Philippe Karoyan

N- and Cα-Methylation in Biologically Active Peptides: Synthesis, Structural and Functional Aspects

Numerous backbone constraints can be used to develop pseudopeptides or pseudomimetics of biologically active peptides. Among those, N- and Calpha-methyl amino acids that can be incorporated by solid-phase peptide synthesis in a bioactive sequence represent important tools to restrict phi and psi angles of peptide backbone. This review will focus on the chemical syntheses of N- and Calpha-methyl amino acids, their effects on peptide conformation and structure, and their role on the peptide stability towards enzymatic degradation and on the biological activities of the resulting analogues.

New strategy for the synthesis of 3-substituted prolines

Abstract Ring formation involving a 5-exo trig cyclization between a zinc enolate and a non activated double bond led to cis diastereoisomer of 3-substituted prolines. This cyclization was achieved with transfer of chirality onto the C-2 carbon when nitrogen was protected by an α-methylbenzyl group. Reprotonation of the lithium enolate of cis derivative yielded the trans diastereoisomer.

Further Delineation of the Two Binding Sites (R*n) Associated with Tachykinin Neurokinin-1 Receptors Using [3-Prolinomethionine11]SP Analogues

Two binding sites are associated with neurokinin-1 substance P receptors in both transfected cells and mammalian tissues. To further delineate the interactions between the crucial C-terminal methionine of substance P and these two binding sites, we have incorporated newly designed constrained methionines,i.e. (2S, 3S)- and (2S,3R)-prolinomethionines. The potencies of these C terminus-modified SP analogues to bind both sites and to activate phosphatidylinositol hydrolysis and cAMP formation have been measured, together with those of their corresponding sulfoxides and sulfones. The molecular nature of these two binding sites and their selective coupling to effector signaling pathways are discussed in the light of current models of receptor activation. The less abundant binding site is coupled to Gq/11 proteins, whereas the most abundant one interacts with Gs proteins in Chinese hamster ovary cells transfected with human neurokinin-1 receptors. The specific orientation of the C-terminal methionine side chain imposed by these constraints shows that macroscopically χ1 and χ2 angles of this crucial C-terminal residue are similar in both binding sites. However, slight but significant variations in the rotation around the Cγ–S bond yield different either stabilizing or destabilizing interactions in the two binding sites. These results highlight the need of such constrained amino acids to probe subtle interactions in ligand-receptor complexes.

Synthesis and Evaluation of Analogues of N-Phthaloyl-l-tryptophan (RG108) as Inhibitors of DNA Methyltransferase 1

DNA methyltransferases (DNMT) are promising drug targets in cancer provided that new, more specific, and chemically stable inhibitors are discovered. Among the non-nucleoside DNMT inhibitors, N-phthaloyl-l-tryptophan 1 (RG108) was first identified as inhibitor of DNMT1. Here, 1 analogues were synthesized to understand its interaction with DNMT. The indole, carboxylate, and phthalimide moieties were modified. Homologated and conformationally constrained analogues were prepared. The latter were synthesized from prolinohomotryptophan derivatives through a methodology based amino-zinc-ene-enolate cyclization. All compounds were tested for their ability to inhibit DNMT1 in vitro. Among them, constrained compounds 16-18 and NPys derivatives 10-11 were found to be at least 10-fold more potent than the reference compound. The cytotoxicity on the tumor DU145 cell line of the most potent inhibitors was correlated to their inhibitory potency. Finally, docking studies were conducted in order to understand their binding mode. This study provides insights for the design of the next-generation of DNMT inhibitors.

β-Homo-amino Acid Scan of Angiotensin IV

Angiotensin IV, a metabolite of angiotensin II, inhibits the enzyme insulin regulated aminopeptidase or IRAP and also, although with lower potency, aminopeptidase-N (AP-N). When both beta (2)-homo amino acid- and beta (3)-homo amino acid substitutions were used, allowed the identification of H-( R)beta (2)hVal-Tyr-Ile-His-Pro-beta (3)hPhe-OH as a potent and stable Ang IV analog with high selectivity for IRAP versus AP-N and the AT1 receptor.

Amino-zinc-ene-enolate cyclization: a short access to cis-3-substituted prolino-homotryptophane derivatives.

Proline chimeras are useful tools for medicinal chemistry and/or biological applications. The asymmetric synthesis of cis-3-substituted prolines can be easily achieved via amino-zinc-ene-enolate cyclization followed by transmetalation of the cyclic zinc intermediate for further functionalization. Syntheses of prolino-homotryptophane derivatives were achieved through Negishi cross-coupling of the zinc intermediate with indole rings. The use of Pd catalyst derived from Fus [(t-Bu3)PH]-BF4 was required to avoid the undesired beta-hydride elimination. Optically pure and orthogonally protected compounds were obtained readily usable for peptide synthesis.

Short asymmetric synthesis of (2S,3S)- and (2S,3R)-3-prolinoglutamic acids: 2-carboxy-3-pyrrolidine-acetic acids (CPAA)

Abstract The asymmetric synthesis of both cis and trans 3-prolinoglutamic acids can be easily achieved in a diastereoselective and enantioselective way via the amino–zinc–enolate cyclisation.

3-Substituted Prolines: From Synthesis to Structural Applications, from Peptides to Foldamers

Among the twenty natural proteinogenic amino acids, proline is unique as its secondary amine forms a tertiary amide when incorporated into biopolymers, thus preventing hydrogen bond formation. Despite the lack of hydrogen bonds and thanks to conformational restriction of flexibility linked to the pyrrolidine ring, proline is able to stabilize peptide secondary structures such as β-turns or polyproline helices. These unique conformational properties have aroused a great interest in the development of proline analogues. Among them, proline chimeras are tools combining the proline restriction of flexibility together with the information brought by natural amino acids side chains. This review will focus on the chemical syntheses of 3-substituted proline chimeras of potential use for peptide syntheses and as potential use as tools for SAR studies of biologically active peptides and the development of secondary structure mimetics. Their influence on peptide structure will be briefly described.

Asymmetric synthesis of (2S,3S)- and (2S,3R)-3-prolinomethionines: 3-methylsulfanylmethyl-pyrrolidine-2-carboxylic acids

Abstract The synthesis of 3-prolinomethionine can be easily achieved in a diastereoselective and enantioselective way via zinc-enolate cyclisation. After transmetallation by CuCN.2LiCl, the zinc-copper derivative was reacted with S -methyl methanesulfonothioate leading in a “one-pot” procedure, to N-(α-methylbenzyl)-3-prolinomethionine benzyl ester. The α-methylbenzyl group was transformed in vinyl-oxycarbonyl and tertiobutyl-oxycarbonyl groups successively. Reprotonation of cis Voc prolinomethionine enolate at low temperature yielded the enantiomerically pure trans diastereoisomer.

Complement factor H inhibits CD47-Mediated resolution of inflammation

SUMMARY Variants of the CFH gene, encoding complement factor H (CFH), show strong association with age‐related macular degeneration (AMD), a major cause of blindness. Here, we used murine models of AMD to examine the contribution of CFH to disease etiology. Cfh deletion protected the mice from the pathogenic subretinal accumulation of mononuclear phagocytes (MP) that characterize AMD and showed accelerated resolution of inflammation. MP persistence arose secondary to binding of CFH to CD11b, which obstructed the homeostatic elimination of MPs from the subretinal space mediated by thrombospsondin‐1 (TSP‐1) activation of CD47. The AMD‐associated CFH(H402) variant markedly increased this inhibitory effect on microglial cells, supporting a causal link to disease etiology. This mechanism is not restricted to the eye, as similar results were observed in a model of acute sterile peritonitis. Pharmacological activation of CD47 accelerated resolution of both subretinal and peritoneal inflammation, with implications for the treatment of chronic inflammatory disease. Graphical Abstract Figure. No Caption available. HighlightsCFH deficiency restricts chronic subretinal mononuclear phagocyte accumulationIn inflammation resolution, MPs are eliminated via a CD47‐dependent mechanismCFH binding to integrin CD11b curbs TSP‐1 activation of integrin‐associated CD47The AMD‐associated CFH variant CFH(H402) potently inhibits microglial cell elimination &NA; Variants in complement factor H (CFH) show strong association to age‐related macular degeneration. Calippe et al. find that CFH binding to mononuclear phagocytes (MP) curbs the CD47‐mediated elimination of MPs that maintains homeostasis in the subretinal space. The AMD‐associated CFH variant CFH(H402) increases subretinal MP accumulation, providing insight into disease etiology.