Pablo Martín-Gago

A tetradecapeptide somatostatin dicarba-analog: Synthesis, structural impact and biological activity

We described here the first tetradecapeptide somatostatin-analogue where the disulfide bridge has been replaced by a carbon-carbon double bond. This analogue was prepared using microwave assisted ring closing metathesis (RCM) using the 2nd generation Grubbs as catalyst. Under our optimized conditions the cyclization between allylGly 3 and 14 proceeded in moderate yield, excellent cyclic/linear ratio and very high Z-double bond selectivity. NMR studies also demonstrated that the conformational flexibility of this peptide is increased in comparison to that of the natural hormone. Remarkably, this alkene-bridged somatostatin analog is highly selective against somatostatin receptors 1 and 5, suggesting that conformational rigidity is not required for the efficient interaction of somatostatin analogues with these two receptors.

SSTR1- and SSTR3-selective somatostatin analogues.

We prepared the two enantiomers of 3‐(3′‐quinolyl)‐alanine (Qla, 1) in multigram scale by asymmetric hydrogenation. These amino acids, protected as Fmoc derivatives, were then used in the solid‐phase synthesis of two new somatostatin 14 (SRIF‐14) analogues 8 a and 8 b, tetradecapeptides in which the tryptophan residue (Trp8) is replaced by one of the two enantiomers of 3‐(3′‐quinolyl)‐alanine (Qla8) and therefore lack the NH bond in residue 8. The selectivity of these new analogues for the somatostatin receptors, SSTR1–5, was measured. Substitution with L‐Qla8 yielded peptide 8 a, which was highly selective for SSTR1 and SSTR3, with an affinity similar to that of SRIF‐14. Substitution by D‐Qla gave the relatively selective analogue 8 b, which showed high affinity for SSTR3 and significant affinity for SSTR1, SSTR2 and SSTR5. The biological results demonstrate that bulky and electronically poor aromatic amino acids at position 8 are compatible with strong activity with SSTR1 and SSTR3. Remarkably, these high affinity levels were achieved with peptides in which the conformational mobility was increased with respect to that of SRIF‐14. This observation suggests that conformational rigidity is not required, and might be detrimental to the interaction with receptors SSTR1 and SSTR3. The absence of an indole N proton in Qla8 might also contribute to the increased flexibility observed in these analogues.

Insights into Structure-Activity Relationships of Somatostatin Analogs Containing Mesitylalanine

The non-natural amino acid mesitylalanine (2,4,6-trimethyl-L-phenylalanine; Msa) has an electron-richer and a more conformationally restricted side-chain than that of its natural phenylalanine counterpart. Taking these properties into account, we have synthesized ten somatostatin analogs containing Msa residues in different key positions to modify the intrinsic conformational flexibility of the natural hormone. We have measured the binding affinity of these analogs and correlated it with the main conformations they populate in solution. NMR and computational analysis revealed that analogs containing one Msa residue were conformationally more restricted than somatostatin under similar experimental conditions. Furthermore, we were able to characterize the presence of a hairpin at the pharmacophore region and a non-covalent interaction between aromatic residues 6 and 11. In all cases, the inclusion of a D-Trp in the eighth position further stabilized the main conformation. Some of these peptides bound selectively to one or two somatostatin receptors with similar or even higher affinity than the natural hormone. However, we also found that multiple incorporations of Msa residues increased the life span of the peptides in serum but with a loss of conformational rigidity and binding affinity.

Peptide aromatic interactions modulated by fluorinated residues: Synthesis, structure and biological activity of Somatostatin analogs containing 3-(3',5'difluorophenyl)-alanine

Somatostatin is a 14-residue peptide hormone that regulates the endocrine system by binding to five G-protein-coupled receptors (SSTR1–5). We have designed six new Somatostatin analogs with L-3-(3′,5′-difluorophenyl)-alanine (Dfp) as a substitute of Phe and studied the effect of an electron-poor aromatic ring in the network of aromatic interactions present in Somatostatin. Replacement of each of the Phe residues (positions 6, 7 and 11) by Dfp and use of a D-Trp8 yielded peptides whose main conformations could be characterized in aqueous solution by NMR. Receptor binding studies revealed that the analog with Dfp at position 7 displayed a remarkable affinity to SSTR2 and SSTR3. Analogs with Dfp at positions 6 or 11 displayed a π-π interaction with the Phe present at 11 or 6, respectively. Interestingly, these analogs, particularly [D-Trp8,L-Dfp11]-SRIF, showed high selectivity towards SSTR2, with a higher value than that of Octreotide and a similar one to that of native Somatostatin.

Arylfluorosulfate-Based Electrophiles for Covalent Protein Labeling: A New Addition to the Arsenal

Abstract Selective covalent modification of a targeted protein is a powerful tool in chemical biology and drug discovery, with applications ranging from identification and characterization of proteins and their functions to the development of targeted covalent inhibitors. Most covalent ligands contain an affinity motif and an electrophilic warhead that reacts with a nucleophilic residue of the targeted protein. Because the electrophilic warhead is prone to react and modify off‐target nucleophiles, its reactivity should be balanced carefully to maximize target selectivity. Arylfluorosulfates have recently emerged as latent electrophiles for selective labeling of context‐specific tyrosine and lysine residues in protein pockets. Here, we review the recent but intense introduction of arylfluorosulfates into the arsenal of available warheads for selective covalent modification of proteins. We highlight the untapped potential of this functional group for use in chemical biology and drug discovery.