Influence of JC virus coding region genotype on risk of multiple sclerosis and progressive multifocal leukoencephalopathy.

Two features of the biology of JC virus make it a particularly suitable candidate for an agent in MS-like disease: its neurotropic capability targeting glial cells as evidenced in progressive multifocal leukoencephalopathy lesions, and its capacity for latency and persistence as illustrated by its behaviour in the kidney. JC virus is chronically or intermittently excreted in the urine by some 40% of the population. The existence of JC virus in multiple coding-region genotypes provides a unique approach to the study of JC virus-induced neurological disease. We have previously shown that a genotype originating in Asia but also present in Europe and the US, called Type 2B, is more frequently found in PML brain than expected based on its prevalence in urine samples from a control population. In contrast, we find that the excretion of JCV in MS patients is similar in both genotype and frequency to that of control individuals, and appears to be regulated by factors unrelated to those that control CNS disease activity.

JC virusType 2: definition of subtypes based on DNA sequence analysis of ten complete genomes.

Five major genotypes of JC virus (JCV) have been defined based on nucleotide differences in the VP1 gene of the DNA sequence. These types are probably a result of virus evolution in geographically isolated population groups. One of the first genotypes identified, Type 2, was found to represent strains of Asian origin. In order to further define the spectrum within Type 2 strains, the entire 5.1 kb genome of nine urinary strains of JCV was amplified by PCR with one pair of primers. These urine samples were obtained in the USA (California and New Mexico) from three European Americans, three Native Americans, two African Americans and one Hispanic American. The complete genome of an Asian JCV strain (Tokyo-1) isolated from progressive multifocal leukoencephalopathy (PML) brain in Japan was also sequenced. Here, we report the analysis of these ten DNA sequences and their deduced protein translations. Two phylogenetically distinct subtypes of Type 2 were found, 2A and 2B, which differ from each other by 0.8-1.1% of the coding region sequence. A 215 bp product amplified with primers in the VP1 gene contains enough sequence information to distinguish the major types and subtypes of JCV and is suitable for application in viral epidemiological studies. The investigation of these genomic variations is of special interest because JCV Type 2 strains are found at a significantly higher frequency in brain tissue of patients with PML than would be predicted from their excretion in a control population.

JC virus type 2B is found more frequently in brain tissue of progressive multifocal leukoencephalopathy patients than in urine from controls.

OBJECTIVES: Previous studies have shown that strains of human polyomavirus JC (JCV) of Asian origin (type 2) are much more highly represented in progressive multifocal leukoencephalopathy (PML) brain than would be expected from their frequency of excretion in urine samples of a comparable control group. The present studies were designed to test whether one subtype of type 2 was preferentially elevated. STUDY DESIGN/METHODS: The statistical relation between JCV subtypes represented in PML brain tissue from 51 probands and those in urine samples from 115 control individuals was examined. RESULTS: The proportion of the JCV subtype 2B in PML brain (36%) was highly significantly increased relative to its occurrence in control urine samples (5.9%; P < .001). Type 1 and its subtypes were not different in the PML brain and control urine cohorts. The number of type 4 strains in PML brains was reduced, although the difference did not reach statistical significance (P = .08). CONCLUSIONS: The results predict that the human immunodeficiency virus (HIV)-positive individuals at highest risk of PML infection are those carrying the JCV genotype known as type 2B. Prospective studies will be required to confirm this finding.

Comparative evaluations of neuroperformance and clinical outcome assessments in chronic progressive multiple sclerosis: I. Reliability, validity and sensitivity to disease progression. Multiple Sclerosis Study Group.

There remains controversy regarding the most sensitive and valid outcome assessments to use in multiple sclerosis (MS) clinical trials. A double blind, placebo controlled, parallel group multicenter clinical trial to evaluate the clinical efficacy of cyclosporine A in chronic progressive MS incorporated several major clinical and performance outcome assessment modalities and a large sample size, both of which provide a unique opportunity to explore the relationship among MS disease status and the various outcome measures over time. The measures included a structured neurological examination, the Kurtzke Functional System scales and Expanded Disability Status Score, and the Incapacity Status Scale from the MS Minimal Record of Disability, the Harvard Ambulation Index, and neuroperformance testing. A test-retest reliability index, principal component analyses and a signal-to-noise ratio metric were used to comparatively evaluate the reliability, validity and sensitivity to disease progression of the various outcome assessments. The goal was to provide a rational basis for selection of behavioral outcome assessments in future MS clinical trials by identifying the primary dimensions of MS measured by the candidate outcome assessments and providing an objective basis for selecting tests that are most sensitive to MS disease and its progression over a two year trial period. We conclude that the components of the major clinical and performance measures show excellent reliability and cross validation. Principal component analyses of all outcome assessments yielded six primary underlying factors for describing disease status in chronic progressive MS that included lower extremity/pyramidal dysfunction, cerebellar/brainstem and upper extremity dysfunction, somatosensory dysfunction, visual dysfunction, mental or intellectual dysfunction and bowel/bladder problems. Signal-to-noise ratios indicated that upper and lower extremity composites of neuroperformance test items provided the most sensitive indicators of MS disease progression in the placebo group over the 2 year trial period.

Characterization of JC virus DNA amplified from urine of chronic progressive multiple sclerosis patients.

Thirty-seven chronic progressive multiple sclerosis (MS) patients, 20 of whom were taking cyclosporine, were examined for excretion of JC virus (JCV) in the urine. Polymerase chain reaction (PCR) amplification of DNA in urinary cell extracts detected JCV in 30% of the MS urines. In the cyclosporine treated group four of 20 (20%) excreted JCV, whereas in the untreated group seven of 17 (41%) excreted JCV. Thus, cyclosporine treatment did not enhance urinary excretion of the virus. A control group consisting of an unselected series of 89 patients donating urine in a general medical clinic and 16 healthy volunteers showed 41% with detectable urinary JCV. Thirty-three percent of the control females excreted JCV (18/54), as did 49% of the control males (25/51). Although the percentage of MS patients excreting detectable virus was not increased compared to the control group, the presence of JCV in the urine provides a convenient source of the virus for further characterization. Genotyping of DNA fragments amplified from the VP1 region indicates mainly the presence of JCV Type 1 in these chronic progressive MS patients. This is also the type that predominates in the control group. An apparent recombinant between Type 1 and Type 3 (African) within the VP1 region, tentatively designated Type 1/3 (or Type 4), was found in both the MS group and the controls. A larger series of MS patients that includes relapsing/remitting disease will be required to determine whether the genotype profile of JCV excreted in the urine of MS patients differs significantly from controls.

Quantitation of intrathecal measles virus IgG antibody synthesis rate: subacute sclerosing panencephalitis and multiple sclerosis.

A method for quantitating specific anti-viral antibodies in serum and cerebrospinal fluid (CSF) is established using enzyme-linked immunosorbent assay (ELISA). Quantitated antibody levels are used to determine intrathecal specific IgG synthesis rate for the particular antibody. Measles virus was used as a model for validating this quantitative technique: a mutated form of measles virus is a cause of subacute sclerosing panencephalitis (SSPE) and there is a possibility that measles virus is related to the cause of multiple sclerosis (MS). Matched serum and CSF samples were assayed. Concentration of anti-measles IgG was determined and intrathecal measles-specific IgG synthesis rate was calculated. For the SSPE samples, measles-specific IgG synthesis rate was elevated and comprised > 20% of the total intrathecal IgG synthesis rate; these results are consistent with the literature. The ELISA method can be performed routinely, providing a quick, simple, reproducible means of quantitating specific antibody concentrations, with sensitivity greater than 1 nanogram per milliliter. With this method, quantitation of IgG antibodies to any other viral antigen can be reliably and precisely determined.

Cryptococcal meningitis/acquired immune deficiency syndrome presenting as a transient neurologic deficit.

A previously healthy man presented with a transient neurologic deficit and neck pain. Lumbar puncture revealed cryptococcal meningitis. He was subsequently found to have acquired immune deficiency syndrome (AIDS). Transient neurologic deficit is an uncommon initial manifestation of cryptococcal meningitis. We suggest AIDS-related infections be considered in the differential diagnosis of transient neurologic deficits.

Quantitative multiple sclerosis plaque assessment with magnetic resonance imaging. Its correlation with clinical parameters, evoked potentials, and intra-blood-brain barrier IgG synthesis.

Magnetic resonance imaging (MRI) of the cerebrum, cerebellum, brain stem, and upper cervical cord was performed in 62 individuals with clinically definite chronic, progressive multiple sclerosis (MS). The total area of MRI-demonstrated lesions was measured from film enlargements for each region using an interactive image analysis system. While the MRI was abnormal in 60 (97%) of 62 patients, the visual-evoked potentials in 51 (85%) of 60 patients, the brain stem auditory-evoked potentials (BAEPs) in 24 (46%) of 52 patients, and the somatosensory-evoked potentials (SSEPs) in 45 (89%) of 54 patients, an abnormal intra-blood-brain barrier (BBB) IgG synthesis rate, IgG oligoclonal bands, or both were found in all 62 patients. The total area of MRI abnormality in the cerebrum was significantly correlated only with the intra-BBB IgG synthesis rate, abnormal visual-evoked potentials, impaired performance on the Symbol Digit Modalities Test (SDMT), and one test of standing duration in the quantitative examination of neurologic function (QENF). The brain stem lesion area correlated with the Kurtzke expanded disability status scale and brain stem functional systems score, the ambulation index, abnormal BAEPs, and impaired performance on the SDMT as well as multiple tests of upper and lower extremity function in the QENF. The cerebellar lesion area correlated with impaired performance on the SDMT and primarily upper extremity testing in the QENF.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiple sclerosis intra-blood-brain-barrier IgG synthesis: effect of pulse intravenous and intrathecal corticosteroids.

Nine severely disabled clinically definite chronic progressive multiple sclerosis (MS) patients who had at least one determination of intra-blood-brain-barrier (BBB) IgG synthesis rate of greater than 7 mg/day (upper limit of normal = 3.3) participated in this study. Seven patients were given 1 gram of methylprednisolone sodium succinate (MP) by intravenous infusion over 30 minutes once a day for 3 days. Statistically significant (p less than .05) reduction in intra-BBB IgG synthesis (mg/day) was seen in 4/7 patients, but in only 2 were normal levels of synthesis rate (less than 3.3 mg/day) attained. Rebound of IgG synthesis to premedication rates occurred within 30 days in 2/4 patients. There was no change in intensity or pattern of cerebrospinal fluid (CSF) oligoclonal IgG bands by isoelectric focusing, immunofixation, and silver staining. A subsequent course of intrathecal methylprednisolone acetate (MPA) (80 mg twice a week for 5 weeks) was given to 5 of the 7 patients and to 2 additional patients not previously treated. In spite of signs of subarachnoid inflammation, a statistically significant depression of intra-BB synthesis, which far exceeded that from the pulse treatment occurred in all 7, including the 2 patients whose intra-BBB IgG synthesis rates were previously resistant to pulse steroid administration. Normal levels of synthesis were rapidly reached in 4/7 patients; however, an IgG synthesis rebound occurred in 3/7 patients which was just as rapid. One out of 7 patients showed a temporary reduction in the number of cathodic IgG oligoclonal bands in the CSF. Two patients required discontinuation of treatment due to aseptic meningitis in one and progressive weakness in the other. Clinically, these severely afflicted patients with fixed deficits remained unchanged with either treatment protocol. While MPA and ACTH have similar initial effect on the central nervous systems (CNS) inflammatory response in MS, the well documented risk of serious adversities with MPA prohibit its clinical use in MS in its present form.

The long march of the cerebrospinal fluid profile indicative of clinical definite multiple sclerosis; and still marching.

Much progress has been made, especially in the last two decades, in laboratory aids to diagnosis and to follow the course of patients with multiple sclerosis (MS). The cerebrospinal fluid (CSF) profile indicative of MS, though not pathognomonic of MS, is present in almost every case of clinical definite MS in a chronic progressive phase (probably also true for early MS). The cardinal aspect of the profile is intra-blood-brain barrier (BBB) IgG synthesis which can be qualitatively detected by determining unique CSF oligoclonal IgG bands and quantitated by rate formula, mg/day. We believe that intra-BBB IgG synthesis is caused by a persistent antigen, most likely a virus, possibly measles. A number of issues about the profile are proposed and opportunities are presented to resolve them.

Copolymer 1 as therapy for multiple sclerosis: the cons.

One of the hallmarks of multiple sclerosis (MS) is intra-blood-brain-barrier (BBB) IgG synthesis, a byproduct of plasma cells located in and around active inflammatory demyelinating plaques. The rate of IgG synthesis can be measured by plugging CSF and blood IgG and albumin concentrations into our equation. When done in conjunction with CSF and serum analyses for IgG oligoclonal bands, 99% of definite MS patients demonstrate intra-BBB IgG synthesis. At autopsy the pathologic criterion of an inactive plaque of demyelination is absence of inflammatory cells. Hence, we propose that modulation downward or eradication of intra-BBB IgG synthesis (ie, a manifestation of reduced white matter inflammation) in a living patient is a reasonable therapeutic criterion and goal of MS therapy. In a preliminary trial of five severely disabled MS patients, we evaluated the effects of copolymer 1 (COP-1) in daily intramuscular doses of 20 mg (2 patients) and twice daily subcutaneous doses of 15 mg (3 patients) on clinical parameters and on intra-BBB IgG synthesis over a 2-month study period. The results of this trial showed no beneficial effect on neurologic function or on inflammatory demyelination, as assessed by monitoring of intra-BBB IgG synthesis.

The basis of intra-blood-brain-barrier IgG synthesis.

We hold that the intra-blood-brain-barrier (BBB) IgG synthesis (SYN) rate can be quantitated reliably and validly. Although several formulas distinguish patients with multiple sclerosis from normal controls equally well, only the SYN rate formula has been validated in humans using isotopic tracer techniques. Our formula for the IgG SYN rate is reducible to Reiber's formula; therefore, both can be used to quantify the IgG SYN rate in a manner not possible using the IgG index. Although our SYN rate formula has been validated for a modest range of BBB abnormalities (cerebrospinal fluid/serum albumin ratios), there is evidence to suggest that it may be used even in patients having severe BBB damage. We question the acceptance of unique cerebrospinal fluid IgG bands as indisputable evidence of intra-BBB IgG SYN in the presence of a modest to severely damaged BBB. Finally, the utility of quantitation and detection of intra-BBB IgG SYN by standardized methods in a group of asymptomatic, normal individuals compared with a group of patients with clinical definite multiple sclerosis is presented.

The current status of multiple sclerosis intra-blood-brain-barrier IgG synthesis.

The status of the central nervous system as an immunologic organ synthesizing IgG in several neurological diseases has been established convincingly in the past two decades. The measurement of intra-BBB IgG synthesis has been accomplished. It has been demonstrated that this synthesis is a potential marker for an abnormal immunologic process within the CNS which if manipulated may be a quantifiable aspect of disease modulation. Modulation of intra-BBB IgG synthesis is feasible, but there is no evidence yet that modulation of this synthesis is an index of effective disease control in MS. Only ACTH and corticosteroids lead to a significant and lasting depression of intra-BBB IgG synthesis, even though clonal eradication is not achieved. Based on our immunopharmacologic studies in MS we have proposed an immunokinetic model. The qualitative analysis of the specificity of IgG synthesized intra-BBB in MS has not led to the etiology of MS, but utilization of quantitative methods may. Questions raised by this review with suggested experiments to advance further our understanding of intra-BBB IgG synthesis as it relates to the etiopathogenesis of MS are included.

Difference in effect of single immunosuppressive agents (cyclophosphamide, CCNU, 5-FU) on peripheral blood immune cell parameters and central nervous system immunoglobulin synthesis rate in patients with multiple sclerosis.

Cyclophosphamide (CY), 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU) and 5-fluorouracil (5-FU) were given in single course schedules to chronic progressive multiple sclerosis (MS) patients clinically stable for 6 months. The following peripheral immune cellular parameters were measured before, during and after each drug administration: white blood count (WBC), polymorphonuclear count (PMN), lymphocyte count, percentage of T cells, T cell response to phytohaemagglutinin (PHA), percentage of B cells, percentage of cells bearing receptors for the Fc portion of immunoglobulin (% FcR cells), killer (K) cell activity defined by antibody-dependent cellular cytotoxicity (ADCC), and natural killer (NK) cell activity. Central nervous system (CNS) immunoglobulin G (IgG) synthesis was also measured. The patients were followed carefully by both quantitative and qualitative methods for any change in their neurologic condition. Selective reduction in NK activity was observed with CY and 5-FU while no significant alteration was seen in %FcR cells and K activity. CY differed from 5-FU in reducing lymphocyte count and B cell percentage while 5-FU decreased the percentage of T cells. CCNU, but not the other drugs, reduced T cell proliferative response to PHA. In addition, CCNU, which is known to penetrate well into the nervous system, caused a modest reduction in CNS IgG synthesis, while 5-FU had an uncertain effect. Clinically the patients were unchanged or continued to progress in their disability. The results suggest an independence of the CNS immune from the systemic immune system in MS in response to many immunosuppressive drugs.