transplanted organ. BKV and JCV seroprevalence and serostability was measured in 441 European solid organ transplanted recipients. Baseline samples were collected on average 24 days post-transplantation and sera were then collected over an 18 months follow-up period on up to six different time points. The overall seroprevalence at baseline for BKV was 97% with very little change over time. Prevalence for JCV was 76% at baseline and increased to 80% at the end of follow-up. BKV seroprevalence was highest in the youngest age group (100%) and decreased with increasing age (92% in the oldest age group; P < 0.0001), while JCV increased with age (69% vs. 81%; P = 0.020). Antibody reactivities for both BKV and JCV increased significantly with time (P = 0.0002 and P < 0.0001, respectively). Among the 406 patients with several samples, 94% were stably seropositive for BKV and 1% remained seronegative during the follow-up. JCV antibody stability was somewhat lower: 67% remained stably seropositive and 13% seronegative. While seroprevalence of BKV and JCV decrease and increase with age, respectively, both polyomaviruses showed significant increasing antibody reactivity over time in organ transplanted recipients at the onset of immunosuppression., (Copyright © 2012 Wiley Periodicals, Inc.)

The hypothesis was examined that physiologic variation of estrogen concentrations during the menstrual cycle can provoke BK virus (BKV) excretion. BKV and JCV viral loads were determined in urine specimens obtained almost daily from 20 healthy, non-pregnant women over 2 months. Asymptomatic urinary shedding of BKV was observed in 123 (12.0%) of 1,021 specimens from 11 (55%) study subjects. Two subjects excreted JCV in their urine, with one subject excreting detectable JCV in all urine specimens. Analysis of 36 complete menstrual cycles revealed no difference in the prevalence of BKV excretion between pre-ovulatory and post-ovulatory phases of the menstrual cycle. The unexpected day-to-day variability in BKV excretion suggests that as yet unidentified factors may contribute to the periodic shedding of BKV by healthy women., (Copyright © 2012 Wiley Periodicals, Inc.)

Asymptomatic polyomaviruria of pregnancy has been documented in point prevalence studies, but little attention has been given to the dynamics of polyomavirus excretion during pregnancy because of its benign course. We tested the hypothesis that the frequency and/or magnitude of polyomavirus excretion would increase as pregnancy progresses. Urine specimens were obtained prospectively from 179 healthy women during uncomplicated pregnancies and 37 healthy non-pregnant women. Real-time polymerase chain reaction was used to determine BK virus (BKV) and JC virus (JCV) viral loads in urine, blood, and rectal and vaginal swabs collected during routine obstetric and gynecologic clinic visits. Asymptomatic urinary shedding of BKV and/or JCV was observed in 384 (48.0%) of 800 specimens from 100 (55.8%) pregnant women. BKV excretion was more common in pregnant than non-pregnant women (41.3% vs. 13.5%, P = 0.0026). The frequency of JCV excretion was no different in pregnant compared to non-pregnant women. The frequency and magnitude of polyomavirus shedding did not vary with gestational age. Post-partum shedding of BKV, but not JCV, rapidly decreased to undetectable levels. Pregnancy-associated BKV excretion begins early in pregnancy and terminates rapidly post-partum. Neither the frequency nor magnitude of BKV or JCV shedding increased with pregnancy progression. Further study into the host factors that regulate pregnancy-associated BKV excretion may allow identification of the host factors that predict susceptibility to BKV-associated diseases in immune compromised patients., (Copyright © 2012 Wiley Periodicals, Inc.)

Objective: A study was undertaken to define the prevalence of anti-JC virus (JCV) antibodies in multiple sclerosis (MS) patients and to evaluate the analytical false-negative rate of a 2-step anti-JC virus antibody assay., Methods: STRATIFY-1 is an ongoing, longitudinal, observational study of relapsing MS patients in the United States who are being treated or considering treatment with natalizumab. Baseline serum and plasma samples were collected for anti-JC virus antibody detection using an analytically validated, 2-step, virus-like particle-based enzyme-linked immunosorbent assay. Urine was collected for JC virus DNA detection., Results: At baseline (n = 1,096), overall anti-JC virus antibody prevalence was 56.0% (95% confidence interval [CI], 53.0-59.0) in STRATIFY-1 patients, with an assay false-negative rate of 2.7% (95% CI, 0.9-6.2). Prevalence was significantly lower in females (53.4%; 95% CI, 49.9-56.8) than males (64.3%; 95% CI, 58.2-70.0) and increased with age, p = 0.0019 and p = 0.0001, respectively. Prevalence was similar in patients regardless of natalizumab exposure or prior immunosuppressant use, p = 0.9709 and p = 0.6632, respectively. STRATIFY-1 results were generally consistent with those observed in another large North American cohort, TYGRIS-US (n = 1,480)., Interpretation: Baseline results from STRATIFY-1 are consistent with other studies utilizing this assay that demonstrate a 50 to 60% prevalence of anti-JC virus antibodies, a low false-negative rate, and an association of increasing age and male gender with increasing anti-JC virus antibody prevalence. Neither natalizumab exposure nor prior immunosuppressant use appear to affect prevalence. Longitudinal data from STRATIFY-1 will confirm the stability of anti-JC virus antibody prevalence over time., (Copyright © 2011 American Neurological Association.)

JC virus (JCV) DNA in the cerebrospinal fluid (CSF) provides the laboratory confirmatory diagnosis of progressive multifocal leukoencephalopathy (PML) in patients whose clinical symptoms and magnetic resonance imaging findings are consistent with PML.The Laboratory of Molecular Medicine and Neuroscience (LMMN), National Institute of Neurological Disorders and Stroke (NINDS), National Institutes of Health (NIH), made the confirmatory laboratory diagnosis in 35 multiple sclerosis (MS) patients treated with natalizumab. Thirteen patients had 3 or more CSF samples taken from weeks to months following PML diagnosis. Seven of the 13 patients demonstrated persistence of JCV DNA in the CSF even though all patients experienced immune reconstitution inflammatory syndrome (IRIS), 11 patients had plasma exchange, and 2 had immunoabsorption. Specific anti-JCV antibody was measured in plasma/sera samples from 25 of the 35 patients. Most of the samples showed moderate to high or rising antibody levels from the time of PML diagnosis. However, plasma from 1 patient at or near the time of PML diagnosis had a titer considered seronegative and 2 other plasma samples from patients had titers considered at baseline for seropositivity. In several PML cases, viral persistence and neurological deficits have continued for several years, indicating that once initiated, JCV infection may not entirely clear, even with IRIS.

Objective: Analyses were conducted to determine the clinical utility of measuring JC virus (JCV) DNA in blood or urine of natalizumab-treated multiple sclerosis (MS) patients to predict the risk of progressive multifocal leukoencephalopathy (PML)., Methods: A total of 12,850 blood and urine samples from nearly 1,400 patients participating in natalizumab clinical trials were tested for JCV DNA using a commercially available quantitative polymerase chain reaction (qPCR) assay. A subset of these samples was also tested using a more sensitive qPCR assay developed at the National Institutes of Health (NIH)., Results: At the time natalizumab dosing was suspended, JCV DNA was detected in plasma by the commercial assay in 4 of 1,397 (0.3%) patients; the NIH assay confirmed these positive samples and detected JCV DNA in an additional 2 of 205 (1%) patients who tested negative with the commercial assay. None of these 6 JCV DNA positive patients developed PML. In a 48-week study testing the safety of natalizumab redosing, JCV DNA was detected in plasma of 6 of 1,094 (0.3%) patients, none of whom developed PML. Urine at baseline and week 48 was assessed in 224 patients; 58 (26%) were positive at baseline, and 55 (25%) were positive after 48 weeks of natalizumab, treatment. JCV DNA was not detected in peripheral blood mononuclear cells from any of these 1,094 patients before or after natalizumab treatment. In 5 patients who developed PML, JCV DNA was not detected in blood at any time point before symptoms first occurred., Interpretation: Measuring JCV DNA in blood or urine with currently available methods is unlikely to be useful for predicting PML risk in natalizumab-treated MS patients.

Objective: A study was undertaken to establish an enzyme-linked immunosorbent assay (ELISA) to detect JC virus (JCV)-specific antibodies in multiple sclerosis (MS) patients, and to evaluate its potential utility for identifying patients at higher or lower risk (ie, risk stratification) of developing progressive multifocal leukoencephalopathy (PML)., Methods: A 2-step assay for detecting and confirming the presence of anti-JCV antibodies in human serum and plasma was developed and demonstrated to be both sensitive and specific. ELISA cutpoints were statistically established using sera from >800 MS patients from natalizumab clinical studies. Subsequently, this assay was used to determine the presence of anti-JCV antibodies in natalizumab-treated PML patients where serum samples were collected 16-180 months prior to the diagnosis of PML., Results: In our evaluation of natalizumab-treated MS patients, 53.6% tested positive for anti-JCV antibodies, with a 95% confidence interval of 49.9 to 57.3%. The false-negative rate of the ELISA was calculated to be approximately 2.5%, with an upper 1-sided confidence limit of 4.4%. Notably, we observed anti-JCV antibodies in all 17 available pre-PML sera samples, which was significantly different from the 53.6% seropositivity observed in the overall MS study population (p < 0.0001)., Interpretation: This 2-step assay provides a means to classify MS patients as having detectable or not detectable levels of anti-JCV antibodies. The finding that all 17 of the pre-PML samples that were available tested seropositive, and none tested seronegative, warrants further research on the clinical utility of the anti-JCV antibody assay as a potential tool for stratifying MS patients for higher or lower risk of developing PML.

The formation and progression of mudulloblastoma (MB) is poorly understood. However, somatic inactivation of pRb/p105, in combination with a somatic or a germ-line TP53 inactivation, leads to MB in a mouse model. Presently, there is no specific evidence of pathway/s alterations for the other two members of the retinoblastoma family, pRb2/p130 and/or p107 in MB. JC virus (JCV) is a human polyomavirus. Although there is no firm evidence that this virus plays a causal role in human neoplasia, it has been clearly proven that JCV is highly oncogenic when injected into the brain of experimental animals. The mechanism of JCV-induced tumorigenesis is not entirely clear. However, several studies relate the oncogenic properties of JCV mainly to its early protein large T-antigen (T-Ag), which is able to bind and inactivate both TP53 and Rb family proteins. Here, we compared the protein expression profiles of p53, p73, pRb family proteins, and PCNA, as main regulators of cell proliferation and death, in different cell lines of mouse primitive neuroectodermal tumors (PNET), either T-Ag-positive or -negative, and in human MB cell lines. Our goal was to determine if changes in the relative expression of these regulators could trigger molecular perturbations underlying MB pathogenesis in mouse and human cells. Our results support that the presence of JCV T-Ag may interfere with the expression of pRb family proteins, specific p73 isoforms, and p53. In turn, this "perturbation" may trigger a network of signals strictly connected with survival and apoptosis., ((c) 2010 Wiley-Liss, Inc.)

The regulatory proteins of polyomaviruses, including small and large T antigens, play important roles, not only in the viral life cycle but also in virus-induced cell transformation. Unlike many other tumor viruses, the transforming proteins of polyomaviruses have no cellular homologs but rather exert their effects mostly by interacting with cellular proteins that control fundamental processes in the regulation of cell proliferation and the cell cycle. Thus, they have proven to be valuable tools to identify specific signaling pathways involved in tumor progression. Elucidation of these pathways using polyomavirus transforming proteins as tools is critically important in understanding fundamental regulatory mechanisms and hence to develop effective therapeutic strategies against cancer. In this short review, we will focus on the structural and functional features of one polyomavirus transforming protein, that is, the small t-antigen of the human neurotropic JC virus (JCV) and the simian virus, SV40., ((c) 2007 Wiley-Liss, Inc.)

Chemokines (chemoattractant cytokines) and their receptors are present in the brain and may play roles in both neurodevelopment and neuropathology. Increased brain levels of monocyte chemoattractant protein-1 (MCP-1), also known as CCL2, are found in patients with human immunodeficiency virus type 1 (HIV-1)-associated dementia and other acute and chronic neurologic diseases. Although the function of CCL2 in the brain is unclear, it is believed that upregulation of this chemokine during neuropathologic or neuroinflammatory conditions leads to recruitment of activated monocytes into the brain, where they differentiate into macrophages producing neurotoxic and inflammatory molecules. We recently showed that human fetal brain-derived progenitor cells are susceptible to HIV-1 and JC virus infection, and that differentiation toward an astrocyte phenotype increased virus production from these cells. In the current study, we found that in the absence of infection, progenitors produced moderate levels of CCL2 (5.6 ng per million cells). Astrocyte differentiation over 3 weeks increased CCL2 protein levels 30-fold in a biphasic manner, whereas neuronal differentiation decreased production 20-fold. Electromobility shift assays (EMSAs) demonstrated increased nuclear NF-kappaB levels within 2 h of initiating astrocyte differentiation, and inhibitors of NF-kappaB activation partially blocked the CCL2 increase in differentiating astrocytes. Transfection of progenitors with mutated CCL2 promoter/CAT reporter constructs showed that the distal promoter region, containing NF-kappaB and NF-I binding sites, is important for differentiation-induced CCL2 upregulation. Together these results suggest that the transcription factor NF-kappaB, and possibly NF-I, contribute to the upregulation of CCL2 chemokine production during the differentiation of human progenitor cells toward an astrocyte phenotype., ((c) 2005 Wiley-Liss, Inc.)

Two related polyomaviruses, JC virus (JCV) and BK virus (BKV), commonly cause lifelong infections in humans, with periodic reactivation manifesting as viral shedding in urine. Because JCV can infect lymphocytes and cause chromosomal damage, it is a plausible candidate to cause non-Hodgkin lymphoma (NHL). To test this hypothesis, we measured IgG antibodies to JCV and BKV capsids using a virus-like particle enzyme immunoassay in 3 separate groups of subjects. First, in a U.S. population-based case-control study of NHL (724 cases, 622 controls), we found lower JCV antibody levels in cases than controls (median optical density = 0.12 vs. 0.21, p < 0.0001); likewise, JCV seroprevalence was lower in cases (49% vs. 59%, adjusted odds ratio [OR] = 0.70, 95% confidence interval [CI] = 0.56-0.87). In contrast, BKV antibody levels did not differ between groups. Second, we found that JCV and BKV antibody levels changed little over time among 24 NHL patients receiving chemotherapy. Third, we evaluated 126 homosexual men, of whom 46 were shedding JCV and 14 were shedding BKV in urine. Antibody levels were much higher in shedders than non-shedders (JCV: median optical density = 0.67 vs. 0.07, p < 0.0001; BKV: 0.87 vs. 0.40, p = 0.003), indicating that these antibodies are a marker for viral replication. Because no deficit of BKV antibody was seen in NHL cases, and because antibody levels did not change materially with chemotherapy, we suggest that the lower levels of JCV antibody observed in NHL patients may not be due entirely to a disease or treatment effect. Additional research is needed to determine whether JCV replication is decreased in individuals with NHL and whether these findings are consistent with an etiologic role for JCV in NHL., (Copyright 2005 Wiley-Liss, Inc)

To investigate if polyomavirus infection during pregnancy is linked to development of neuroblastoma in the child, serum samples of 115 index mothers from the pregnancy where the child eventually developed neuroblastoma were identified and matched with serum samples from 8 control mothers per index mother. The samples were tested for specific IgG and IgM antibodies to BK and JC virus using enzyme immunoassays based on purified yeast-expressed virus-like particles (VLPs). The serum samples as well as 10 neuroblastoma cell lines were also analyzed using Real Time (TaqMan) PCR for detection and quantification of BK virus DNA. The BK virus IgG seroprevalence was similar among index mothers (80%) and control mothers (83%) [OR 0.8; 95% confidence interval (95% CI): 0.5-1.3]. BK virus IgM was also not associated with neuroblastoma risk (OR was OR = 0.6; 95% with CI, 0.2-1.9). Also JC virus had no association, neither for IgG (OR = 0.9; 95% CI, 0.6-1.4) nor for IgM (OR = 0.9; 95% CI, 0.4-1.9). All serum samples and all neuroblastoma cell lines were negative for BKV DNA. In summary, a comprehensive cohort using both serology and polyomavirus DNA detection found no evidence for association between BKV or JCV polyomaviruses and neuroblastoma.

Molecular studies suggest that the simian polyomavirus SV40 is present in the human population, possibly introduced in contaminated polio vaccine. However, no recent seroepidemiological data exist in England on SV40 or on the two human polyomaviruses, BKV and JCV. A comparative age seroprevalence study was undertaken on 2,435 residual sera from 1991 by haemagglutination inhibition (HI) for BKV and JCV, and virus neutralisation for SV40. The overall rates of seropositivity for BKV and JCV were 81% and 35%, respectively, and each was significantly related to age (P < 0.001). BKV seroprevalence reached 91% at 5-9 years of age, but JCV seroprevalence reached only 50% by age 60-69 years. There was a highly significant association between BKV antibody titre and age (P < 0.001), titres decreasing linearly at a rate of 8.7% per 10 years (95% CI = 7.4-10% drop). Significantly more males than females had antibody to JCV (P = 0.013). In individuals under 40 years of age there was a significant negative association between the presence of antibody to BKV and JCV (P < 0.001). By contrast, the antibody prevalence to SV40 remained at 1.3-5% throughout all age groups and titres were low. There was a significant positive association between the presence of antibody to SV40 and antibody to both BKV (P < 0.001) and JCV (P = 0.009), and also to the geometric mean titre (GMT) of BKV antibody (P = 0.011). The results indicate that BKV and JCV are transmitted by different routes. There is no serological evidence that SV40 entered the human population during the past 80 years, and the possibility of cross-reaction with BKV or JCV antibody must be considered., (Copyright 2003 Wiley-Liss, Inc.)

The human polyomavirus, JC virus (JCV), provides an excellent model system to investigate the reciprocal interaction of the immune and nervous systems. Infection with JCV occurs during childhood and the virus remains in the latent state with no apparent clinical symptoms. However, under immunosuppressed conditions, the virus enters the lytic cycle and upon cytolytic destruction of glial cells, causes the fatal demyelinating disease of the central nervous system (CNS), named progressive multifocal leukoencephalopathy (PML). In this short review, we discuss the molecular pathogenesis of PML by highlighting the role of the immune system in modulating JCV gene activation and replication, and the latency/reactivation of this virus upon immunosuppression. Further, due to the higher incidence of PML among AIDS patients, we further elaborate on the cross-talk between JCV and HIV-1 by direct and indirect pathways that lead to enhanced expression of the JCV genome., (Copyright 2002 Wiley-Liss, Inc.)

Progressive multifocal leukoencephalopathy (PML) is a fatal, demyelinating disease caused by JC virus (JCV) in patients with severe immunosuppression. We studied the JCV-specific cellular and humoral immune response in 7 healthy donors (HD), 6 human immunodeficiency virus-1 (HIV-1)-infected patients without PML (HIV), 4 HIV-1-negative patients with PML (PML), and 8 HIV-1-positive patients with PML (HIV/PML). As antigens, recombinant virus-like particles of the major structural protein VP1 (VP1-VLP) of JCV, tetanus toxoid (TT), or the mitogen phytohemagglutinin (PHA) were used. Proliferation of peripheral blood mononuclear cells (PBMC) after stimulation with the VP1-VLP was significantly suppressed in PML and HIV/PML patients compared to HD. After antigen stimulation the production of interferon-gamma (IFN-gamma) was reduced in PML, in HIV/PML, and in HIV patients. The production of interleukin-10 (IL-10), however, was elevated in HIV/PML patients. Neither proliferation nor cytokine production correlated with the presence of JCV DNA in PBMC. The immunoglobulin G serum antibody titer to the VP1-VLP was slightly elevated in HIV, elevated in PML, and highly elevated in HIV/PML patients compared to HD. The development of PML appears to coincide with a general impairment of the Th1-type T-helper cell function of cell-mediated immunity.

By screening consecutive autopsy cases with an antibody that recognizes human polyomaviruses, we found a case of malignant lymphoma in which the virus infection was confined to epithelia of the renal calyces, renal pelvis, ureter, and urinary bladder. The virus was confirmed as BK virus by a specific monoclonal antibody against BK virus T antigen, and numerous virus particles were identified by electron microscopy. The results showed that BK virus is a human urotheliotrophic virus.

Seven monoclonal antibodies (MAbs) to polyomavirus JC were produced, and one was selected for use in immunofluorescence (IF) tests on brain material from patients with suspected progressive multifocal leucoencephalopathy (PML). The selected MAb (5.12.2) reacted by IF with JC-infected primary human foetal glial (PHFG) cell cultures (titre 1/200,000) but not with BK-infected human embryo lung (HEL) fibroblasts (titre less than 1/20). Its haemagglutination-inhibition (HI) titre was high (greater than 1/5 x 10(6)) against JC virus but low (less than 1/5) against BK virus. A diagnosis of PML was confirmed on brain biopsy or at postmortem in four patients, two of whom were also infected with human immunodeficiency virus (HIV). In one of the patients, specific detection of JC virus antigen had not been possible using our routine high titred JC and BK human sera due to interference by JC antibody present in the cerebrospinal fluid (CSF). Viral antigen was, however, detected with the MAb 5.12.2.

A set of recombinant DNAs, representing several isolates of JCV, has been constructed. Some of these cloned DNAs transform nonpermissive primary hamster brain cells. Analysis of the early viral mRNAs and proteins isolated from these transformants revealed similarities with the large and small T messages and proteins of SV40. The region of the JCV genome from 0.63 to 0.73 map units was sequenced by the Maxam-Gilbert technique. This segment of DNA specifies several regulatory elements and the amino-terminal portion of the early viral proteins. Comparisons with the analogous regions in the polyomaviruses SV40 and BKV, confirm the close evolutionary relationship of these three viruses. Similarities include palindromic and symmetrical sequences near the origins of DNA replication, binding sites for the large T proteins, and AT-rich region (the Goldberg-Hogness or TATA box), and a large tanden duplication or triplication to the late side of the replication origin (however, these sequences differ). Homology between the sequences coding for the early proteins is also evident (27 of the first 33 amino acids are shared). Of greater interest are features of the JCV genome which differ from those of other polyomaviruses. Absent in JCV and BKV are sequences which resemble the third T antigen-binding site of SV40. In addition, a set of sequences present in JCV and BKV DNA (33 nucleotides in JCV, 22 nucleotides in BKV), and located near a 17 base pair palindrome shared by all three viruses, is missing in SV40 DNA. Another sequence, GGGXGGAG, which is repeated several times in many polyomaviruses and adenoviruses, and which is thought to play a role in DNA replication and/or transcription, is not found in the JCV sequence presented. Finally, the tandem repeat of JCV, unlike those of BKV and SV40, includes the Goldberg-Hogness sequence.

In 139 patients with breast or lung carcinoma, malignant melanoma, and Hodgkin's or non-Hodgkin's lymphoma, JC and BK virus serology (hemagglutination inhibition) and urinary polyomavirus excretion (cytology and immunofluorescence microscopy) were studied. Overall, 18 of 70 patients with paired sera (26%) had titer increases against JC or BK virus, and 11 of 114 patients (10%) had evidence for urinary excretion. The infection rate was highest in patients with Hodgkin's or non-Hodgkin's lymphoma (approximately 40%). Serum HAI antibody titers against JC and BK viruses appeared similar to age-matched controls in each patient group--with the exception of decreased BK titers at diagnosis in patients with resected malignant melanoma and increased JC virus antibody titers at diagnosis in patients with poor-prognosis non-Hodgkin's lymphoma (IC-2). The biologic significance of these observations remains to be determined. Initial antibody titers against JC or BK virus were not of prognostic value for subsequent survival in any of the tested patient groups. Both nonspecific immunotherapy and aggressive, multidrug chemotherapy had surprisingly little effect on serum HAI titers to JC or BK virus. Patients with poor-prognosis non-Hodgkin's lymphoma appear especially suitable for further investigation of JC and BK virus infections. Study of nonbrain, nonurinary-tract tissues may disclose other parenchymal sites of polyomavirus replication in these patients.

Current techniques for the measurement of BK papovavirus (BKV) specific IgM include sucrose density gradient centrifugation followed by hemagglutination inhibition (HAI) or indirect immunofluorescent (IF) staining of BKV infected cells using a fluorescein conjugated anti-human IgM antibody. These techniques are cumbersome and labor intensive and do not lend themselves to testing large numbers of sera. A solid phase radioimmunoassay (RIA) was developed to facilitate the measurement of BKV IgG and IgM in large numbers of sera. Solid phase antigen was prepared by adsorbing CsCl purified BKV antigen to polyvinyl chloride microtiter plates. Following reaction with serum, bound immunoglobulin was detected with iodinated goat anti-human IgG or IgM. RIA for the measurement of BKV IgG was sensitive with titers approaching 10(-6). Determination of IgG titers by RIA and HAI showed good agreement (P less than 0.01, correlation coefficient = 0.74). Measurement of BKV IgM was not affected by the presence of BKV IgG as evidenced by sucrose density gradient fractionation of IgM positive sera, removal of IgG by treatment with S. aureus protein A, and addition of BKV IgG to BKV IgM. Rheumatoid factor (RF) gave false positive IgM titers in the presence of BKV IgG when RF titers were greater than or equal to 1:640 by latex agglutination testing and BKV IgG levels exceed 1:256 by HAI. False positives due to RF could be eliminated by treatment of sera with sheep anti-human IgG antisera. RIA for BKV IgM was specific as sera containing JCV-, cytomegalovirus (CMV)-, rubella-, or hepatitis B core antibody (anti HBc)-IgM were negative by RIA. RIA detected BKV IgM in several sera from renal dialysis or allograft patients with titers ranging from 1:400 to 1:128,000 and demonstrated that BKV IgM persisted in sera of renal allograft patients for as long as 343 days post transplantation.

We have examined both the naturally occurring and passage-induced variation in the genome of the human polyomavirus, JCV. JCV DNA was extracted directly from diseased brain tissue of ten cases of progressive multifocal leukoencephalopathy (PML) and the DNA population from any one case was homogeneous with respect to length and restriction endonuclease cleavage patterns. However, a comparison of cloned JCV DNAs revealed that the viral DNAs derived from different cases of PML were not identical. We identified a hypervariable region near the origin of DNA replication, and we demonstrated that there are two genetically distinguishable subtypes of the virus. After growth in vitro, the DNA from certain isolates of JCV became heterogeneous in size. Deletions of up to 12% of the genome occurred after as few a three low multiplicity passages in human glial cells. Most of the deletions mapped within the region presumed to code for T antigen. In addition, we examined the physical properties of JCV from brain and, as expected, they were typical of the polyomavirus genus.

Antibody responses to JCV viral and T antigens were determined in owl monkeys following infection with JCV. Antibody to JCV virion antigen was present in all inoculated monkeys, arising within one months. These antibodies gradually diminished in all nontumor-bearing monkeys and in 10/13 tumor-bearing monkeys over the ensuing 12 months. Antibody to T antigen became evident later in most inoculated monkeys but had a marked variable course and did not correlate well with tumor production. The antibody patterns suggest that JCV is basically nonpermissive in owl monkeys and that the viral genome is retained in a prolonged persistent stage before progressing rapidly into a tumor.

Human polyomavirus BK-like isolates were subjected to restriction endonuclease analysis with the enzymes EcoRI and Hind III. End-point dilution was used to obtain homogeneous virus pools for DNA analysis and to remove JC virus from a mixed stock. The results of Hind III digestion suggested that two subgroups could be distinguished. Several BK-like isolates were purified and rabbit antisera raised. The isolates were compared with each other and with BK and JC viruses by haemagglutination-inhibition (HI) and by neutralisation. JC virus was serologically distinct, but all the other isolates showed some cross-reactivity. Two subgroups were again evident: GS and PG were with prototype BK in subgroup 1, and MG and IV were in subgroup 2. Two isolates, AS and SB, reacted with isolates of both subgroups 1 and 2 but were distinct from one another: their genome was similar to subgroup 1 isolates. Typing by HI or by neutralisation may form a basis for grouping BK-like polyomavirus isolates.

Serologic studies were performed to determine if BKV and JCV are transmitted congenitally and to assess if the virus present in donor kidney contributes to virus activity in renal transplant recipients. Fourteen of 100 normal women showed antibody rise to BKV or JCV during pregnancy; in all instances, these were reactivation infections occurring in initially seropositive women. BKV-and JCV-specific antibodies of the IgM class were not detected in over 300 umbilical cord sera. Thus, there was no evidence of congenital transmission of BKV or JCV. BKV-seronegative renal transplant recipients who received kidneys from BKV-seropositive donors had a frequency of BKV infection which was about four times greater than that in BKV-seronegative recipients receiving kidneys from BKV-seronegative donors. These data suggest that BKV in donor kidney contributes to primary BKV infections in renal transplant recipients.