JC Virus DNA Research Articles

JC virus-associated central nervous system diseases in HIV-infected patients in Brazil: clinical presentations, associated factors with mortality and outcome

INTRODUCTION: Several presentations of neurologic complications caused by JC virus (JCV) in human immunodeficiency virus (HIV)-infected patients have been described and need to be distinguished from the "classic" form of progressive multifocal leukoencephalopathy (PMl). The objectives of this study were: 1) to describe the spectrum and frequency of presentations of JCV-associated central nervous system (CNS) diseases; 2) identify factors associated with in-hospital mortality of patients with JCV-associated CNS disease; and 3) to estimate the overall mortality of this population. MATERIAL AND METHODS: This was a retrospective study of HIV-infected patients admitted consecutively for JCVassociated CNS diseases in a referral teaching center in São Paulo, Brazil, from 2002 to 2007. All patients with laboratory confirmed JCV-associated CNS diseases were included using the following criteria: compatible clinical and radiological features associated with the presence of JCV DNA in the cerebrospinal fluid. JCV-associated CNS diseases were classified as follows: 1) classic PMl; 2) inflammatory PMl; and 3) JC virus granule cell neuronopathy (GCN). RESULTS: We included 47 cases. JCV-associated CNS diseases were classified as follows: 1) classic PMl: 42 (89%); 2) inflammatory PMl: three (6%); and 3) JC virus GCN: four (9%). Nosocomial pneumonia (p = 0.003), previous diagnosis of HIV infection (p = 0.03), and imaging showing cerebellar and/or brainstem involvement (p = 0.02) were associated with in-hospital mortality. overall mortality during hospitalization was 34%. CONCLUSIONS: Novel presentations of JCV-associated CNS diseases were observed in our setting; nosocomial pneumonia, previous diagnosis of HIV infection, and cerebellar and/or brainstem involvement were associated with in-hospital mortality; and overall mortality was high.

JC virus-associated central nervous system diseases in HIV-infected patients in Brazil: Clinical presentations, associated factors with mortality and outcome

IntroductionSeveral presentations of neurologic complications caused by JC virus (JCV) in human immunodeficiency virus (HIV)-infected patients have been described and need to be distinguished from the “classic” form of progressive multifocal leukoencephalopathy (PML). The objectives of this study were: 1) to describe the spectrum and frequency of presentations of JCV-associated central nervous system (CNS) diseases; 2) identify factors associated with in-hospital mortality of patients with JCV-associated CNS disease; and 3) to estimate the overall mortality of this population. Material and methodsThis was a retrospective study of HIV-infected patients admitted consecutively for JCV-associated CNS diseases in a referral teaching center in Sao Paulo, Brazil, from 2002 to 2007. All patients with laboratory confirmed JCV-associated CNS diseases were included using the following criteria: compatible clinical and radiological features associated with the presence of JCV DNA in the cerebrospinal fluid. JCV-associated CNS diseases were classified as follows: 1) classic PML; 2) inflammatory PML; and 3) JC virus granule cell neuronopathy (GCN). ResultsWe included 47 cases. JCV-associated CNS diseases were classified as follows: 1) classic PML: 42 (89%); 2) inflammatory PML: three (6%); and 3) JC virus GCN: four (9%). Nosocomial pneumonia (p=0.003), previous diagnosis of HIV infection (p=0.03), and imaging showing cerebellar and/or brainstem involvement (p=0.02) were associated with in-hospital mortality. Overall mortality during hospitalization was 34%. ConclusionsNovel presentations of JCV-associated CNS diseases were observed in our setting; nosocomial pneumonia, previous diagnosis of HIV infection, and cerebellar and/or brainstem involvement were associated with in-hospital mortality; and overall mortality was high.

0854 The sensitivity of JC virus DNA detection vs JC virus antibody testing in a natalizumab-treated group of relapsing MS patients

IntroductionNatalizumab treatment for relapsing remitting multiple sclerosis (RRMS) is associated with a risk of progressive multifocal leukoencephalopathy (PML) due to reactivation of latent JC virus (JCV). We prospectively screened for...

144 Case report of severe reactivation of multiple sclerosis in a patient who had recently discontinued Natalizumab and who did not have progressive multifocal encephalopathy (PML)

BackgroundNatalizumab has been shown to be a cost-effective treatment for rapidly evolving severe relapsing–remitting multiple sclerosis (MS). However concern has arisen over the increased risk of PML with duration of...

Value of MRI as a surrogate marker for PML in natalizumab long-term therapy

Five years after the introduction of natalizumab into therapy of active multiple sclerosis (MS), the number of natalizumab-associated progressive multifocal leukoencephalopathy (PML) cases has reached more than 150, with a mortality rate of 20% [1]. There is clear evidence that delay in making the diagnosis of PML is a critical parameter for prognosis. Despite the development of JC-virus (JCV) testing as a putative surrogate marker, there is still a critical need for better detection of early stages of PML. Here we report an asymptomatic case of PML with an initially negative JCV DNA PCR test in cerebrospinal fluid recognized by frequent MRI scans and describe the further benign disease course. A 28-year-old previously healthy Caucasian female was diagnosed with MS in 1995. She was treated with IFN-b-1a (Rebif 22 l) and experienced no relapses until 2007. In December 2007 the patient suffered from a severe relapse with dysarthria and paraparesis. These symptoms subsided after methylprednisolone therapy; however, a control MRI in February 2008 showed new contrast-enhancing brainstem lesions. Owing to disease progression natalizumab therapy was started in March 2008. Annual MRI controls from 2008 through 2010 revealed no new lesions. An elective control MRI in March 2011 showed a new hyperintense T2 lesion (1.0 9 0.5 cm) in the white matter of the left frontal lobe (Fig. 1b). PML infection was suspected; however, high-sensitivivity JCVPCR (university laboratory) was negative, and treatment was continued. The subsequent MRI in May 2011 revealed slight enlargement of the lesion with a mild increase of signal at the borders after contrast injection, suggesting peripheral enhancement (Fig. 1c). These changes were erroneously interpreted as a new MS lesion, and a possible switch of the therapy was discussed with the patient. Yet a control MRI in July 2011 revealed marked progression of the lesion (5 9 3 cm, Fig. 1d). Therefore, the patient was admitted to our hospital with suspected PML. The patient exhibited no neurological symptoms on admission. Family members did not report any behavioral changes; she continued working and planned traveling for holidays. The only possibly new finding was a slight deterioration of short-term verbal memory in neuropsychological testing. The JCV-PCR revealed 6230 copies/ml in CSF. JCV serology was also positive. The natalizumab therapy (total of 37 infusions) was discontinued, and plasma exchange immediately started. During the last control visit (3 weeks after PLEX) the patient was still asymptomatic, and MRI demonstrated an unchanged white matter lesion in the frontal lobe without contrast enhancement. Our case report illustrates several interesting points. Firstly, the radiological manifestation of PML can be present before clinical deterioration. Depending on the localization of lesions, patients may stay asymptomatic for quite a long period of time. Our patient had no deterioration for at least 4 months after the first changes in MRI had been detected. Secondly, PML can manifest as a monofocal condition and may be difficult to differentiate from a new MS lesion. The observation that PML lesions generally occur in areas not previously affected by multiple sclerosis may be helpful [2]. Thirdly, the first CSF analysis may be I. Ayzenberg (&) N. Trampe R. Gold K. Hellwig Department of Neurology, St. Josef Hospital, Ruhr University Bochum, Bochum, Germany e-mail: ilya_ayzenberg@yahoo.com

Plasma levels of JC virus are sensitivity and specific for detecting and predicting progressive multifocal leukoencephalopathy in HIV patients

Aims: HIV-1 infection represents the most common immunosuppressive condition associated with progressive multifocal leukoencephalopathy (PML). Materials & methods: Nested PCR and quantitative real-time PCR (qPCR) for JC virus (JCV) DNA was performed on serial plasma samples obtained from 14 HIV patients with PML and 27 matched controls. Results: JCV large T antigen (LT) DNA was detected via qPCR in 11 out of 14 (79%) PML patients at disease onset and four out of 27 (15%) controls (p < 0.001). JCV LT qPCR was associated with PML diagnosis, duration of known HIV infection, absence of a prior AIDS-defining illness and absence of combination antiretroviral therapy (p < 0.001; R2 = 0.35). JCV LT qPCR was more likely to be positive in the 8 months prior to PML diagnosis compared with earlier samples (p = 0.01). Conclusion: Detection of JCV DNA in plasma of HIV infected patients via qPCR may represent a valuable test for identifying patients at risk of developing PML.

Individuals infected with JC polyomavirus do not present detectable JC virus DNA in oropharyngeal fluids

JC virus (JCV) is ubiquitous in the human population. Primary infection normally occurs during childhood and is followed by a lifelong persistent infection. The main mode of transmission remains unknown. Several authors have hypothesized that JCV transmission occurs through the respiratory route, and that respiratory secretions could represent a possible source of viral particles. The present study intended to evaluate oropharyngeal fluids from patients infected with JCV, in order to ascertain if respiratory secretions could indeed constitute a source of exposure to this polyomavirus. Oropharyngeal washing samples from 25 patients co-infected with JCV and human immunodeficiency virus type 1 were evaluated for the presence of JCV DNA. Regardless of the titre of antibodies or the presence of viral urinary excretion, JCV genome was not detected in oropharyngeal samples collected from any of the patients infected with JCV included in this study, which may suggest that oropharyngeal fluids are an unlikely source for JCV infection.

History and current concepts in the pathogenesis of PML

The JC virus (JCV), first described in 1971, is responsible for initiation of progressive multifocal leukoencephalopathy (PML), a disease characterized by demyelinating plaques and a classic triad of symptoms consisting of cognitive impairment, visual deficits, and motor dysfunction. To establish a diagnosis of PML, evidence of the presence of JCV DNA in pathologic tissue is necessary. The host range for productive infection of JCV is controlled by factors in the cell nucleus that bind to the viral promoter, initiating transcription of mRNA for the coordinated synthesis of viral proteins. Oligodendrocytes, astrocytes, and CD34+ and CD19+ cells of the immune system have the necessary binding proteins in sufficient concentration to allow lytic infection to occur. A strong link between JCV infection in cells of the immune system and cells of the nervous system points to the importance of the tissue origin of JCV latency, the bone marrow that harbors CD34+ cells. The emergence of PML in patients treated with natalizumab and other immune-altering agents supports this observation and provides new insights into the pathogenic mechanisms of JCV infection.

A rational approach to PML for the clinician

The first step in the management of progressive multifocal leukoencephalopathy (PML) is awareness of the disease. Patients vulnerable to PML are those with immunosuppression, either through their disease or use of immune-modulating therapy. In patients susceptible to PML who exhibit focal neurologic signs and symptoms, brain magnetic resonance imaging can detect the telltale PML brain lesions--subcortical white matter hyperintense areas on T2-weighted images and fluid-attenuated inversion recovery sequences and hypointensity on T1-weighted images, typically without enhancement. Demonstration of JC virus DNA by ultrasensitive polymerase chain reaction in cerebrospinal fluid is diagnostic for PML. Immune restoration whenever possible is the cornerstone of treatment. Highly active antiretroviral therapy has dramatically improved the prognosis for patients infected with human immunodeficiency virus. Alternatively, restoration of immunity is frequently attended by the immune reconstitution inflammatory syndrome which can be clinically devastating or even fatal. In the case of natalizumab-associated PML, withdrawal of therapy and prompt institution of plasmapheresis to desaturate target receptors provides the best chance for long-term survival.

Anti‐John Cunnigham virus antibody prevalence in multiple sclerosis patients: Baseline results of STRATIFY‐1

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. 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. 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). 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.

The high incidence of JC virus infection in urothelial carcinoma tissue in Taiwan

Human polyomaviruses, JC virus (JCV) and BK virus (BKV), usually remain latent in kidney and urothelial tissue after primary infection. Infection with human polyomavirus has still not been correlated conclusively with malignancy of kidney and urothelial tissue. The present study investigated further the possible relationship between JCV/BKV infection and urothelial carcinoma. Tissue samples were examined from 33 urothelial carcinomas and 5 renal cell carcinomas for JCV/BKV infection, using nested PCR with primers common to both JCV and BKV. The viral genotypes were further verified by endonuclease digestion and DNA sequencing following the PCR. In addition, immunohistochemistry and Western blotting were also performed to detect viral large tumor protein (LT) and the late capsid protein (VP1) in the tissue samples. The results from nested PCR showed that 90.1% (30/33) of the urothelial carcinomas samples and all of the renal cell carcinomas samples (5/5) were JCV DNA positive. Both archetypal and re-arranged JCV genotypes were detected. On the other hand, BKV DNA was detected in only one (3%) of the urothelial carcinoma tissue samples. The immunohistochemical results showed that 30% (10/33) of urothelial carcinoma tissues was stained positive for large tumor antigen (LT). However, the structural protein VP1 was not detectable in any of the tissue samples examined. The present study demonstrated that JCV is highly prevalent in urothelial carcinoma tissue as is the expression of large tumor antigen. Therefore, the findings support the hypothesis that JCV infection is associated with urothelial carcinoma.

Presence of JC Virus DNA in the Tumor Tissue and Normal Mucosa of Patients with Sporadic Colorectal Cancer (CRC) or with Positive Family History and Bethesda Criteria

JC virus (JCV) may infect the gastrointestinal tract in childhood, and, by encoding a gene for T-antigen (T Ag), can initiate chromosomal instability in epithelial cells. We looked for JCV DNA in the cancer tissue of patients with sporadic colorectal cancer (CRC, Group A) and with positive family history and Bethesda criteria (Group B). We hypothesized that the role of JCV may be different between these two groups. Fifty-six patients were randomly selected from our database, 30 in Group A and 26 in Group B. DNA was isolated from the tumor, normal mucosa, and plasma, and JCV DNA sequences were looked for with specific polymerase chain reaction (PCR) assays for T Ag primers. Immunohistochemistry for hMLH1, hMSH2, hMSH6, and PMS2 was performed on paraffin-embedded tissue. In Group A, T Ag was demonstrated in 6 (20.00%) and 3 (10.00%) of the tumors and adjacent normal mucosa, respectively (P = 0.094). In Group B, the corresponding observations were 10 (38.46%) and 6 (23.07%), respectively (P < 0.001). Immunohistochemistry for hMLH1, hMSH2, hMSH6, and PMS2 was performed in all of the Group A and B patients. All patients of Group A (100%) showed expression of these proteins, while only 19 patients of Group B did so (73.1%), P = 0.009. JCV T Ag DNA was found in 20, 28.5, and 42.1% of the tumors in Group A, Group B with negative staining for DNA repair genes, and Group B with a positive staining, respectively (NS). CRC patients with positive family history have a higher incidence of JCV T Ag, but this did not correlate with specific DNA repair gene mutations. We could not conclude that, on the background of genetic mutation in one of the DNA repair genes, JCV acts as the missing link in the chain of events leading to CRC.

Sequencing and Analysis of JC Virus DNA From Natalizumab-Treated PML Patients

Background. Progressive multifocal leukoencephalopathy (PML) in natalizumab-treated MS patients is linked to JC virus (JCV) infection. JCV sequence variation and rearrangements influence viral pathogenicity and tropism. To better understand PML development, we analyzed viral DNA sequences in blood, CSF and/or urine of natalizumab-treated PML patients.Methods. Using biofluid samples from 17 natalizumab-treated PML patients, we sequenced multiple isolates of the JCV noncoding control region (NCCR), VP1 capsid coding region, and the entire 5 kb viral genome.Results. Analysis of JCV from multiple biofluids revealed that individuals were infected with a single genotype. Across our patient cohort, multiple PML-associated NCCR rearrangements and VP1 mutations were present in CSF and blood, but absent from urine-derived virus. NCCR rearrangements occurred in CSF of 100% of our cohort. VP1 mutations were observed in blood or CSF in 81% of patients. Sequencing of complete JCV genomes demonstrated that NCCR rearrangements could occur without VP1 mutations, but VP1 mutations were not observed without NCCR rearrangement.Conclusions. These data confirm that JCV in natalizumab-PML patients is similar to that observed in other PML patient groups, multiple genotypes are associated with PML, individual patients appear to be infected with a single genotype, and PML-associated mutations arise in patients during PML development.

Rituximab-Associated Progressive Multifocal Leukoencephalopathy in Rheumatoid Arthritis

To describe the development of progressive multifocal leukoencephalopathy (PML) in patients with rheumatoid arthritis (RA) treated with rituximab. Case study. Clinical care for patients with rheumatologic diseases. Most were referred to academic centers for care after diagnosis (Washington University, St Louis, Missouri; Karolinska Insitute, Stockholm, Sweden; and Royal Melbourne Hospital, Melbourne, Australia) while one was cared for in a neurology practice in Dallas, Texas, with consultation by an academic neurovirologist from the University of Colorado in Denver. Four patients developing PML in the setting of rituximab therapy for RA. Rituximab therapy. Clinical and pathological observations. Four patients from an estimated population of 129 000 exposed to rituximab therapy for RA are reported in whom PML developed after administration of this drug. All were women older than 50 years, commonly with Sjögren syndrome and a history of treatment for joint disease ranging from 3 to 14 years. One case had no prior biologic and minimal immunosuppressive therapy. Progressive multifocal leukoencephalopathy presented as a progressive neurological disorder, with diagnosis confirmed by detection of JC virus DNA in the cerebrospinal fluid or brain biopsy specimen. Two patients died in less than 1 year from PML diagnosis, while 2 remain alive after treatment withdrawal. Magnetic resonance scans and tissue evaluation confirmed the frequent development of inflammatory PML during the course of the disease. These cases suggest an increased risk, about 1 case per 25 000 individuals, of PML in patients with RA being treated with rituximab. Inflammatory PML may occur in this setting even while CD20 counts remain low.

Assessment of JC virus DNA in blood and urine from natalizumab‐treated patients

Assessment of JC virus DNA in blood and urine from natalizumab‐treated patients

Progressive multifocal leukoencephalopathy and other forms of JC virus disease

Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease of the brain caused by the JC virus (JCV). PML usually occurs via reactivation of JCV when an immune system becomes compromised. A diagnosis of PML is normally made on the basis of distinguishing neurological features at presentation, characteristic brain MRI changes and the presence of JCV DNA in cerebrospinal fluid. PML has a 3 month mortality rate of 20-50%, so prompt intervention is essential. Currently, reconstitution of the immune system affords the best prognosis for this condition. When PML is first suspected, and where possible, immunosuppressant or immunomodulatory therapy should be suspended or reduced. If PML is associated with a protein therapy that has a long half-life the use of plasma exchange to accelerate the removal of the drug from the circulation may aid the restoration of immune system function. Rapid improvements in immune function, however, might lead to transient worsening of the disease. In this Review, we critically appraise the controversies surrounding JCV infection, and provide practical management guidelines for PML.

JC virus DNA in the peripheral blood of renal transplant patients: A 1‐year prospective follow‐up in France

There is little information on JC virus (JCV) infection in renal transplant patients. A long-term prospective follow-up study was conducted to assess the incidence of JCV DNA in the blood of 103 adult renal transplant patients enrolled prospectively between 1 January and 31 December 2006. Patients were monitored until April 2008. JCV DNA was quantified by a real-time polymerase chain reaction in whole blood samples collected regularly for at least 1 year post-transplant. JCV was detected in seven patients (6.8%) (31/1,487 whole blood samples) at a median time of 139 days post-transplant. The median JC virus load of the first positive DNA blood sample was 3.4 log(10) copies/ml (1.9-5.7 log(10) copies/ml). Induction therapy were either anti-CD25 monoclonal antibodies (n = 5) or antithymocyte globulins (n = 2). Post-transplant immunosuppressive treatment included steroids with tacrolimus/mycophenolate mofetil (MMF) (n = 2), or ciclosporin/MMF (n = 1), or belatacept/MMF (n = 4). Two patients were also treated with rituximab. All seven patients infected with JCV had other viral infections(s): BK virus (3), Epstein-Barr virus (2), Cytomegalovirus (1) or both BK virus and Epstein-Barr virus (1). Three patients had BKV-associated nephropathy and decoy cells shedding. JCV infection was not associated with acute rejection episodes or nephropathy, regardless of the virus load. No patient developed progressive multifocal leukoencephalopathy during follow-up. Thus the incidence of JCV infection in renal transplant patients was low and not associated with any specific clinical manifestations. JCV replication must still be diagnosed and differentiated from BK virus infection because of its non-aggressive course.

CD34+ progenitor cells mobilized by natalizumab are not a relevant reservoir for JC virus

Background: Progressive multifocal leukoencephalopathy (PML) is associated with natalizumab treatment in patients with multiple sclerosis (MS). It has been hypothesized that natalizumab mobilizes JC virus (JCV)-infected haematopoietic progenitor cells mediating viraemia and subsequently this disease. Objective: The objective of this study was to investigate peripheral haematopoietic progenitor cells for evidence of JCV DNA in MS patients treated with natalizumab. Methods: We assessed JCV and cytomegalovirus (CMV) DNA in magnetically separated CD34+ haematopoietic progenitor cells, peripheral blood mononuclear cells and plasma of 67 natalizumab-treated patients with MS and six PML patients. Results: Viral DNA was not detectable in CD34+ haematopoietic progenitor or peripheral blood mononuclear cells from any sample. Two plasma samples from patients with MS while undergoing natalizumab treatment were JCV-positive. In one case clinically manifest PML developed 8 months thereafter. Conclusions: Our findings do not support the hypothesis that natalizumab mobilizes JC virus-infected CD34+ cells from the bone marrow mediating JC viraemia. Notably, JC viraemia was detected in one patient with MS prior to developing clinical PML. This warrants further study.

JC virus persistence following progressive multifocal leukoencephalopathy in multiple sclerosis patients treated with natalizumab

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.

Assessment of JC virus DNA in blood and urine from natalizumab‐treated patients

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). 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). 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. 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.