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Oxidosqualene Cyclase (OSC) Inhibitors for the Treatment of Dyslipidemia

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Oxidosqualene Cyclase (OSC) Inhibitors for the Treatment of Dyslipidemia LAUREATES: AWARDS AND HONORS SCS FALL MEETING 2004 72 CHIMIA 2005, 59, No. 3 Chimia 59 (2005) 72–76 © Schweizerische Chemische Gesellschaft ISSN 0009–4293 Oxidosqualene Cyclase (OSC) Inhibitors for the Treatment of Dyslipidemia Henrietta Dehmlowa§*, Jean Ackermanna, Johannes Aebia, Denise Blum-Kaelina, Alexander Chucholowskib, Philippe Coassoloa, Peter Hartmana, Manfred Kansya, Hans Peter Märkia, Olivier Morandc, Elisabeth Von der Marka, Narendra Pandaya, Armin Rufa, Ralf Thomaa, and Tanja Schulz-Gascha §Mettler Toledo Award Winner (Oral Presentation) Abstract: Novel inhibitors of oxidosqualene cyclase (OSC) for the treatment of dyslipidemia are reported. Starting point for the chemistry program was a set of compounds derived from a fungicide project which, in addition to high affinity for OSC from Candida albicans, also showed high affinity for the human enzyme (hOSC). Here the evaluation process of different scaffolds is outlined for two representative series, the phenyl substituted benzo[d]isothiazoles and the aminocyclohexanes. The most promising compounds derived from the latter series were further profiled in vivo and showed promising properties with respect to modulation of lipid parameters. Keywords: Dyslipidemia · Medicinal chemistry · Oxidosqualene cyclase · Structure-activity relationship Introduction observed [2][6], and the clinical benefit as- HMG-CoA reductase inhibition, should not sociated with the lowering of LDL-C levels lead to an undesired reduction of isoprenoid One of the most common cases of death in is now well established for both primary as intermediates of cholesterol synthesis and the western world is artherosclerotic car- well as secondary prevention [2][7][8]. their metabolites, e.g. coenzyme Q10 [9]. diovascular disease, accounting for over Current standard therapies for the phar- Further more, OSC inhibition has an indi- 30% of deaths [1]. In epidemiological and macologic treatment of lipid abnormalities rect, negative feed-back regulatory mecha- prospective studies several risk factors effectively modulate only one of the lipid nism [12] involving 24(S),25-epoxycholes- have been identified which promote the de- parameters. So the most widely used lip- terol which potentiates the primary inhibi- velopment of atherosclerosis [2–5]. These id-modifying drugs, statins (3-hydroxy-3- tory effect and reduces the accumulation of include, in addition to life habit risk factors methyl gluturyl coenzyme A (HMG-Co-A) mono-oxidosqualene in the liver. In addi- such as obesity, smoking and physical in- reductase inhibitors) very effectively reduce tion, potential OSC inhibitors could also activity, elevated plasma levels of low-den- plasma LDL-C by 30–45% at standard dose indirectly activate liver X receptor (LXR) sity lipoprotein cholesterol (LDL-C), low while affecting the plasma triglyceride level dependent pathways via 24(S),25-epoxy- plasma high-density lipoprotein cholesterol to a lesser extent. On the other hand, fibrates cholesterol, a known potent natural agonist (HDL-C), and elevated concentrations of (fibrate ester derivatives) reduce plasma lev- of the nuclear receptor LXR [13][14]. plasma triglyceride (TG)-rich lipoproteins. els of triglycerides more effectively, but not A strong correlation for the occurrence of LDL-C levels. Although statins are well tol- coronary heart disease (CHD) cases with erated at standard doses and adverse events Starting Point and Strategy LDL-C and TG levels in plasma has been are rare, side effects reported at high doses are elevated liver enzyme levels, myopathy A set of compounds derived from a fun- and rhabdomyolisis. gicide project [15][16] which in addition to In this context 2,3-oxidosqualene:lano- high affinity for OSC from Candida albi- sterol cyclase (OSC; E.C. 5.4.99.7) inhi- cans, also showed high affinity for hOSC bition, which would allow a concomitant [9][17] served as a starting point for the *Correspondence: Dr. H. Dehmlowa lowering of both LDL-C and TG levels, was chemistry program. Common structural fea- Tel.: + 41 61 688 9040 identified as a potential target to improve tures of all of these inhibitors are an amine Fax: + 41 61 688 9748 current therapies [9]. OSC, a microsomal residue and an electrophilic carbonyl-C- E-Mail: [email protected] aPharma Division membrane-associated enzyme consisting atom embedded in a benzophenone system. Preclinical Research of 732 aa with a size of 83.4 kDa, is loca- As neither an X-ray structure of hOSC [18] F. Hoffmann-La Roche Ltd ted down-stream of HMG-CoA reductase nor of related enzymes such as the prokary- CH–4070 Basel bChemBridge Research Laboratories, LLC (CRL) in the biosynthetic pathway of choleste- otic counterpart of OSC squalene hopene 16981 Via Tazon rol. It catalyzes the selective cyclization cyclase (SHC) [19][20] were known at the San Diego, CA 92127, USA of the linear substrate 2,3-oxidosqualene cActelion Pharmaceuticals Ltd start of the project, the optimization process Gewerbestrasse 16 (mono-oxidosqualene, MOS) to lanosterol was guided by a pharmacophore model. CH–4123 Allschwil [10][11]. Inhibition of OSC, in contrast to This model was derived by superimposing LAUREATES: AWARDS AND HONORS SCS FALL MEETING 2004 73 CHIMIA 2005, 59, No. 3 the structure of the folded, high energy con- former of mono-oxidosqualene (mimicking an opened protosterol) and 1 (Ro 48-8071) a c Amine --- Spacer --- --- Core as depicted in Fig. 1b. Later analysis of the F O X-ray of 1 in complex with hOSC support- ed this initial model. The essential interac- N tions of 1 with the protein are depicted in O Br Fig. 1c and 1d. The hydrogen bond between the nitrogen of the N,N-methylallylamine 1 (Ro 48-8071) moiety and the carboxylate of the catalytic acid Asp 455 demonstrates the importance of the basic amine residue of the inhibi- tors. The electron-deficient aromatic part b d of the benzophenone system is embedded in a pocket formed by Phe 521 and His 232. Furthermore, the carbonyl moiety of δ+ the benzophenone indirectly interacts with δ+ + Ile338 via a water molecule. δ The goal of the optimization was to iden- tify hOSC inhibitors with increased potency δ+ both in vitro and in vivo, increased metabol- δ+ ic stability and higher drug exposure in the target organs as compared to 1. As the car- bonyl moiety of the benzophenone residue was considered a potential site of metabo- lism [21] one approach consisted of replac- ing the benzophenone by other core struc- Fig. 1. a) 1 Ro 48-8071; b) Superimposed structures of the folded, high energy conformer of tures. Several alternative core structures oxidosqualene (mimicking the opened protosterol) and 1 (Ro 48-8071); c) Binding interactions with including heterocyclic derivatives such as the amine residue; d) Binding interactions with the benzophenone moiety phenyl substituted benzo[d]isothiazoles, benzisoxazoles, dioxo-benzo[d]isothiazole, benzo[b]thiophenes, indazoles, isoxazoles and indoles as well as substituted indolines, piperidines, anilines and aminocyclohex- anes were explored [17][22]. For most core scaffolds rapid evaluation of different spac- ers and amine substituents was possible by parallel synthesis. The process is depicted for two representative classes, the substi- tuted benzo[d]isothiazoles and aminocy- clohexane derivatives. Chemistry The synthesis for benzo[d]isothiazoles is outlined in Scheme 1 [17]. Starting from 3-fluoro-anisole (2), Friedel-Crafts acyla- tion with 4-bromo benzoyl chloride (3) in the presence of AlCl3 gave the desired benzophenone 4. Reaction of 4 with benzyl- mercaptane in the presence of potassium t-butylate yielded the thioether derivative 5 which upon treatment with sulfuryl chlo- ride and ammonia gave benzo[d]isothiazole (6) [23]. Subsequent cleavage of the meth- ylether with boron tribromide gave phenol 7, which was subjected to alkylation with an excess of α,ω-dibromoalkanes to yield the intermediates 8 that were converted to the final amines 9 by reaction with different Scheme 1. Synthesis of heteroaromatic compounds – benzo[d]isothiazole derivatives. Reagents and primary or secondary amines. conditions: a) AlCl3, PhNO2, 0 °C to rt, 42%; b) KOt-Bu, BnSH, THF, rt; c) SO2Cl2, CH2Cl2, rt; d) NH3, Furthermore, intermediates 8 could be EtOH, rt; e) BBr3, CH2Cl2, –78 °C to rt, 73% (from 4); f) α,ω-dibromoalkane, 10% aq. NaOH, N(Bu)4Br, 1 2 oxidized with KMnO4 to give the bromo- CH2Cl2, rt, 70%; g) NHR R , DMA, rt, 70–88%; (h) KMnO4/SiO2, CH2Cl2, rt, 80%. LAUREATES: AWARDS AND HONORS SCS FALL MEETING 2004 74 CHIMIA 2005, 59, No. 3 alkyloxy dioxo-benzo[d]isothiazoles 10 which were further converted to the final products 11 by amination. Analogous to compounds 9, the benzo[d]isothiazoles 13 were derived from benzophenone 12. For the aminocyclohexane derivatives with oxygen-linked spacers the synthesis is depicted in Scheme 2. The synthesis start- ed from trans-4-aminocyclohexanol (14) which was converted to the BOC-protected trans-4-methylaminocyclohexanol (15) by treatment with ZCl, reduction with LAH followed by BOC-protection. 15 was alkyl- ated under phase transfer conditions with α,ω-dihaloalkanes to yield the bromide 16. Alternatively, 15 was treated with α,ω-diha- loalkanes in the presence of NaH in DMF or especially for shorter spacer lengths with Scheme 2. Synthesis of aminocyclohexane derivatives with oxygen-linked spacers. Reagents and conditions: a) ZCl, Na2CO3, THF, H2O, rt, 98%; b) LAH, THF, 0 °C to rt, 80%; c) (BOC)2O, iPrOH, the in situ generated haloalkane-triflates 1 2 CH2Cl2, rt, 98%; d) α,ω-dibromoalkane, Bu4NHSO4, NaOH, H2O, rt, 76%; e) NHR R , DMA, rt, 92%; (from the corresponding haloalkanols with 3 3 f) TFA, CH2Cl2, rt, 91%; g) R SO2Cl, iPr2NEt, CH2Cl2, rt, 50–80%; h) R OCOCl, iPr2NEt, CH2Cl2, rt, trifluoromethansulfonic anhydride/2,6- 60–75%. di-tert-butylpyridine [24]). The reaction of bromide 16 with primary or secondary amines followed by cleavage of the BOC of the ester to the aldehyde 21 could be yielded the building block 24 as TFA salt.
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