In contrast, unprotected individuals would have a severe primary JCPyV infection, resulting in high levels of JCPyV capsid antibody and a future risk of PML. antibodies to BKPyV capsids are an immunological marker of protection against development of PML. Further studies are needed to define the mechanism. Keywords:Human immunodeficiency virus, progressive multifocal leukoencephalopathy, BK polyomavirus, JC polyomavirus, viral serology, cohort study == Introduction == Progressive multifocal leukoencephalopathy (PML) is a severe demyelinating disorder of the central nervous system caused by lytic infection of oligodendrocytes with JC polyomavirus (JCPyV) (Brew et al., 2010). PML occurs on a background of conditions associated with T-cell deficiencies (Viscidi RP and Shah KV, 2010). The majority of cases reported in the past 3 decades have occurred in human immunodeficiency virus (HIV) infected patients (Christensen et al., 2010). More recently, the immunomodulatory drug, natalizumab, and less commonly other monoclonal antibodies targeting immune modulatory antigens, have been associated with rare cases of PML (Rossi Thioridazine hydrochloride et al., 2014b). Exposure to JCPyV is common in the general population but PML is a rare disease, even among patients at risk due to an underlying predisposing condition. Thus, most individuals must be protected against the development of PML. Protection is likely to be immunologically mediated since all conditions associated with PML involve an immunodeficiency state. However, the nature of protective immunity is unknown. As a measure of exposure to Thioridazine hydrochloride JCPyV, antibodies to the VP1 major capsid protein have been shown to be a risk predictor for PML associated with natalizumab therapy (Gorelik et al., 2010), and we have shown that JCPyV seropositivity is associated with a nonsignificant trend for increased risk of PML in HIV-infected patients (Viscidi et al., 2011a). JCPyV is genetically related to a second human polyomavirus, BK polyomavirus (BKPyV) (Bennett et al., 2012). A negative correlation has been observed between levels of antibody to JCPyV and BKPyV capsids (Kean et al., 2009;Egli et al., 2009;Hamilton et al., 2000;Knowles et al., 2003). This unexplained phenomenological observation lead us to hypothesize that high levels of antibody to BKPyV will be a biomarker for protection against the development of PML. An opportunity to investigate pre-diagnostic markers is provided by Thioridazine hydrochloride prospective cohort studies of HIV infected subjects. We performed a nested case control study, conducted within the Multicenter AIDS Cohort Study (MACS), to examine whether antibody to BKPyV capsids are predictors of the subsequent diagnosis of PML. BKPyV genotypes are known to be serologically distinct and the major genotypes circulating in human populations are type 1 and 4; therefore, we tested for antibody to the capsid proteins of these two major serotypes (Pastrana et al., 2013). == Methods and Materials == == Study population == The MACS is an ongoing, prospective cohort study of HIV/AIDS in homosexual and bisexual men in the United States (Detels et al., 2012). Initial recruitment for the MACS began in April 1983 and up to January 2001, a total of 2,239 HIV-positive men have been enrolled and 584 HIV-negative men at study entry have become HIV-1 infected. Collectively, these men have contributed over 34,491 HIV-positive person-years of observation. The study was approved by local ethical review boards, and written informed consent was obtained from all participants. Demographic and clinical data and plasma samples were collected at follow-up visits every 6 months. A total of 32 patients were diagnosed with PML between 1985 and 2000. Seven patients were excluded from the present study because plasma samples were not available during the time interval of interest, 0.5 to 2 years prior to PML diagnosis. The time interval was selected based on sample availability and the desire to evaluate biomarkers in a clinically useful time frame. PML was diagnosed by histological examination of brain tissue in 7 of 25 (28%), a combination of radiographic and clinical signs in 8 (32%), radiology alone in 2 (8%), and clinical diagnosis alone in 8 (32%) patients. Each case was matched to three HIV seropositive participants, who did EIF4G1 not develop PML. Matching criteria were (1) CD4+ T-cell count (+50 cells per microliter) at the study entry visit to account for duration of HIV infection prior to entry into the study and (2) PML-free time from baseline. Cases for which PML was the AIDS defining event were further matched to HIV-positive controls on rate of CD4+ T-cell decline from baseline (+50cells per microliter per year). If PML was not the AIDS-defining event of a case, an HIV-positive control was selected who had an AIDS-defining event within one calendar year of the AIDS-defining event of the case and comparable PML-free time from AIDS. A total of 81 controls were identified. The 25 cases contributed a total of 66 plasma specimens with 16 (64%) cases providing three or more specimens..