CNS Antigen-Specific B Cell and Antibody Assays in Multiple Sclerosis

Abstract

Embodiments of this invention include methods for detecting in vitro the presence in peripheral blood mononuclear cells (PBMCs), and in serum or plasma, of antibodies reactive to and of lymphocytes that are responsive to CNS antigens associated with Multiple Sclerosis (MS). These CNS antigens include, but are not limited to whole brain lysate and the myelin antigens myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), MOG peptides (MOGps), proteolipid protein (PLP), and PLP peptides (PLPps). PBMCs obtained from patients with active MS produce antibodies specific for CNS antigen, and causes T-lymphocytes to produce T-lymphocyte-specific cytokines, including interferon gamma (IFN-γ), interleukin-2 (IL-2), or interleukin-17 (IL-17). In contrast, PBMCs from subjects without MS do not produce such responses. The magnitude of the CNS antigen-specific response in patients with definite MS or in patients with CIS or RIS can indicate the likelihood that a given patient will develop a relapse and can indicate the responsiveness to and the success of immune modulatory treatment.

1 . A method for detecting a cell that produces an antibody against a CNS-specific antigen, comprising the steps:
 a) providing a cell culture well having a surface and a cell culture medium therein;   b) attaching a CNS-specific antigen to said surface;   c) introducing a sample of peripheral blood mononuclear cells (PBMCs), CNS cells, or cell liquor from a human being into said cell culture medium;   d) permitting said PBMCs to produce an antibody against said CNS-specific antigen; and   e) detecting the presence of said antibody using an anti-antibody specific reagent.   
     
     
         2 . The method of  claim 1 , said step of detection being carried out using a method selected from the group consisting of enzyme linked immune spot assay (ELISPOT), enzyme-linked immunoassay (ELISA), bead array, or protein array. 
     
     
         3 . The method of  claim 1 , said CNS-specific antigen selected from the group consisting of whole-brain lysate, human myelin antigens, human proteolipid protein (hPLP), human myelin basic protein (hMBP), myelin oligodendrocyte glycoprotein (hMOG), neuronal antigen, a peptide from hMOG (hMOGp) containing a sequence of hMOG, and a MOG/PLP fusion protein. 
     
     
         4 . The method of  claim 1 , said method performed with PBMCs or cells isolated from the CNS. 
     
     
         5 . The method of  claim 1 , where said CNS-specific hMOG antigen is SEQ ID NO.1. 
     
     
         6 . The method of  claim 1 , where said CNS-specific MOGp antigen is human MOG 35-55 (SEQ ID NO.2). 
     
     
         7 . The method of  claim 1 , where said CNS-specific antigen MBP is MBP isoform 3, 21.5 kDa (SEQ ID NO.3). 
     
     
         8 . The method of  claim 1 , where said CNS-specific proteolipid protein antigen is ΔPLP4 (SEQ ID NO.4). 
     
     
         9 . The method of  claim 1 , where said CNS-specific MOG/PLP antigen is fusion protein MP4 (SEQ ID NO.6). 
     
     
         10 . A method for treating a patient having multiple sclerosis (MS), comprising the steps:
 a) providing a cell culture well having a surface and a cell culture medium therein;   b) attaching a CNS-specific antigen to said surface;   c) introducing a sample of peripheral blood mononuclear cells (PBMCs), CNS cells, or cell liquor from a human being into said cell culture medium;   d) permitting said PBMCs to produce an antibody against said CNS-specific antigen;   e) detecting the presence of said antibody using an anti-antibody specific reagent; and   f) if said antibody is detected in step e), administering to said patient an immune modulating agent.   
     
     
         11 . The method of  claim 10 , wherein said immune modulating agent is a B-cell depleting agent. 
     
     
         12 . The method of  claim 11 , wherein said B-cell depleting agent is an anti-CD20 antibody. 
     
     
         13 . The method of  claim 10 , further comprising administering to said patient a therapeutic agent selected from the group consisting of glatiramer acetate, interferon beta-1a, interferon beta-1b, mitoxantrone, natalizumab, and FTY720 fingolimod.