## 2 Results and Discussion
### Chemistry  
The synthesis of linear peptides [Ala\({}^{41}\)]MOG\({}_{35-55}\), [Ala\({}^{41,46}\)]MOG\({}_{35-55}\) and [TyrOMe\({}^{40}\)]MOG\({}_{35-55}\) as well as of the cyclic analogues cyclo(46-55)MOG\({}_{35-55}\) and cyclo(41-55)MOG\({}_{35-55}\) was carried out by using the Fmoc/tBu methodology, utilizing the 2-chlorotrityl chloride resin [33; 34; 35; 36] that was previously used for the synthesis of novel cyclic amide-linked analogues of peptides based on the MBP protein [10; 30; 31]. The side chain of amino acids were protected as follows: Trt for Asn, His, Pbf for Arg, tBu for Tyr (or TyrOMe), Ser, Glu and Boc for Lys, Trp. In the case of cyclic analogues, in which the side chain amine group of Lys at position 41 or 46 and \(C\)-terminal carboxyl group formed an amide bond, we used Mtt in linear precursor peptide to protect Lys. This group was removed upon treatment  
\begin{table}
\begin{tabular}{c c c c c c} \hline No. & Peptide & \(\mathbf{t_{R}}\) (min) & HPLC & Purity \% \# & HPLC & Purity \% \# & \(\mathbf{M_{cak.}}\) (Da) \\ \hline
1 & [Ala\({}^{41}\)]MOG\({}_{35-55}\) & 10.23 & 75.3 & 98.2 & 2505.27 \\
2 & [Ala\({}^{41,46}\)]MOG\({}_{35-55}\) & 10.89 & 74.3 & 98.9 & 2420.21 \\
3 & [TyrOMe\({}^{40}\)]MOG\({}_{35-55}\) & 09.16 & 58.2 & 98.7 & 2603.35 \\
4 & cyclo(46–55)MOG\({}_{35-55}\) & 10.57 & 29.1 & 97.7 & 2545.97 \\
5 & cyclo(41–55)MOG\({}_{35-55}\) & 09.96 & 31.8 & 97.9 & 2545.97 \\ \hline \end{tabular}  
* It is referred to crude (unpurified) peptides, after the final deprotection, based on the HPLC chromatograms;
* It is referred to purified peptides, used in EAE experiments, based on the HPLC chromatograms.  
\end{table}
Table 1: Analytical data of the synthesized linear and cyclic peptides.  
Figure 4: Co-administration of linear APLs, [Ala\({}^{41,46}\)]MOG\({}_{35-55}\) and [Ala\({}^{41}\)]MOG\({}_{35-55}\), ameliorate the clinical symptoms of rat MOG\({}_{35-55}\)-induced EAE in C57 BL/6 mice. Mean clinical scores of MOG-EAE are shown for all groups of mice against time (\(n=3\) for the cyclo (41–55) MOG\({}_{35-55}\) and \(n=5\) for other groups). Statistical significance after pair-wise comparisons (using Student’s \(t\) test) of each experimental group with the non-vaccinated control (PBS) group is shown (* \(p<0.05\)).  
of protected peptide-resin with the splitting mixture dichloromethane (DCM)/1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) (6/4) that used to cleave the peptide from the resin. The use of the 2-chlorotrityl chloride resin, as well as of mild cleaving conditions, allowed the protected peptides to release from the resin and the subsequent cyclization of the desired protected peptides. Cyclization was achieved using _O_-(benzotriazol-1-yl)-_N_,\(N\),_N_-tetramethylruriomu tetrafluoroborate (TBTU), 1-hydroxy-7-azabenzotriazole (HOAt) and 2,4,6-collidine (2,4,6-trimethylpyridine) allowing fast reaction and high yield cyclization product. The final deprotection of each peptide was achieved using trifluoroacetic acid (TFA) in DCM in presence of scavengers. The oxidation of thioether of Methionine to sulfoxide was revealed as a small peak, just before the main peak in analytical HPLC (Figure 1A). The oxidative product, in each peptide, was removed during the purification of synthesized analogues. Moreover, the TFA solution in final deprotection resulted to low yield of oxidation. The purification was achieved using semi preparative, reversed-phase HPLC, the peptide purity was assessed by analytical HPLC and by electron spray ionization mass spectrometry (ESI-MS). The purity of the synthesized peptides were verified by analytical HPLC (Figures 1-3), the identification and characterization of the final peptides were performed by ESI-MS (Table 1 and Figures 1-3).