Review
Enzymatic synthesis of chiral intermediates for Omapatrilat, an antihypertensive drug

https://doi.org/10.1016/S1389-0344(01)00068-5Get rights and content

Abstract

Biocatalytic processes were used to prepare chiral intermediates required for the synthesis of Omapatrilat 1 by three different routes. The synthesis and enzymatic conversion of 2-keto-6-hydroxyhexanoic acid 3 to l-6-hydroxynorleucine 2 was demonstrated by reductive amination using beef liver glutamate dehydrogenase. To avoid the lengthy chemical synthesis of the ketoacid 3, a second route was developed to prepare the ketoacid by treatment of racemic 6-hydroxy norleucine [readily available from hydrolysis of 5-(4-hydroxybutyl) hydantoin 4] with d-amino acid oxidase from porcine kidney or Trigonopsis variabilis followed by reductive amination to convert the mixture completely to l-6-hydroxynorleucine in 98% yield and 99% enantiomeric excess (e.e.). The enzymatic synthesis of (S)-2-amino-5-(1,3-dioxolan-2-yl)-pentanoic acid (allysine ethylene acetal, 5) was demonstrated using phenylalanine dehydrogenase (PDH) from T. intermedius. Phenylalanine dehydrogenase was cloned and overexpressed in Escherichia coli and Pichia pastoris. Using PDH from E. coli or P. pastoris, the enzymatic process was scale-up to prepare kg quantity of allysine ethylene acetal 5. The reaction yields of >94% and e.e. of >98% were obtained for allysine ethylene acetal 5. An enzymatic process was developed for the synthesis of [4S-(4a,7a,10ab)]1-octahydro-5-oxo-4 [[(phenylmethoxy)carbonyl]amino]-7H-pyrido-[2,1-b] [1,3]thiazepine-7-carboxylic acid [BMS-199541-01]. The enzymatic oxidation of the ε-amino group of lysine in the dipeptide dimer N2-[N[[(phenyl-methoxy)carbonyl] l-homocysteinyl] l-lysine)-1,1-disulphide [BMS-201391-01] to produce BMS-199541-01 using a novel l-lysine ε-aminotransferase (LAT) from Sphingomonas paucimobilis SC 16113 was demonstrated. This enzyme was overexpressed in E. coli and a process was developed using the recombinant enzyme.

Introduction

Currently much attention is being focused on the interaction of small molecules with biological macromolecules. The search for selective enzyme inhibitors and receptor agonists/antagonists is key for target-oriented research in the pharmaceutical and agrochemical industries. Increased understanding of the mechanism of drug interactions on a molecular level has led to a strong awareness of the importance of chirality as the key to the efficacy of many drug products and agrochemicals. The production of optically active chiral intermediates is a subject of increasing importance in the pharmaceutical industry. Increasing regulatory pressure to market homochiral drugs [1], [2] has led to the use of chemical or chemo-enzymatic synthesis of chiral compounds. The advantages of biocatalysis over chemical catalysis are that enzyme-catalyzed reactions are often highly stereoselective and regioselective and can be carried out at ambient temperature and atmospheric pressure, thus by avoiding extreme conditions one can minimize problems of isomerization, racemization, epimerization, and rearrangements that may occur during typical chemical processes. Biocatalytic processes catalyzed by microbial cells and enzymes derived therefrom can be immobilized and reused for many cycles. In addition, enzymes can be overexpressed to make biocatalytic processes economically efficient. The idea of designing biocatalysts that would act specifically in any desired reaction of interest will change the face of synthesis. Tailor-made enzymes with modified activity and the preparation of thermostable and pH stable enzymes produced by random and site-directed mutagenesis have lead to the production of novel stereoselective biocatalysts. The use of enzymes in organic solvents has led to hundreds of publications on enzyme-catalyzed asymmetric synthesis and resolution processes. Molecular recognition and selective catalysis are key chemical processes in life which are embodied in enzymes. A number of review articles [3], [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15], [16], [17], [18] have been published on the use of biocatalysis in organic synthesis. This chapter provides different approaches towards the use of microbial enzymes for the synthesis of chiral intermediates for Omapatrilat 1 (Fig. 1) by three different routes. Omapatrilat 1 is an antihypertensive drug which acts by inhibiting angiotensin-converting enzyme (ACE) and neutral endopeptidase [19]. ACE a zinc-containing carbopeptidase, catalyzes the conversion of the decapeptide angiotensin I (AI) to the octapeptide angiotensin II (AII). AII is a potent vasoconstrictor which also triggers the release of aldosterone, a sodium-retaining steroid. Thus ACE raises blood pressure by increasing both vascular resistance and fluid volume. Effective inhibitors of ACE have been used not only in the treatment of hypertension but in the clinical management of congestive heart failure [20]. Neutral endopeptidase (NEP), like ACE, is a zinc metalloprotease and is found in high concentration in the brush border of the renal proximal tubule. NEP is highly efficient in degrading atrial natriuretic peptide (ANP), a 28-amino acid peptide secreted by the heart in response to atrial distension. ANP has opposing hormonal actions to those of AII. By interaction with its receptor, ANP promotes the generation of cGMP via guanylate cyclase activation, thus resulting in vasodilatation, natriuresis, diuresis, and inhibition of aldosterone [21]. Therefore simultaneous potentiation of ANP via NEP inhibition and attenuation of AII via ACE inhibition should lead to complementary effects in the management of hypertension and congestive heart failure [22], [23].

Section snippets

Synthesis of key intermediates for omapatrilat

The enzymatic and/or microbial synthesis of single enantiomers of three chiral intermediates by three different routes to Omapatrilat 1 were developed as described below.

Conclusion

The production of optically active chiral intermediates is a subject of increasing importance to the pharmaceutical industry. Increasing regulatory pressure to market homochiral drugs [1], [2] has led to the use of alternative approaches, including biocatalysis for the synthesis of chiral compounds. Organic synthesis has been one of the most successful scientific disciplines and has enormous practical utility. One can ask question then why biocatalysis? What has biocatalysis to offer to

Acknowledgements

The author would like to acknowledge Drs Ronald Hanson, Amit Banerjee, Venkata Nanduri, Jeffrey Howell, Steven Goldberg, Robert Johnston, Paul Cino, Mary-Jo Donovan, Dana Cazzulino, Shanker Swaminathan, Lawrence Parker, John Venit, Thomas LaPorte, Thomas Tully, John Wasylyk, Laszlo Szarka, Michael Montana, Sushil Srivastava, Jerome Moniot and Raphael Ko for research collaboration during this work. The author would also like to acknowledge and thank Drs Richard Mueller and John Venit for their

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