© Copyright 2001 by the American Chemical Society
`
`Volume 44, Number 6
`
`March 15, 2001
`
`Expedited Articles
`
`A Novel Series of Highly Potent Benzimidazole-Based Microsomal Triglyceride
`Transfer Protein Inhibitors
`
`Jeffrey A. Robl,* Richard Sulsky, Chong-Qing Sun, Ligaya M. Simpkins, Tammy Wang, John K. Dickson, Jr.,
`Ying Chen, David R. Magnin, Prakash Taunk, William A. Slusarchyk, Scott A. Biller, Shih-Jung Lan,
`Fergal Connolly, Lori K. Kunselman, Talal Sabrah, Haris Jamil, David Gordon, Thomas W. Harrity, and
`John R. Wetterau
`The Bristol-Myers Squibb Pharmaceutical Research Institute, P.O. Box 5400, Princeton, New Jersey 08543-5400
`
`Received November 21, 2000
`
`A series of benzimidazole-based analogues of the potent MTP inhibitor BMS-201038 were
`discovered. Incorporation of an unsubstituted benzimidazole moiety in place of a piperidine
`group afforded potent inhibitors of MTP in vitro which were weakly active in vivo. Appropriate
`substitution on the benzimidazole ring, especially with small alkyl groups, led to dramatic
`increases in potency, both in a cellular assay of apoB secretion and especially in animal models
`of cholesterol lowering. The most potent in this series, 3g (BMS-212122), was significantly
`more potent than BMS-201038 in reducing plasma lipids (cholesterol, VLDL/LDL, TG) in both
`hamsters and cynomolgus monkeys.
`
`Introduction
`Despite major advances in pharmaceutical and surgi-
`cal treatments, coronary heart disease (CHD) remains
`the major cause of death in the industrialized world.1,2
`Elevated LDL cholesterol has been identified as a key
`risk factor for CHD, and statin therapy for hypercho-
`lesterolemia has been shown to lower the risk for CHD
`by approximately one-third.3 It remains to be deter-
`mined whether more aggressive LDL-lowering therapy,
`or impact on related lipid risk factors such as elevated
`triglycerides and low HDL, can lead to further reduc-
`tions in CHD risk. Currently, there has been an
`increased focus on the treatment of concomitant hyper-
`triglyceridemia, and statins generally have a modest
`impact on triglyceride levels.
`Microsomal triglyceride transfer protein (MTP) is a
`key factor in the assembly of VLDL, the direct precursor
`to LDL.4 In a recent publication, we outlined the design
`
`(609) 818-
`* To whom correspondence should be addressed. Tel:
`5048. Fax: (609) 818-3550. E-mail: Jeffrey.Robl@bms.com.
`
`and pharmacology of BMS-201038 (1), a potent inhibitor
`of MTP, and described its robust efficacy in lowering
`plasma cholesterol and triglycerides in both Golden
`Syrian hamsters and WHHL rabbits.5 Based on its
`efficacy and safety profile, BMS-201038 was advanced
`into clinical trials. Our focus was subsequently directed
`toward the identification of a suitable back-up clinical
`candidate to BMS-201038. A substantial amount of
`structure-activity studies (SAR) were performed incor-
`porating various carbocycles and heterocycles, generi-
`cally depicted by structure 2 (Chart 1), as replacements
`for the piperidine ring in BMS-201038. A driving force
`for this change was a concern with the ubiquitous
`presence of piperidine pharmacophores in ligands for
`G-protein coupled receptors. Within the chemical pro-
`gram, particular emphasis was placed on the incor-
`poration of basic functionalities which would aid in
`the solubility of these rather lipophilic molecules.
`Heteroaryl scaffolds were particularly attractive since
`additional substituents could readily be built upon these
`cores. Approximately 20 piperidine replacements were
`
`10.1021/jm000494a CCC: $20.00 © 2001 American Chemical Society
`Published on Web 02/21/2001
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`852 Journal of Medicinal Chemistry, 2001, Vol. 44, No. 6
`
`Robl et al.
`
`Chart 1
`
`Scheme 1a
`
`a (a) SeO2, 2.4 N HCl, 80 °C (95%); (b) HNO3, H2SO4, 10 °C (91%); (c) 57% HI, 50 °C (88%); (d) 2,4-pentanedione, 5 N HCl, EtOH,
`reflux, 1 h (R ) Me, 98%); 98% HCO2H, reflux (R ) H, 95-97%); n-C3H7C(OMe)3, PPTS, dioxane, reflux (R ) n-Pr, 77%); i-C3H7CO2H,
`4 N HCl, reflux (R ) i-Pr, 100%); (e) K2CO3, DMF, rt; (f) H2, Pd/C, EtOH, rt; (g) EDAC, TEA, CH2Cl2, rt (when Z ) OH) or TEA, CH2Cl2,
`0 °C (when Z ) Cl).
`
`explored in addition to changes within the linker group
`(A) and the terminal amide functionality. This manu-
`script highlights in vitro and in vivo SARs of a select
`group of compounds which incorporate a benzimidazole
`core as the central heteroaryl ring, resulting in an
`exceptionally potent series of novel MTP inhibitors.
`
`Chemistry
`
`Generation of the benzimidazole precursors and in-
`corporation within the BMS-201038 scaffold are de-
`picted in Scheme 1. In the case where W is hydrogen,
`the nitrodiamine 6 was obtained commercially. Where
`W was methyl, the requisite nitrodiamines were pre-
`pared following the procedures described by Grivas et
`al.6 For example, treatment of 4 with SeO2 in 3 N HCl
`afforded the corresponding selenocycle which underwent
`nitration to afford regioisomer 5 exclusively in good
`overall yield. Conversion to the diamine 6 (W ) Me) was
`effected by treatment with HI in concentrated HCl.
`Functionalized nitrobenzimidazoles 7 were easily pre-
`pared from the corresponding nitrodiamines 6 exploiting
`several methodologies: either by reacting the diamines
`
`with the corresponding acid under acidic conditions (as
`with formic or isobutyric when R ) H or i-Pr, respec-
`tively), by reaction with an orthoacetal (as when R )
`Pr), or by reaction with 2,4-pentanedione (as when R )
`Me). Alkylation of 7 with fluorenylamide 8 (readily
`prepared in two steps from 9-fluorenylcarboxylic acid)
`in DMF with K2CO3 as base often provided mixtures of
`regioisomers (e.g. 3.2:1 when R ) H, 7:1 when R ) Me)
`which could be readily separated by flash chromatog-
`raphy. In all cases, and especially with alkyl substitu-
`ents at R, the desired alkylation regioisomer 9 (as
`shown) was the predominant product. Simple catalytic
`hydrogenation provided amines 10 which were subse-
`quently acylated with 4¢-(trifluoromethyl)-2-biphenyl-
`carboxylic acid via EDAC activation or by reaction with
`the corresponding acyl chloride to afford the final
`products 3a-g.
`
`Results and Discussion
`A comparison of these compounds shows that potency
`in vitro, and especially in vivo, was critically dependent
`on both the length of the alkyl chain joining the fluo-
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`Benzimidazole-Based MTP Inhibitors
`
`Journal of Medicinal Chemistry, 2001, Vol. 44, No. 6 853
`
`Table 1. Inhibition of MTP in Vitro and the in Vivo Efficacy Responses in Lowering TC in Hamsters and Cynomolgus Monkeys for
`Compounds 1 and 3a-g
`
`n
`
`R
`
`W
`
`compda
`1
`
`MTP TG transfer
`assay: IC50 (nM)b
`8
`
`apoB secretion HepG2
`cells: IC50 (nM)b
`0.8
`
`TC lowering in hamsters:
`% @ mg/kg or ED50 (mg/kg)c
`2.3
`
`3a
`3b
`
`3c
`3d
`
`3e
`3f
`3g
`
`0 H
`1 H
`
`1 H
`1 Me
`
`H
`H
`
`Me
`H
`
`H
`i-Pr
`1
`n-Pr H
`1
`1 Me
`Me
`
`10
`5
`
`1
`1
`
`4
`2
`1
`
`nde
`0.24
`
`0.02
`0.15
`
`0.06
`nd
`0.03
`
`TC lowering in monkeys:
`% @ mg/kg or ED50 (mg/kg)d
`-88% @ 10
`-74% @ 5
`-65% @ 2.5
`-35% @ 1.25
`ED50 ) 2 mg/kg
`nd
`-52% @ 5
`-19% @ 2.5
`-68% @ 2.5
`-80% @ 5
`-51% @ 1.25
`-39% @ 2.5
`nd
`-83% @ 1.25
`-76% @ 1
`-43% @ 0.3
`-28% @ 0.1
`ED50 ) 0.38 mg/kg
`a All spectral data were consistent with the assigned structures. b The TG transfer and apoB secretion assays were performed as described
`in ref 6. c ED50 represents dose (mg/kg) required for 50% lowering of plasma TC in standard-diet-fed Golden Syrian hamsters 16 h postdosing
`or percent TC lowering at dose (mg/kg) (compound administered once daily orally for 3 days). d ED50 represents dose (mg/kg) required for
`50% lowering of plasma TC in standard-diet-fed cynomolgus monkeys 16 h postdosing or percent TC lowering at dose (mg/kg) (compound
`administered once daily orally for 7 days). e nd, not determined.
`renylbenzimidazole moieties as well as the substituents
`on the benzimidazole ring (Table 1). Compounds were
`evaluated for inhibition of human MTP activity in vitro
`(triglyceride transfer assay) and in HepG2 cells (apoB
`secretion assay). In vivo activity was determined in both
`hamsters (3 days, po, qd, standard diet) and cynomolgus
`monkeys (7 days, po, qd, standard diet) as compared to
`the clinical agent BMS-201038 (1). Direct replacement
`of the piperidine moiety in 1 with an unsubstituted
`benzimidazole group, affording 3b, resulted in a slight
`enhancement of in vitro activity in both the triglyceride
`transfer and HepG2 apoB secretion assays. Unfortu-
`nately, this modification attenuated the ability of 3b to
`lower cholesterol in both the rodent and primate models.
`Shortening the linker group to three carbons to afford
`3a essentially abolished activity in the hamster, despite
`reasonable activity in the transfer assay. Significant
`enhancements in potency were realized by incorporation
`of small substituents at the R and/or W positions of
`the benzimidazole ring. Due to sensitivity limitations
`of the transfer assay, the secretion assay was primarily
`utilized to differentiate among the most potent com-
`pounds (3c-g). Compounds 3c-g were 5-40-fold more
`active than 1 in the apoB secretion assay, which on the
`basis of our experience usually translates to greater
`potency in vivo. Although larger alkyl substituents at
`R (isopropyl and n-propyl, 3e,f, respectively) diminished
`cholesterol lowering in the hamster and monkey as
`compared to piperidine 1, substitution with methyl (3d)
`significantly enhanced in vivo potency as compared to
`3b. A similar boost in potency was observed by the
`addition of methyl at W (3c). Remarkably, incorporation
`of methyls at both R and W to give 3g resulted in a
`synergistic effect in vivo, affording a 43- and 13-fold
`enhancement in potency in the hamster and monkey,
`respectively, over 3b.
`The significant potency of 3g in these models war-
`ranted detailed evaluation of this compound in vivo. The
`dose-response for 3g upon 3-day treatment in standard
`chow-fed hamsters (oral administration once daily) is
`depicted in Figure 1. At the lowest dose tested (0.1 mg/
`
`-11% @ 15
`12
`
`3.8
`2.5
`
`-60% @ 8
`-44% @ 15
`0.28
`
`Figure 1. Effect of 3 days of treatment with 3g on hamster
`plasma lipids. Golden Syrian hamsters were fed a standard
`diet (Purina 5001, which contains 0.02% TC and 4.5% TG)
`during the course of once daily oral treatment with 3g. Fasting
`plasma lipid levels were determined 16 h after the last dose.
`The percent decreases were determined by comparing the lipid
`levels of drug-treated animals to those of the control-treated
`animals. Error bars indicate SEM; p values: *<0.1, **<0.01,
`***<0.001.
`
`kg), administration of 3g resulted in a 37%, 37%, 51%,
`and 18% decrease in total cholesterol (TC), triglycerides
`(TG), VLDL/LDL, and HDL, respectively, as compared
`to vehicle controls. Significantly greater efficacy was
`observed at the highest dose tested (1 mg/kg). The HDL
`decrease was unlikely due to a direct effect on HDL
`production because the in vitro IC50 for the inhibition
`of secretion of apolipoprotein AI with 3g in HepG2 cells
`was found to be >500 (cid:237)M (highest concentration tested).
`No elevation of liver function tests (AST and ALT) were
`detected at any dose. An equally dramatic effect was
`observed in cynomolgus monkeys after once daily oral
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`854 Journal of Medicinal Chemistry, 2001, Vol. 44, No. 6
`
`Robl et al.
`
`kinetic testing of 3g in rats showed this compound
`possessed good oral bioavailability (81%) with a terminal
`elimination half-life of 4.8 h. In cynomolgus monkeys,
`oral bioavailability was somewhat reduced (23%) but
`half-life (6.9 h) was slightly greater.
`
`Conclusion
`We have demonstrated that the piperidine of BMS-
`201038 can be replaced by a benzimidazole ring system
`to provide a novel and highly potent class of MTP
`inhibitors. Molecular overlays indicate that the benz-
`imidazole system provides a similar disposition of the
`flanking fluorenyl- and biphenylamide groups, suggest-
`ing that the heterocycle serves as a central scaffold. The
`enhanced potency of the benzimidazole-based inhibitors
`with small substituents at R and W suggests that
`orienting the biphenylamide at a position orthogonal to
`the central ring is desirable for optimal activity.
`Currently, statin therapy is the first-line treatment
`for patients with hyperlipidemia and elevated choles-
`terol levels, although their effects on TG lowering is
`modest and variable. Preclinically, MTP inhibitors have
`dramatic effects not only on plasma cholesterol and LDL
`levels but on TG levels as well, offering the potential
`for greater efficacy and plasma lipid control in both
`hypertriglyceridemia and mixed hyperlipidemia. Com-
`pound 3g has demonstrated superior potency both in
`vitro and especially in vivo as compared to the clinical
`agent BMS-201038. Indeed, it is the most potent MTP
`inhibitor described to date. On this basis, as well as its
`acceptable pharmacokinetic and safety profile, 3g (BMS-
`212122) was brought forward for development as a
`potentially superior back-up agent to BMS-201038.
`
`Experimental Section
`All reactions were carried out under a static atmosphere of
`argon and stirred magnetically unless otherwise noted. All
`reagents used were of commercial quality and were obtained
`from Aldrich Chemical Co. or Sigma Chemical Co. Melting
`points were obtained on a Hoover Uni-melt melting point
`apparatus and are uncorrected. Infrared spectra were recorded
`on a Mattson Sirius 100-FTIR spectrophotometer. 1H (400 Mz)
`and 13C (100 Mz) NMR spectra were recorded on a JEOL
`GSX400 spectrometer using Me4Si as an internal standard.
`All flash chromatographic separations were performed using
`E. Merck silica gel 60 (particle size, 0.040-0.063 mm). Reac-
`tions were monitored by TLC using 0.25-mm E. Merck silica
`gel plates (60 F254) and were visualized with UV light or 5%
`phosphomolybdic acid in 95% EtOH. BMS-201038 was syn-
`thesized at the Bristol-Myers Squibb Pharmaceutical Research
`Institute (Princeton, NJ).
`5-Methyl-4-nitro-2,1,3-benzoselenadiazole (5). To a
`stirred solution of 48.95 g (0.40 mol) of 4 in 500 mL of 2.4 M
`hydrochloric acid at 80 °C under argon was added a warm
`solution of 88.77 g (0.80 mol) of selenium dioxide in 300 mL of
`water dropwise over the course of 30 min. After an additional
`90 min, the reaction was cooled to room temperature and the
`solids were collected, washing with water. The brown solids
`were dried in vacuo at 50 °C to give the intermediate
`selenocycle (75.10 g) in 95% yield: TLC Rf 0.59 (CH2Cl2); mp
`67-69 °C; 1H NMR (DMSO-d6) (cid:228) 2.39 (s, 3H), 7.35 (dd, 1H),
`7.57 (s, 1H), 7.69 (d, 1H); 13C NMR (DMSO-d6) (cid:228) 21.13, 120.96,
`122.38, 132.54, 139.61, 158.72, 160.14. To a stirred solution
`of 72.00 g (0.365 mol) of the intermediate in 180 mL of 98%
`sulfuric acid at 10 °C was added a cold solution of 108 mL of
`2:1 98% sulfuric acid/70% nitric acid over 1 h. The temperature
`of the reaction mixture was not allowed to rise above 20 °C.
`After an additional 60 min, the reaction was poured as a thin
`stream into 750 g of ice with rapid stirring. The fine yellow
`
`Figure 2. Plasma TC and TG concentrations in cynomolgus
`monkeys treated orally once daily with 3g for 7 days. Fasting
`plasma lipid levels were measured 16 h after the last dose (0.1,
`0.3, or 1 mg/kg/day) of 3g. The percent decreases were
`determined by comparing the lipid levels of drug-treated
`animals to those of the control-treated animals. Error bars
`indicate SEM; p values: **<0.01, ***<0.001.
`
`treatment for 7 days (Figure 2). At the highest dose
`tested (1 mg/kg), TC, TG, VLDL/LDL, and HDL were
`lowered 73%, 71%, 84%, and 68%, respectively. Toler-
`ability was good in this study, but an approximate 2-fold
`increase in plasma ALT, AST, and CPK levels was
`observed. The significance on the modest increases in
`liver function enyzmes in the monkey is unclear in light
`of the magnitude of the lipid-lowering response. Other
`hypolipidemics, such as the stains, have shown a similar
`effect in humans.
`To confirm the decrease in plasma lipids is due to an
`inhibition of lipoprotein production, as would be antici-
`pated for an MTP inhibitor, a dose-response study of
`3g was performed in the fasted Triton rat assay, which
`measures the effect on hepatic apoB lipoprotein produc-
`tion.7 Intravenous injection of Triton WR1339, a non-
`ionic surfactant, has been shown to block the clearance
`of plasma TG-rich lipoproteins, enabling the measure-
`ment of their production in vivo. To ascertain the effects
`of MTP inhibition on TG secretion rates, Sprague-
`Dawley rats were fasted overnight for 18 h and orally
`dosed with 3g 1 h before intravenous injection of Triton
`WR1339 (250 mg/kg). The secretion rate was deter-
`mined by calculating the amount of TG accumulated
`during the first 2.5 h after the Triton injection. In this
`assay, 3g was equipotent to 1 upon oral administration
`(ED50 ) 0.15 mg/kg) but was nearly 4-fold more potent
`when given intravenously (ED50 ) 0.04 mg/kg). At a
`dose of 1 mg/kg (po) and 0.3 mg/kg (iv), TG secretion
`rate was inhibited 94% and (cid:24)100%, respectively, thus
`demonstrating that 3g is a potent inhibitor of hepatic
`lipoprotein production in vivo.
`Based on the excellent in vivo profile of 3g in models
`of lipid lowering, this compound was further evaluated
`for its pharmacokinetic properties. Standard pharmaco-
`
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`Benzimidazole-Based MTP Inhibitors
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`Journal of Medicinal Chemistry, 2001, Vol. 44, No. 6 855
`
`slurry was filtered and the collected solids were washed five
`times with 200-mL portions of cold water. The moist cake was
`heated in 500 mL of ethanol to near boiling and then cooled
`to room temperature and the solid collected. Drying in vacuo
`at 50 °C gave 5 as a yellow solid (80.70 g) in 91% yield: TLC
`Rf 0.47 (CH2Cl2); mp 190-192 °C; 1H NMR (DMSO-d6) (cid:228) 2.46
`(s, 3H), 7.62 (d, 1H), 8.01 (d, 1H); 13C NMR (DMSO-d6) (cid:228) 16.98,
`125.44, 131.59, 131.88, 141.65, 150.59, 158.21. Anal. Calcd for
`C7H5N3O2Se: C, 34.73; H, 2.08; N, 17.36; Se, 32.61. Found:
`C, 34.96; H, 1.97; N, 17.35; Se, 32.59.
`4-Methyl-3-nitro-1,2-benzenediamine (6, W ) Me). To
`a stirred solution of hydriodic acid (25.0 mL, 57%, 189 mmol,
`stabilized with 1.5% hypophosphorous acid) at room temper-
`ature in argon was added 5.00 g (20.7 mmol) of 5. The reaction
`vessel was placed in an oil bath preheated to 50 °C and the
`resulting deep red solution was vigorously stirred for 2 h. After
`cooling to room temperature the reaction mixture was poured
`into a stirred slurry of 24 g (0.2 mol) of sodium hydrogen sulfite
`in 50 mL of water. The resulting light yellow slurry was
`treated with an ice-cold solution of sodium hydroxide (7.5 g,
`188 mmol) in 50 mL of water. Additional 6 M sodium hydroxide
`was added until the aqueous slurry was brought to pH 8. The
`resulting deep red slurry was filtered and the filtrate extracted
`three times with 200-mL portions of chloroform. The solids
`from the filtration were dissolved in 300 mL of chloroform and
`washed once with 50 mL of water. The organic extracts were
`combined, dried (Na2SO4) and evaporated to give 6 as a deep
`red solid (3.04 g) in 88% yield: TLC Rf 0.38 (1:49 Et2O:CH2-
`Cl2); mp 132-133 °C; 1H NMR (CDCl3) (cid:228) 2.38 (s, 3H), 3.35 (br
`s, 2H), 4.92 (br s, 2H), 6.49 (d, 1H), 6.74 (d, 1H); 13C NMR
`(CDCl3) (cid:228) 20.14, 120.11, 120.27, 125.71, 133.34, 133.46.
`2,5-Dimethyl-4-nitro-1H-benzimidazole (7, W ) R ) Me).
`To a refluxing solution of 1.00 g (6.00 mmol) of 6 in 27 mL of
`ethanol and 7.2 mL of 5 M hydrochloric acid under argon was
`added 1.20 g (12.0 mmol) of 2,4-pentanedione over the course
`of 5 min. After an additional 60 min at reflux, the reaction
`was cooled and partially evaporated to remove ethanol. The
`residue was treated with saturated NaHCO3 solution to pH
`7, filtered, washed with water and dried in vacuo at40 °C to
`give 7 (R ) W ) Me) as a tan solid (1.12 g) in 98% yield: TLC
`Rf 0.20 (EtOAc); mp 232-234 °C; 1H NMR (CDCl3) (cid:228) 2.70 (s,
`3H), 2.83 (s, 3H), 7.19 (d, 1H), 7.84, (d, 1H); 13C NMR (CDCl3)
`(cid:228) 15.08, 22.13, 97.13, 104.91, 125.34, 126.26, 131.41, 152.68.
`9-(4-Bromobutyl)-N-(2,2,2-trifluoroethyl)-9H-fluorene-
`9-carboxamide (8, n ) 1). To a solution of 9-fluorene-
`carboxylic acid (50 g, 240 mmol) in THF (1200 mL) at 0 °C
`was added dropwise a solution of n-butyllithium (2.5 M, 211
`mL, 530 mmol) in THF. The yellow reaction was stirred at 0
`°C for 1 h, then 1,4-dibromobutane (31.3 mL, 260 mmol) was
`added dropwise over 30 min. The reaction was stirred at 0 °C
`for 30 min, then warmed to room temperature for 30 h. The
`reaction was extracted with water (3 (cid:2) 750 mL). The combined
`aqueous layers were extracted with ethyl ether (800 mL). The
`aqueous layer was made acidic with HCl solution (1 N, 500
`mL), then extracted with dichloromethane (3 (cid:2) 750 mL). The
`combined organic layers were dried over MgSO4. Evaporation
`gave the corresponding C-9 alkylated acid (71 g, 85%) as a
`white solid. To a solution of the intermediate acid (60 g, 173
`mmol) and DMF (100 (cid:237)L) in CH2Cl2 (600 mL) under argon at
`0 °C was added oxalyl chloride (104 mL, 2.0 M in CH2Cl2, 208
`mmol) dropwise. The reaction was stirred at 0 °C for 10 min,
`then warmed to room temperature and stirred for 1.5 h. The
`reaction was concentrated in vacuo to give the crude acid
`chloride as a yellow oil. To a suspension of 2,2,2-trifluoroethyl-
`amine hydrochloride (25.9 g, 191 mmol) in CH2Cl2 (500 mL)
`at 0 °C under argon was added triethylamine (73 mL, 521
`mmol) followed by dropwise addition of a solution of the crude
`acid chloride in CH2Cl2 (15 mL). The reaction was stirred at
`0 °C for 1 h, diluted with CH2Cl2 (500 mL), and washed with
`water (2 (cid:2) 300 mL), 1 N HCl (2 (cid:2) 300 mL), saturated NaHCO3
`(2 (cid:2) 300 mL), and brine (2 (cid:2) 300 mL), then dried over MgSO4.
`Evaporation gave 80 g of a oil which was purified by flash
`chromatography on silica gel (2.5 kg). The crude product was
`loaded in a mixture of CH2Cl2 and hexane, and eluted with a
`
`step gradient of 10% EtOAc/hexane (4 L) to 15% EtOAc/hexane
`(2 L) to 20% EtOAc/hexane (4 L). Pure fractions were combined
`and evaporated to give 8 (n ) 1) (52.5 g, 71%) as a white
`solid: TLC Rf 0.32 (2:8 EtOAc:hexanes); mp 88-92 °C; 1H
`NMR (CDCl3) (cid:228) 0.83 (m, 2H), 1.68 (m, 2H), 2.42 (m, 2H), 3.21
`(t, 2H), 3.68 (m, 2H), 5.36 (t, 1H), 7.36-7.59 (m, 6H), 7.78 (d,
`2H); 13C NMR (CDCl3) (cid:228) 22.27, 32.70, 32.99, 40.65 (q), 62.17,
`120.51, 124.07, 128.21, 128.71, 140.96, 144.88, 173.29.
`9-[4-(2,5-Dimethyl-4-nitro-1H-benzimidazol-1-yl)butyl]-
`N-(2,2,2-trifluoroethyl)-9H-fluorene-9-carboxamide (9g,
`W ) R ) Me, n ) 1). To a stirred slurry of 1.80 g of 8 (9.41
`mmol) in 15 mL of DMF at room temperature under argon
`was added 1.75 g (33 mmol) of potassium carbonate. After 1
`h, 4.26 g (10.0 mmol) of 7 was added and the reaction stirred
`for 86 h. The reaction mixture was quenched with 30 mL of
`water. The liquids were decanted away from the formed
`gummy solid, which was then washed with water. The semi-
`solid residue was triturated with 40 mL of ether. The resulting
`granular solid was chilled and filtered. The collected solid cake
`was washed with water, transferred to a round-bottom flask
`and evaporated from toluene. The dried residual solids were
`triturated with hot ethyl acetate and filtered to give 4.02 g of
`9g (80%) as a white solid: TLC Rf 0.20 (3:17 Et2O:CH2Cl2);
`mp 181-183 °C; 1H NMR (CDCl3) (cid:228) 0.77 (m, 2H), 1.52 (m, 2H),
`2.25 (s, 3H), 2.41 (m, 2H), 2.47 (s, 3H), 3.71 (m, 2H), 3.82 (t,
`2H), 5.91 (t, 1H), 7.00 (d, 1H), 7.11 (d, 1H), 7.26 (t, 2H), 7.32
`(t, 2H), 7.46 (d, 2H), 7.62 (d, 2H); 13C NMR (CDCl3) (cid:228) 13.20,
`17.74, 20.94, 29.32, 35.49, 40.26 (q), 43.37, 61.65, 111.94,
`119.94, 123.63 (q), 123.75, 124.00, 124.19, 127.72, 128.23,
`134.84, 135.24, 139.61, 140.45, 144.42, 153.64, 172.86.
`9-[4-(2,5-Dimethyl-4-amino-1H-benzimidazol-1-yl)butyl]-
`N-(2,2,2-trifluoroethyl)-9H-fluorene-9-carboxamide (10g,
`W ) R ) Me, n ) 1). A stirred slurry of 1.05 g (1.96 mmol) of
`9g and 200 mg of 10% palladium-on-charcoal in 40 mL of
`ethanol was purged with argon and evacuated three times.
`Hydrogen was introduced to the partially evacuated solution
`via a balloon. After 14 h, the reaction mixture was purged with
`argon and passed through a 0.45-(cid:237)m nylon filter, washing with
`dichloromethane. The filtrate was evaporated and then re-
`evaporated twice from dichloromethane to give 10g as a white
`foam (0.958 g) in 99% yield: TLC Rf 0.13 (EtOAc); 1H NMR
`(CDCl3) (cid:228) 0.80 (m, 2H), 1.53 (m, 2H), 2.19 (s, 3H), 2.27 (s, 3H),
`2.42 (m, 2H), 3.71 (m, 4H), 4.06 (br s, 2H), 5.66 (t, 1H), 6.40
`(d, 1H), 6.82 (d, 1H), 7.40 (m, 4H), 7.48 (d, 2H), 7.69 (d, 2H);
`13C NMR (CDCl3) (cid:228) 13.31, 16.41, 21.27, 29.46, 35.77, 40.49 (q),
`43.32, 62.00, 98.31, 112.79, 120.30, 123.72 (q), 124.05, 124.48,
`128.04, 128.54, 131.27, 133.77, 135.30, 140.76, 144.75, 148.62,
`173.11.
`9-[4-[2,5-Dimethyl-4-[[[4¢ -(trifluoromethyl)[1,1¢ -biphen-
`yl]-2-yl]carbonyl]amino]-1H-benzimidazol-1-yl]butyl]-N-
`(2,2,2-trifluoroethyl)-9H-fluorene-9-carboxamide (3g).
`To a slurry of 1.72 g (6.47 mmol) of 4¢-(trifluoromethyl)-2-
`biphenylcarboxylic acid (11, Z ) OH), in 15 mL of dichloro-
`methane at room temperature was added 0.85 mL (9.74 mmol)
`of oxalyl chloride followed by 75 (cid:237)L of DMF. After 1 h, the
`resulting solution was evaporated and re-evaporated from
`dichloromethane. The residue was dissolved in 10 mL of
`dichloromethane and the solution was added dropwise to a
`solution of 3.21 g (6.34 mmol) of 10g and 1.00 mL (7.17 mmol)
`of triethylamine in 15 mL of dichloromethane at room tem-
`perature. After 90 min, the reaction mixture was diluted with
`100 mL of ethyl acetate and washed once with saturated
`sodium bicarbonate solution. The organic phase was dried
`(MgSO4) and evaporated. Purification by flash chromatography
`on silica gel (5 (cid:2) 25-cm column, ethyl acetate) gave, after
`recrystallization from dichloromethane/hexanes, 3g as a white
`crystalline solid (3.86 g) in 81% yield: TLC Rf 0.34 (EtOAc);
`mp 127-129 °C; 1H NMR (CDCl3) (cid:228) 0.81 (m, 2H), 1.56 (m, 2H),
`2.26 (s, 3H), 2.36 (s, 3H), 2.42 (m, 2H), 3.69 (dq, 2H), 3.81 (t,
`2H), 5.36 (t, 1H), 6.89 (d, 1H), 6.99 (d, 1H), 7.35 (m, 1H), 7.41
`(m, 2H), 7.44 (m, 2H), 7.49 (d, 4H), 7.53 (dd, 1H), 7.59 (d, 1H),
`7.69 (t, 3H), 7.75 (d, 2H), 7.85 (dd, 1H); 13C NMR (CDCl3) (cid:228)
`13.53, 18.43, 21.35, 29.61, 35.76, 40.57 (q), 43.56, 62.02, 107.43,
`120.44, 123.72 (d) 124.77, 124.92 (q), 124.99 (d) 125.24, 127.45,
`
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`856 Journal of Medicinal Chemistry, 2001, Vol. 44, No. 6
`
`128.00, 128.17, 128.71, 129.27, 129.34 (d), 130.25, 130.38,
`136.14, 133.69, 136.14, 137.84, 138.95, 140.79, 144.03, 144.66,
`150.66, 167.79, 173.12. Anal. Calcd for C43H36F6N4O2(cid:226)CH2Cl2:
`C, 63.46; H, 4.50; N, 6.58; F, 13.38. Found: C, 63.28; H, 4.30;
`N, 6.56; F, 13.10.
`Supporting Information Available: Melting point
`and HRMS data for compounds 3a-g, 5, 6, and 8. This
`material is available free of charge via the Internet at http://
`pubs.acs.org.
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