Research On The Design, Synthesis And Long-action Of PEGylated Tα1 Analogs

In order to improve the pharmacological and pharmacokinetic properties of thymosin alpha l(Tα1), poly(ethylene glycol) (PEG) modification (PEGylation) was performed in this thesis. By assaying the bioactivities, secondary structures and pharmacokinetic profiles of synthetic compounds, the influences of macromolecular modification on peptides were systematically elucidated, which provided some conceptions and good advices for developing the PEG modification research of peptides.The chosen sites and site-directed modifications are of extreme importance. The secondary conformations of peptides exert remarkable influence on the bioactivity. Thus several sites of Tα1 were selected at the N-terminus, C-terminus, α-helix region, β-turn region and random coil regions, according to SAR results of literatures. And [Cys^x]Tα1 (x: the chosen sites) analogs were synthesized first to conveniently induce the cavalent attachment mPEG₅₀₀₀-MAL to Tal analoges. Summarizing the finished work as follows:1. Nine [Cys^x]Tα1 were synthesized with solid phase peptide synthesis (SPPS), purified by RP-HPLC or Flash column, and confirmed by ESI-MS. And the corresponding PEGylated products, [Cys^x(mPEG₅₀₀₀-MAL)]Tα1, were subsequently synthesized in solution, purified by RP-HPLC, confirmed by MALDI-TOF-MS.2. Immunoactivities of the synthetic compounds were tested in vitro, and the results indicate that the cystein substitutions at some positions take great effects on the activity, but PEGylations don't influence much. All compounds, but TC8, TC24, and PT8, can stimulate lymphocytes to product IFN-γ, moreover, TC17, TC, CT, TCP and PCT even better than Tal. In the proliferative experiment, except CT, TC16, TC24 and PT24, all compounds can increase the proliferative responses of the mitogen stimulated lymphocytes.3. Results of metabolic evaluation in vivo showed that PEGylated products indeed improve the metabolic profiles of the peptide drug, and the influences of PEGylation differ from the modified sites. Administration 24h later, the values of AUC of the compounds PT8, PT11, PT16, PT17, PT21 and PT24 are 8²0 times of Tα1, but...