John Cunningham virus (JCPyV), a member of the Polyomaviridae family, consists of a circular double-stranded DNA genome composed of an early and a late regions physically separated by the non-coding control region (NCCR). The DNA sequence of the NCCR distinguishes two forms of JCPyV, the designated archetype and the prototype which results from a rearrangement of the archetype sequence. JCPyV is the etiological agent of the progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the brain. PML was originally a rare complication of hematological malignancies; however, the incidence of PML has increased during the HIV epidemic in the 1980s. Recently, PML cases are increasing among patients treated with monoclonal antibodies. Although JCPyV infection is widespread in the population, the mode of transmission is not yet well defined. Virus might enter the mouth or nose and presumably spread by the hematogenous route from the primary site of infection to secondary sites such as kidneys, lymphoid tissues and brain to establish focal areas of infection or persistence. When alteration of the immune system occurs, viral infection emerges. Although rearrangement of NCCR of archetype JCPyV is thought to be an important event in the pathogenesis of PML, little is known regarding what induces this rearrangement. To date, the cell culture systems to propagate JCPyV archetype have been very limited in their availability and robustness. Prior to this study, it has been demonstrated that JCPyV archetype DNA replicates in COS-7 cells expressing SV40 TAg and HIV-1 Tat. Based on these observations, the present study is addressed: 1) to produce an in vitro model in COS-7 and SVGp12 cells to study JCPyV DNA replication, 2) to analyze NCCR rearrangements during the viral cycle, 3) to define NCCR rearrangements as possible viral biomarkers for an early PML diagnosis.
It is important to be able to grow the CY archetype form of JCPyV to understand the natural history of infection and develop means to prevent transmission and reactivation. An in vitro model could be useful in order to study the possibility of JCPyV NCCR rearrangement in sites of viral persistence, such as the tubular epithelia of kidneys. It is also important to finally understand the nature of JCPyV infection in cell types supporting productive viral infection, and which kind of cells could be implicated in the process of JCPyV rearrangement. This is the missing piece in the pathogenesis puzzle of PML: in fact, the importance of NCCR rearrangements in the onset of PML has been demonstrated by the discovery of highly rearranged sequences in patients with this condition. However, it is not yet understood where and when these rearrangements occur, if during primary infection or during the persistence of the virus in the host.
The main aim of the study is: 1) to produce an in vitro model in COS-7 and SVGp12 cells to study JCPyV DNA replication, 2) to analyze the NCCR rearrangements during the viral cycle and 3) to define NCCR rearrangements which could be translated in clinical practice as possible viral biomarkers for an early PML diagnosis.
Although the hypothesis that immune-deficiency, either diseases or therapeutic drug-related, is a predisposing factor for PML, there are a number of unsolved issues including the pathogenetic mechanisms related to the interaction of JCV infection/reactivation with the host and the absence of a stable ¿in vitro¿ cellular model to verify the factors influencing the latency and the reactivation state of the virus. Moreover, as the etiological agent of PML, JCV is widely diffused in the human healthy world population and as there is no known treatment or cure for PML, the precise understanding of the molecular mechanisms and of the viral/host factors influencing the reactivation of JCV is needed.
Therefore, it could be helpful to define new biomarkers that will lead to more precise diagnosis, to more timely therapeutic intervention, to optimization of the use of healthcare resources and reduction of the direct and indirect costs of the disease. Possible new insights obtained with the results of this research, may open new perspectives in the pathogenesis of PML.