B CELL RESPONSES IN MULTIPLE SCLEROSIS

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Increased IgG and oligoclonal bands (OGBs) are found exclusively in the CSF of patients with infectious and inflammatory diseases of the CNS, particularly multiple sclerosis (MS). Although their specificity in MS is unknown, in infectious diseases of the CNS, OGBs are specific for the agent that causes disease, thus providing a rationale for our hypothesis that OGBs in MS brain and CSF are directed against the antigen that causes disease. To distinguish between a specific versus a random humoral response, we sequenced IgG heavy (V/H) and light (V/L) chain variable regions expressed in multiple acute MS plaques and found a restricted response consisting primarily of V/H4 germline genes. Some V/H sequences were over-represented, clonally expanded, and displayed a non-random accumulation of somatic mutations, features indicative of an antigen driven B cell response. A parallel analysis of subacute sclerosing panencephalitis (SSPE) brain also revealed over-represented and somatically mutated V/H and V/L sequences. We co-expressed these sequences in mammalian expression vectors and showed that the recombinant IgG was specific for measles virus (the cause of SSPE) in infected tissue culture cells. We will exploit our success with recombinant measles virus specific antibody to identify measles antigen in SSPE brain. Information accumulated from SSPE will be used to synthesize recombinant antibodies from candidate MS sequences to demonstrate their immunologic specificity, for either protein or carbohydrate antigens. Once specificity is established, we will screen phage display cDNA libraries from MS brain to identify and characterize cDNAs whose protein products react with MS recombinant antibodies or with IgG purified from acute MS plaques. We will also continue to sequence V/H regions from acute MS plaques to determine if restricted use of specific V/H4 or other family germlines is characteristic of MS. We are well-prepared to conduct these studies because we have: (a) pathologically verified acute MS brains and non-MS neurologic disease brains; (b) a demonstrated ability to identify disease- relevant IgG sequences from human brain, and to use these sequences to generate recombinant IgG specific for the agent that causes disease; (c) the expertise in molecular biology to construct and screen complex cDNA libraries; and (d) CSF containing OGBs from patients with MS and other CNS inflammatory diseases. Identification of an MS-specific antigen will have wide application, not only for early definitive diagnosis, but also for developing strategies for modulation, if not prevention, of disease.