Molecular Modeling, Synthesis, and Biological Evaluation of Macrocyclic Calpain Inhibitors

Abstract

The introduction of a macrocycle into a biologically active peptide can increase potency [1,2] and selectivity [3] by reducing the entropic penalty of inhibitor-enzyme binding. The incorporation of macrocycles in peptides has been used to mimic secondary structure, such as extended b-strand-like and bent b-turn-like conformations. [3,4] The challenge is to devise protocols for the design, evaluation, and synthesis of macrocyclic templates that provide access to families of inhibitors. [5] Such templates should 1) have a well-defined conformation that can be readily assessed by computational screening, 2) be readily synthesized from natural building blocks, and 3) be easily modified to target an enzyme for a specific biological application.Herein we present studies on the 16-19-membered macrocycles 1-4 (Scheme 1) designed to be constrained into a b-strand-like geometry, a conformation universally adopted by inhibitors and substrates on binding to a protease. [6] We report a versatile approach based on ring-closing metathesis (RCM) to these orthogonally protected templates as well as computational analysis of their potential to form a b strand and of their binding to a target protease. The templates were converted into aldehydes 1 d-4 d as potential inhibitors of the calcium-activated cysteine protease calpain. The alcohols 1 c-4 c were evaluated to establish whether the macrocycles negate the need for a reactive warhead. Calpain was chosen as a challenging target for the design of selective inhibitors, [7] and because its link to cortical cataracts [8] provides an opportunity to assess the in vivo efficacy of our approach. The acyclic tripeptide analogues 5 a-d were also investigated to establish the importance (or unimportance) of the macrocyclic constraint for potency and selectivity of inhibition.Conformational searches [9] were carried out on 1 a-d to 5 a-d to assess their ability to adopt a b-strand conformation. The resulting ensembles of low-energy conformers were examined by XCluster (see the Supporting Information for detailed results). For the 16-membered macrocycles 1 a-d, all conformers (> 99 %) within 12 kJ mol À1 of their global minima adopt a b-strand conformation (see Figure 1 for Scheme 1. Macrocyclic templates and acyclic peptides. Dihedral angles F and Y that describe a b strand are shown. Boc = tert-butoxycarbonyl, Cbz = carbobenzyloxy. Figure 1. Overlaid conformers of the two clusters of 1 d.

Dynamic Docking of Conformationally Constrained Macrocycles: Methods and Applications

“…In their search for compounds to inhibit the cysteine protease calpain, toward the prevention of cataractogenesis, Stuart et al designed constrained bridged peptides with the goal of holding the ligand in a β-strand-like conformation. 58,59 The designed compounds were then subjected to an MCMM conformational search in order to determine the likelihood that the ligands would maintain the desired conformation, indicated by the percentage of conformers with a characteristic β-strand shape. Interestingly, though macrocyclic ligands with a 16member macrocycle possessed nearly 100% β-strand character, the most potent compound in the series, CAT811, displayed only 8% β-strand character.…”

Fixierung cyclischer Peptide: Mimetika von Proteinstrukturmotiven

“…B. Angiotensin-umwandelndes Enzym (ACE), neutrale Endopeptidase NAP (wie 25), Thermolysin, Aminopeptidase, MMP1, -3, -8, -9 (wie 26), TNFa-umwandelndes Enzym (TACE)), und Cystein-Proteasen wie Calpaine (wie 27). [31] [32] Es gibt eines großes, noch unerschlossenes Potenzial für den Entwurf von b-Strang-und b-Faltblatt-Mimetika, mit denen die Protein-Protein-Wechselwirkungen gestçrt oder die Bindung eines der Partner nachgeahmt werden soll. [6,7] PPI spielen sich oft an großen polaren Regionen der Oberfläche mit nur flachen Einbuchtungen ab, die oft nicht effektiv von kleinen hydrophoben Molekülen besetzt werden…”

“…Mit 84 wurde ein Antagonist des onkogenen ternären NOTCH-1-Transkriptionsfaktor-Komplexes entwickelt. Das 15 Aminosäuren lange Peptid von MAML1 [21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36] Abbildung 15. Verschieden lange Alkyllinker zwischen den Positionen i!i + 3, i!i + 4 und i!i + 7.…”

Synthesis and Evaluation of Macrocyclic Peptide Aldehydes as Potent and Selective Inhibitors of the 20S Proteasome

“…18 A SAR by Abell et al on macrocyclic peptide aldehyde inhibitors of ovine calpain found that the addition of an aldehyde to a peptide macrocycle can increase potency as much as ∼1000-fold. 19 The results of Abell et al combined with the proteasome's flexible S 2 and S 4 pockets suggest that a macrocyclic peptide aldehyde with a suitable peptide sequence will yield a potent and selective inhibitor of the 20S proteasome. Other desirable pharmacological properties introduced by the macrocycle include increased metabolic stability, cellular uptake, and tissue distribution.…”