## 2 Results and Discussion
### Inhibitory Activity of MOG APIs in the EAE. The rat MOG\({}_{35-55}\) (rMOG\({}_{35-55}\)) peptide (35 \(\upmu\)g) induces a chronic disease with a peak clinical score by day 16 after the immunization (Figure 4). Co-injection of linear APL analogues (350 \(\upmu\)g of the [Ala\({}^{41,46}\)]MOG\({}_{35-55}\) or [Ala\({}^{41}\)]MOG\({}_{35-55}\)) with the encephalitogenic peptide MOG\({}_{35-55}\) (35 \(\upmu\)g) at the time of EAE induction (time zero) significantly reduced disease symptoms and incidence, with the linear [Ala\({}^{41}\)]MOG\({}_{35-55}\) peptide completely preventing the onset of the disease, as compared to PBS (phosphate buffered saline) control (Figure 4). The linear analogue with TyrPOMe (methyl ether instead of hydroxyl group) at position 40 [TyrOMe\({}^{40}\)]MOG\({}_{35-55}\) and the cyclic analogues cyclo(46-55)MOG\({}_{35-55}\) and cyclo(41-55)MOG\({}_{35-55}\) did not significantly lower the clinical symptoms of EAE (Figure 4). This study was an initial attempt to determine which of the designed linear and cyclic modified peptide analogues of human MOG\({}_{35-55}\) epitope has the greatest efficacy in controlling EAE clinical symptoms. Our decision to use a large quantity of APLs (350 \(\upmu\)g, 10-fold excess) in co-injection with the rat MOG\({}_{35-55}\) epitope was based on our previous studies [10,31] in which the antagonism of MBP\({}_{87-99}\) modified linear and cyclic peptides in Lewis rats was achieved using more than 15-fold excess. MOG is a candidate autoanti in MS and the MOG\({}_{35-55}\) epitope represents one of the most immunomodant regions in humans and induces chronic EAE in C57 BL/6 mice. There is gathering evidence that mutations of disease-associated epitopes (APLs) can actively inhibit disease maybe through modification of interactions with the T cell receptor of encephalitogenic T-cells. Development of novel alternative molecules that mimic the immunomodulatory activity of MOG\({}_{35-55}\) epitope and maintaining an advantage over regular peptides in terms of stability is a necessary step before these molecules can be used for therapeutic purposes. There are several approaches in the development of such molecules. One is the design and synthesis of non-peptide mimetics with the same biological activity as the parent peptide or with antagonist activity. Another approach is the design of constrained cyclic peptide analogues. The cyclic peptide analogues could be designed using various lengths of bridges for their best flexibility in order to reach and enhance their binding to potential receptors. The selected bridging-atoms should allow for significant side-chain interactions with the receptor in order to be conserved. Cyclic analogues of MOG\({}_{35-55}\) offer several advantages compared to its linear counterparts such as increased resistance to metabolic degradation and conformational restriction. A potent constrained peptide analogue, offers important structural information regarding its bioactive conformation for further drug design. MOG\({}_{35-55}\) peptide (based on the rat sequence of MOG with Ser at position 42 instead of Pro in human MOG) induces chronic EAE while the MOG\({}_{35-55}\) human epitope was used for the design of APLs [32]. The amino acids Arg\({}^{41}\), Phe\({}^{44}\), Arg\({}^{46}\) and Val\({}^{47}\) of the rat MOG\({}_{38-51}\) epitope represent primary TCR contact residues while the Gly\({}^{38}\) and Phe\({}^{39}\) minor TCR contact sites [37]. Moreover, the development of spontaneous EAE in transgenic mice that express the MOG-specific TCR in the absence of MOG is possible, due to other neurological proteins that share similar TCR contact sites with the immunodominant MOG\({}_{35-55}\) epitope [38]. The TCR contact sites replacement of either Arg with Ala at positions 41, 46 or Tyr with Tyr(OMe) at position 40, results in linear peptide analogues with inhibitory effects with the [Ala\({}^{41}\)]MOG\({}_{35-55}\) to present the best inhibition in the EAE model.