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Studies of 17 analoges of 3 (SJA6017) in an in silico calpain model are reconciled to measured IC(50) values against ovine calpain. The studies validate the potential of the "model" and criteria established for inhibition as a tool to select structures for synthesis to test as calpain inhibitors. Using this screening methodology of virtual libraries led us to synthesize several inhibitors including macrocycle 33, which in vitro sheep eye lens culture experiments showed to substantially slow opacification.

A series of Val-Leu based peptidic aldehydes containing either a furan or thiophene at the N-terminus was prepared and assayed against ovine m-calpain. In general, potency is favoured by a 2-substituted (rather than 3-substituted) heterocycle, a thiophene rather than a furan, and a shorter chain length at the N-terminus. Molecular docking experiments provide some rationale for these observations.

We report the synthesis of macrocycles 1-6 via ring closing metathesis of dienes 7-12. Addition of chlorodicyclohexylborane to thermal and microwave assisted RCM of dienes 8 and 9 markedly improves the yield. The geometric isomers of macrocycles 1-3 and 5 have been assigned using X-ray crystallography and NMR.

The design and elaboration of a series of macrocyclic templates that exhibit a propensity to adopt a beta-strand-like peptide-backbone conformation led to potent and selective inhibitors of calpain 2. Macrocycle 1 retarded calcium-induced opacification in an ovine-lens culture assay and is a lead compound for the development of a drug for cataract treatment. Cbz=carbobenzyloxy.

A series of N-heterocyclic dipeptide aldehydes 4-13 have been synthesised and evaluated as inhibitors of ovine calpain 1 (o-CAPN1) and ovine calpain 2 (o-CAPN2). 5-Formyl-pyrrole 9 (IC(50) values of 290 and 25nM against o-CAPN1 and o-CAPN2, respectively) was the most potent and selective o-CAPN2 inhibitor, displaying >11-fold selectivity. The amino acid sequences of o-CAPN1 and o-CAPN2 have been determined. Because of the lack of available structural information on the ovine calpains, in silico homology models of the active site cleft of o-CAPN1 and o-CAPN2 were developed based on human calpain 1 (h-CAPN1) X-ray crystal structure (PDB code 1ZCM). These models were used to rationalise the observed SAR for compounds 4-13 and the selectivity observed for 9. The o-CAPN2 selective inhibitor 9 (CAT0059) was assayed in an in vitro ovine lens culture system and shown to successfully protect the lens from calcium-induced opacification.

The photoswitchable N-terminal diazo and triazene-dipeptide aldehydes 8a−d, 10a,b, and 17a,b present predominantly as the (E)-isomer, which purportedly binds deep in the S3 pocket of calpain. All compounds are potent inhibitors of m-calpain, with 8b being the most active (IC50 of 35 nM). The diazo-containing inhibitors 8a, 8c, and 10a were irradiated at 340 nm to give a photostationary state enriched in the (Z)-isomer, and in all cases, these were less active. The most water soluble triazene 17a (IC50 of 90 nM) retards calpain-induced cataract formation in lens culture.

Three new classes of conformationally constrained peptidomimetics, derived from modified α- and β-amino acids, have been prepared by ring closing metathesis (RCM). The first involves Cα–N′ cyclisation of the peptidic diene (23, 24, 26), the second Cβ2–N′ cyclisation (27, 28, 29), and the third N–Cβ2 cyclisation (30). The key C-centred olefin of the dienes was introduced by stereoselective α-allylation of either an α- or β-amino acid. The normal favourable influence of a tertiary amide linker in the diene towards RCM is negated by significant steric congestion, and the combination of a secondary amide linker and α,α-disubstitution promotes ring contraction on RCM.

Ring closing metathesis and cross metathesis approaches to a new macrocyclic peptidomimetic aldehyde 2 have been developed, with the former route being the most convenient. Aldehyde 2 is a potent inhibitor of calpain II (IC50 of 45 nM) with comparable activity to the benchmark acyclic inhibitor SJA6017 4. Both compounds contain an N-terminal 4-fluorophenylsulfonyl group. The P2 Ile analogue of 2 (16) is significantly less active (IC50 of 2000 nM) which reflects an unusually subtle importance of the P2 residue for active site binding.

Olefin metathesis, a process by which alkylidene groups on alkenes are exchanged, is reviewed with reference to the historical development of functional catalysts and microwave catalysis. The advent of new catalysts and improved reaction conditions has paved the way for the development of new and versatile domino processes that combine either different metathesis events or a key metathesis reaction with another reaction in a particularly atom economic and ‘green’ sense. We review this area as a convenient route to some otherwise difficult to prepare systems.