**4. Nonmyelin autoAbs**

#### **4.1 DNA abzymes and antibody proteases**

Catalytic antibodies are endowed with a capacity to hydrolyze an antigenic substrate. (Paul S. 1998; Paul et al., 2005; Friboulet et al., 1999) This has moved antibodies to the level of physiological functionality by providing such antibodies with the ability to mediate direct catalytic and indirect cytotoxic effects on the targets. This property is buried in the Fab fragment of the immunoglobulin molecule. Antibody proteases (antibodies with proteolytic activity) were found in most autoimmune conditions, particularly in MS, accomplishing sequence-specific proteolytic cleavage of the myelin antigens and controlling the degradation of the myelin sheath. The levels of proteolytic activity of the antibody proteases revealed significant correlations with the clinical activity of the course over the disease, and thus with the disability of the patients. The most attractive point is that, in contrast to canonical proteases, for antibody proteases, there is an extra set of cleavage sites in the targeted autoantigens (for sequence-specific proteolytic cleavage) focused predominantly at the immunodominant sites. (Gabibov et al., 2002)

Physiological interaction of autoAb with a living cell is mediated by Fc fragment (Wilkinson et al., 2001). DNA-abzymes are able to realize both cytotoxicity mechanisms-complementdependent cell lysis and K-cell-mediated lysis. (Fishelson et al., 2001) Fab fragment is not involved in these reactions. This fundamentally distinguishes Ab-mediated classical and DNA-mediated mechanisms of cytotoxicity due to direct involvement of Fab fragment into the catalytic attack on the target cell genome. Catalytic and cytotoxic activities of DNA-

Proteolipid protein (PLP) is the most abundant protein in CNS compact myelin (about 50%), highly hydrophobic and evolutionarily conserved across species. There are two main transcripts, the full-length 276 amino acid isoform and DM-20, an isoform that lacks 35 amino acids and is mainly expressed in brain and spinal cord prior to myelination, but also in peripheral lymphoid organs such as the thymus, where full-length PLP is barely found. Interestingly, the major encephalittogenic and immunodominant PLP peptide (139-154) is contained in full-length PLP but not in DM-20. This observation is thought to account for the encephalittogenicity and immunodominance of the PLP (139-154) peptide, since it is essentially not available for thymic negative selection and consequently a high precursor frequency of PLP (139-154)-specific T cells has been observed even in naive unprimed

Myelin oligodendrocyte glycoprotein, a 218 amino acid trancemembrane glycoprotein of the IG superfamily, is much less abundant (0.01 - 0.05%) than MBP and PLP, and also different from the two major myelin proteins in being located not in compact myelin but exposed on the outermost surface of the oligodendrocyte membrane. Because of this "strategic" location, it is directly accessible to antibodies and believed to be particularly relevant as a target for both cellular and humoral immune responses in MS. MOG, is expressed relatively late during myelination and is only found in the brain/spinal cord and the retina but not in peripheral nerve. Furthermore, MOG expression is either completely or almost completely lacking in peripheral lymphoid tissues, although MOG transcripts have been seen in nonhuman primate peripheral nerve and a few samples of human tonsils and thymus.

Catalytic antibodies are endowed with a capacity to hydrolyze an antigenic substrate. (Paul S. 1998; Paul et al., 2005; Friboulet et al., 1999) This has moved antibodies to the level of physiological functionality by providing such antibodies with the ability to mediate direct catalytic and indirect cytotoxic effects on the targets. This property is buried in the Fab fragment of the immunoglobulin molecule. Antibody proteases (antibodies with proteolytic activity) were found in most autoimmune conditions, particularly in MS, accomplishing sequence-specific proteolytic cleavage of the myelin antigens and controlling the degradation of the myelin sheath. The levels of proteolytic activity of the antibody proteases revealed significant correlations with the clinical activity of the course over the disease, and thus with the disability of the patients. The most attractive point is that, in contrast to canonical proteases, for antibody proteases, there is an extra set of cleavage sites in the targeted autoantigens (for sequence-specific proteolytic cleavage) focused predominantly at

Physiological interaction of autoAb with a living cell is mediated by Fc fragment (Wilkinson et al., 2001). DNA-abzymes are able to realize both cytotoxicity mechanisms-complementdependent cell lysis and K-cell-mediated lysis. (Fishelson et al., 2001) Fab fragment is not involved in these reactions. This fundamentally distinguishes Ab-mediated classical and DNA-mediated mechanisms of cytotoxicity due to direct involvement of Fab fragment into the catalytic attack on the target cell genome. Catalytic and cytotoxic activities of DNA-

**3.2 Proteolipid protein** 

**3.3 Myelin oligodendrocyte glycoprotein** 

**4.1 DNA abzymes and antibody proteases** 

the immunodominant sites. (Gabibov et al., 2002)

**4. Nonmyelin autoAbs** 

animals.

abzymes are closely related. This allows to assume on a hypothesis on a new mechanism of the contribution of autoAbs into the pathogenesis of autoimmune disorders. Such mechanism acts independently of complement and cytotoxic T cells; it requires a catalytically active Fab fragment but ignores Fc fragment whose structure is deprived of even buried resources providing direct cytotoxic effect. (Ponomarenko et al., 2000)

Two different mechanisms of DNA-abzymes cytotoxic potential utilization are establishedby means of direct cytotoxic effect on a target cell involving the catalytically active Fab fragment and by means of apoptosis due to high affinity of DNA-binding autoAbs for membrane receptors providing the cell with features of possible target. Cross-reactive with such such cells, DNA-abzymes can provoke their degradation resulting in the development of different syndromal manifestations: lupus nephritis in glomerule endothelium crossreactivity, articular syndrome in synovial cross-reactivity, extracardial manifestations and MCS progression in CM cross-reactivity. (Rekvig et al., February 2004; Raz et al., 1993)

Cross-reactivity of DNA-binding autoAbs with glutamate receptors accumulating at neural cells is of a special interest. Such autoAbs with catalytic and cytotoxic activities can initiate neural cells apoptosis providing the possibility for the development of CNS autoimmune degenerative disorders (Kotzin et al., 2001). The penetration of DNA-binding autoAbs (and DNA-abzymes) into a cell results in activation of cytotoxicity mechanisms, apoptosis induction, etc.

#### **4.2 Molecular mechanisms of the involvement of DNA-abzymes in the development of different autoimmunity conditions and pathogenesis of autoimmune disorders**

One antigen may generate up to 102-104 different antibody molecules, a number that may further increase by somatic mutagenesis. Therefore, it seems feasible that different DNAbinding and other non-catalytic antibodies, as well as antibodies with catalytic activities can be synthesized in the course of immune response, either directed against the substrate or as antiidiotypic antibodies to enzymes hydrolyzing nucleic substrates. It is certainly difficult to predict the clinical significance of these catalytic activities, but it is likely that they modify the pathogenesis or clinical process of these autoimmune diseases.

The interest to catalytic (i.e., DNA-hydrolyzing activity) of the autoantibodies is kept on growing up and is strongly supported by the new data including those illustrating cytotoxic activities of the biocatalysts and evidence that abzyme-mediated cytotoxic effects observed in human SLE (system lupus erythematosis) and mouse SLE-like syndromes are caspasedependent and thus apoptosis-related.
