Syntheses of potent and selective MMP-13 inhibitors with increased metabolic stability

Abstract

MMP-13 inhibitors A dysregulation of matrix metalloproteinase (MMP)-13 has been correlated with pathophysiological conditions like human carcinomas, rheumatoid arthritis, and osteoarthritis. This makes MMP-13 a suitable target for drug development. Since the family of MMPs comprises more than 23 different members in humans, a potential drug candidate should selectively and potently inhibit MMP-13 in order to avoid side effects resulting from a lack of selectivity. Additionally, the desired drug candidate should have a suitable pharmacokinetic profile. To realize the aim of developing potent and selective inhibitors of MMP-13 that are metabolically more stable, the chemical structure of lead compound 41, a potent and selective picomolar inhibitor of MMP-13 from an earlier work of KALININ et al., was extensively varied to access structurally diverse inhibitors (Figure I). First, to avoid a possible O-dealkylation, the oxymethylene moiety of the lipophilic side chain of 41 was replaced with ethylene, vinylene, and acetylene moieties (Figure I). These varied lipophilic side chains were accessed from starting materials like 1,4-diiodobenzene (43), 1-bromo-4-iodobenzene (80), and 4-iodoaniline (49) by employing reactions like C-C couplings, catalytic hydrogenation, semi-reduction of alkynes, diazotization, and desilylation. The respective fluorinated derivatives were obtained by reacting the various hydroxylated side chains with DAST. Starting from hydroxyproline derivative 57, amide 59, the central building block of the synthesis, was obtained by benzoylation, MITSUNOBU, and saponification reactions. Thus, hydroxamic acids I were obtained after SONOGASHIRA coupling reactions of the various side chains with aryl iodide 59 and aminolyses with hydroxylamine. In order to replace the carbonyl function of 41 with a sulfonyl moiety, which is more stable to hydrolysis, proline derivative 57 was transformed into diastereomeric sulfonamides II via sulfonylation, MITSUNOBU, and saponification reactions. Hence, sulfonamide-based hydroxamic acids 72 and 73 were afforded by C-C couplings with ether-based terminal alkyne 71 and aminolyses. To further broaden structure-activity relationships, the lipophilic chain of 41 was moved from position 1 to position 4 of the pyrrolidine ring where it was anchored via benzyloxy, benzylamino, and phenyltriazolyl moieties. At the same time, different residues like methyl, benzyl, ethyl, benzoyl, acetyl, and formyl were substituted onto the pyrrolidine nitrogen. Additionally, N-unsubstituted proline derivatives were synthesized (Figure I). Hydroxamic acids III with all these modifications were accessed in chiral pool syntheses starting from methyl (2S,4R)-4-hydroxypyrrolidine-2-carboxylate hydrochloride (57) and using reactions like acylation, alkylation, Boc protection, WILLIAMSON ether syntheses, copper-catalyzed azide-alkyne cycloadditions, reductive aminations, aza-WITTIG reactions, hydrogenation, C-C couplings, and aminolyses. The newly synthesized inhibitors I with varied side chains were revealed to be less potent and less selective MMP-13 inhibitors than lead compound 41 (IC50 = 0.07 nM), though they were single-digit nanomolar inhibitors. The most potent in the series was hydroxamic acid 66 (IC50 = 1.7 nM) possessing a trans-alkenyl side chain. Fluorination was found to not significantly affect MMP-13 inhibitory activity, thus, producing inconsistent outcomes for both potency and selectivity. The sulfonamides II exhibited moderate MMP-13 inhibitory activity, being inferior to lead compound 41 in potency and selectivity. The (4S)-configured sulfonamide 73 was more potent than its (4R)-configured diastereomer 72. The triazole-based, ether-based, and 4-aminoproline based hydroxamic acids III exhibited no MMP-13 inhibitory activity. These findings suggested that (4S)-configuration in position 4 of the pyrrolidine ring, the oxymethylene moiety of 41, and the pyrrolidine nitrogen as anchoring point for the lipophilic side chains are pivotal for activity.

LpxC inhibitors Bacterial infections caused by multi-drug resistant bacteria cause thousands of deaths every year. Though much progress has been made in the treatment of infections caused by multi-drug resistant Gram-positive organisms, the treatment of infections caused by multi-drug resistant Gram-negative organism is elusive due to the development of resistance. To overcome this challenge, there is a need to develop new antibacterial agents that target enzymatic pathways that have not been exploited before. Gram-negative bacteria possess a unique outer membrane of which lipid A is an essential component. Lipid A, the hydrophobic membrane anchor of the lipopolysaccharides, is very important for bacterial viability and survival, hence, targeting the enzymes involved in lipid A biosynthesis provides an attractive strategy for the development of new antibacterial agents. Of the nine enzymes being involved in lipid A biosynthesis, the UDP-3-O-((R)-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC), catalyzing the second overall and first irreversible step of this biosynthetic route, is a promising target for antibacterial drug development. Hence, as part of this project, LpxC inhibitors were developed using selected intermediates of the elaborated syntheses of the previously described MMP-13 inhibitors. As such, sulfonamide-based hydroxamic acids IV as well as ether-based, triazole-based, and 4-aminoproline-based hydroxamic acids V were synthesized (Figure II) by using the same reactions and conditions used for accessing the MMP-13 inhibitors, except that the morpholine-based terminal alkyne 103 was used in the SONOGASHIRA coupling reactions to build up the lipophilic side chain (Figure II). The antibacterial and LpxC inhibitory activities of the newly synthesized compounds were evaluated. All the newly synthesized LpxC inhibitors were found to be less potent than lead compound CHIR-090 (34) (Ki = 0.008 μM), a threonine-based hydroxamic acid. The sulfonamide-based hydroxamic acids IV (Ki = 1.4 µM and 0.72 µM) were potent LpxC inhibitors with the (4S)-configured derivative 79 being a better LpxC inhibitor and antibacterial agent than its diastereomer 78. The (4S)-configured ethers were better LpxC inhibitors than their respective (4R)-configured diastereomers. A concurrent trend was observed for antibacterial activity of the ethers against E. coli D22 except for the N-unsubstituted derivatives that exhibited similar activity. Among the series of ethers, the (4S)-configured N-benzyl-substituted derivative 141 was the most potent LpxC inhibitor (Ki = 0.582 µM) and antibacterial agent. All the ether-based hydroxamic acids were inactive against E. coli BL21 (DE3). The triazoles had weak antibacterial activity. In spite of this, they displayed promising LpxC inhibitory activity with the benzamide-based triazoles being better LpxC inhibitors than the N-benzyl-substituted triazoles. Regarding the 4-aminoprolines, the benzamide-based derivative 217 was a strong LpxC inhibitor while the N-benzyl-substituted derivative 227 had very little to no activity. A reversed trend was observed for the 4-aminoprolines with regard to the susceptibility of E. coli D22. Like the ethers and triazoles, the 4-aminoproline derivatives 217 and 227 (MICs ˃64) had no activity against E. coli BL21