Anja Johansson

Acyl sulfonamides as potent protease inhibitors of the hepatitis C virus full-length NS3 (protease-helicase/NTPase): A comparative study of different C-terminals

Synthesis and inhibitory potencies of three types of protease inhibitors of the hepatitis C virus (HCV) full-length NS3 (protease-helicase/NTPase) are reported: (i) inhibitors comprising electrophilic serine traps (pentafluoroethyl ketones, alpha-keto acids, and alpha-ketotetrazoles), (ii) product-based inhibitors comprising a C-terminal carboxylate group, and (iii) previously unexplored inhibitors comprising C-terminal carboxylic acid bioisosteres (tetrazoles and acyl sulfonamides). Bioisosteric replacement with the tetrazole group provided inhibitors equally potent to the corresponding carboxylates, and substitution with the phenyl acyl sulfonamide group yielded more potent inhibitors. The hexapeptide inhibitors Suc-Asp-D-Glu-Leu-Ile-Cha-Nva-NHSO(2)Ph and Suc-Asp-D-Glu-Leu-Ile-Cha-ACPC-NHSO(2)Ph with K(i) values of 13.6 and 3.8 nM, respectively, were approximately 20 times more potent than the corresponding inhibitors with a C-terminal carboxylate and were comparable to the carboxylate-based inhibitor containing the native cysteine, Suc-Asp-D-Glu-Leu-Ile-Cha-Cys-OH (K(i)=28 nM). The acyl sulfonamide group constitutes a very promising C-terminal functionality that allows for prime site optimization.

Diffusion and ionic conductivity in Li(CF3SO3)PEG10 and LiN(CF3SO2)2PEG10

Self-diffusion of the cation, the anion and the polymer chain in the low-molecular-weight polymer electrolyte systems Li(CF3SO3)PEG10 and LiN(CF3SO2)2PEG10 has been studied as a function of temperature using nuclear magnetic resonance spectroscopy. The diffusion behaviour is very similar in the two systems, where the cation is diffusing attached to the polymer chain, while the anion is moving slightly decoupled. However, all species are slightly more mobile in LiN(CF3SO2)2PEG10 than in Li(CF3SO3)PEG10, due to the plasticizing effect of the N(CF3SO2)2− ion, which also explains the higher conductivity for the LiN(CF3SO2)2PEG10 system. The temperature dependence of the diffusion coefficients is of Arrhenius type while the ionic conductivity follows the Vogel-Tammann-Fulcher relation. The diffusion has been compared with the conductivity by applying the Nernst-Einstein relation: a slight discrepancy is found for Li(CF3SO3)PEG10, while the agreement is very good for LiN(CF3SO2)2PEG10. Spin-lattice relaxation experiments have shown that the polymer chains undergo local conformational transformations, providing an additional diffusion mechanism for the ions.

Tetrapeptides as Potent Protease Inhibitors of Hepatitis C Virus Full-Length NS3 (Protease-Helicase/NTPase)

A library of tetrapeptides was evaluated for Hepatitis C Virus NS3 protease inhibitor activity in an in vitro assay system comprising the native bifunctional full-length NS3 (protease-helicase/NTPase) protein. Tetrapeptides with K(i) values in the high nanomolar range were identified, for example Suc-Chg-Glu-2-Nal-Cys (K(i)=0.27+/-0.03 microM) and Suc-Dif-Glu-Glu-Cys (K(i)=0.40+/-0.10 microM). Furthermore, it was shown that the inhibitory potencies are not affected significantly by assay ionic strength. As suggested by molecular modelling, potential binding interactions of the tetrapeptide inhibitors with the helicase domain might explain the data and structure-activity relationships thus obtained. Hence, we postulate that the full-length NS3 assay is a relevant system for inhibitor identification, offering new opportunities for inhibitor design.

Inhibition of hepatitis C virus NS3 protease activity by product-based peptides is dependent on helicase domain.

Structure activity relationships (SARs) of product-based inhibitors of hepatitis C virus NS3 protease were evaluated using an in vitro assay system comprising the native bifunctional full-length NS3 (protease-helicase/NTPase). The results were compared to previously reported data derived from the corresponding NS3 protease domain assay. Shortening the length of the protease inhibitors from hexapeptides to tripeptides revealed that the decrease in potency was much less when determined in the assay system with the full-length NS3 protein. Disagreements in SARs at different positions (P5 P2) were also discovered. Taken together, the results suggest that the impact of the helicase domain upon protease inhibitor binding is substantial.

Vinyl Sulfide Cyclized Analogues of Angiotensin II with High Affinity to the AT1-Receptor

We have an interest in developing alternative methods for cyclization of peptides. The use of aldehyde precursors as building blocks for the synthesis of bicyclic Ang II analogues was previously reported by us [1,2]. We herein present the aldehyde precursor 1 and its use in a novel cyclization method, creating the vinyl sulfide bridged Ang II analogues 3 and 4 (Figure 1).

Different Types of P1 Residues in Peptide-Based Inhibitors of Hepatitis C Virus Full-Length NS3 Protease

In a project aimed at the design and synthesis of inhibitors to the hepatitis C virus (HCV) NS3 serine protease, peptides containing different types of P1 residues were compared: 1) pentafluoroethyl ketones, since substrate-based peptides containing electrophilic groups in the P1 position are classical inhibitors of serine proteases; 2) tetrazoles, since HCV NS3 protease is uniquely inhibited by its N-terminal cleavage products [1] and tetrazoles are known metabolically stable bioisosteres for carboxylic acids; 3) ketotetrazoles, since peptide α-ketoacids work as electrophilic inhibitors of the HCV NS3 protease [2] and oc-ketotetrazoles should serve as α-ketoacid bioisosteres and; 4) carboxylic acids, as reference compounds.