Production of a recombinant hybrid molecule of cholera toxin-B-subunit andproteolipid-protein-peptide for the treatment of experimental encephalomyelitis

Mucosal administration of experimental autoimmune encephalomyelitis (EAE)-s pecific autoantigens can reduce the onset of disease. To examine whether ch olera toxin-B-subunit (CTB)-conjugated EAE-specific T-cell epitope can redu ce development of the autoimmune disease in mice, we produced a recombinant hybrid molecule of CTB fusion protein linked with proteolipid-protein (PLP )-peptide139-151(C140S) at levels up to 0.1 gram per liter culture media in Bacillus brevis as a secretion-expression system. Amino acid sequencing an d GM1-receptor binding assay showed that this expression system produced a uniformed recombinant hybrid protein. EAE was induced in SJL/J mice by syst emic administration with the PLP-peptide. When nasally immunized 5 times wi th 70 mug rCTB PLP-peptide hybrid protein, mice showed a significantly supp ressed development of ongoing EAE and an inhibition of both the PLP-peptide -specific delayed-type hypersensitivity (DTH) responses and leukocyte infil tration into the spinal cord. In contrast, all mice given the PLP-peptide a lone or the PLP-peptide with the free form of CTB did not suppress the deve lopment of EAE and DTH responses. These results suggest that nasal treatmen t with the recombinant B. brevis-derived hybrid protein of CTB and autoanti gen peptide could prove useful in the control of multiple sclerosis. (C) 20 01 John Wiley & Sons, Inc. Biotechnol Bioeng 74: 62-69, 2001.