Identification of Selective Acyl Sulfonamide ... - ACS Publications

Article Cite This: J. Med. Chem. 2019, 62, 908−927

pubs.acs.org/jmc

J. Med. Chem. 2019.62. Downloaded from pubs.acs.org by EASTERN KENTUCKY UNIV on 01/27/19. For personal use only.

Identification of Selective Acyl Sulfonamide−Cycloalkylether Inhibitors of the Voltage-Gated Sodium Channel (NaV) 1.7 with Potent Analgesic Activity Shaoyi Sun,† Qi Jia,† Alla Y. Zenova,† Michael S. Wilson,† Sultan Chowdhury,†,∥ Thilo Focken,† Jun Li,‡ Shannon Decker,† Michael E. Grimwood,† Jean-Christophe Andrez,† Ivan Hemeon,† Tao Sheng,†,⊥ Chien-An Chen,§,# Andy White,§,∇ David H. Hackos,‡ Lunbin Deng,‡ Girish Bankar,† Kuldip Khakh,† Elaine Chang,† Rainbow Kwan,† Sophia Lin,† Karen Nelkenbrecher,† Benjamin D. Sellers,‡ Antonio G. DiPasquale,‡ Jae Chang,‡ Jodie Pang,‡ Luis Sojo,† Andrea Lindgren,† Matthew Waldbrook,† Zhiwei Xie,† Clint Young,† James P. Johnson,† C. Lee Robinette,† Charles J. Cohen,† Brian S. Safina,‡ Daniel P. Sutherlin,‡ Daniel F. Ortwine,*,‡ and Christoph M. Dehnhardt*,† †

Xenon Pharmaceuticals Inc., 200-3650 Gilmore Way, Burnaby, British Columbia V5G 4W8, Canada Genentech Inc., 1 DNA Way, South San Francisco, California 94080-4990, United States § ChemPartner, Building No. 5, 998 Halei Road, Zhangjiang Hi-Tech Park, Pudong New Area, Shanghai 201203, P. R. China ‡

S Supporting Information *

ABSTRACT: Herein, we report the discovery and optimization of a series of orally bioavailable acyl sulfonamide NaV1.7 inhibitors that are selective for NaV1.7 over NaV1.5 and highly efficacious in in vivo models of pain and hNaV1.7 target engagement. An analysis of the physicochemical properties of literature NaV1.7 inhibitors suggested that acyl sulfonamides with high fsp3 could overcome some of the pharmacokinetic (PK) and efficacy challenges seen with existing series. Parallel library syntheses lead to the identification of analogue 7, which exhibited moderate potency against NaV1.7 and an acceptable PK profile in rodents, but relatively poor stability in human liver microsomes. Further, design strategy then focused on the optimization of potency against hNaV1.7 and improvement of human metabolic stability, utilizing induced fit docking in our previously disclosed X-ray cocrystal of the NaV1.7 voltage sensing domain. These investigations culminated in the discovery of tool compound 33, one of the most potent and efficacious NaV1.7 inhibitors reported to date.

■

INTRODUCTION Drugs that block voltage-gated sodium channels (NaVs) are widely used therapeutically as antiarrhythmics, anticonvulsants, anesthetics, and analgesics. Non-subtype-selective sodium channel blockers such as mexiletine and carbamazepine have been used for the treatment of neuropathic pain conditions. Unfortunately, this class of drugs is known to exhibit doselimiting side effects in patients, such as somnolence, dizziness, and nausea, that limit their utility as analgesic agents.1,2 These adverse effects are thought to be derived in large part from a lack of selectivity on the NaV family of channels. Nine NaV isoforms are known (NaV 1.1−NaV 1.9), and there are indications3 that isoform selective blockers may have an improved therapeutic index compared to nonselective compounds presently in use. In particular, NaV1.7 is a highly validated target for the treatment of pain conditions based on human genetic studies.4 Congenital insensitivity to pain (CIP), caused by a genetic deficiency of the SCN9A gene encoding © 2018 American Chemical Society

NaV1.7, is a rare autosomal recessive disorder characterized by the complete absence of pain sensation typically associated with noxious stimuli.5−10 Individuals with CIP show no deficits in motor or cognitive function.11 Conversely, gain-of-function mutations in SCN9A result in painful conditions, including inherited erythromelalgia (IEM),12−15 paroxysmal extreme pain disorder,16 and small fiber neuropathies.17 These findings indicate that NaV1.7 has an essential role in mediating pain signaling in humans and suggest that selective NaV1.7 antagonism may be a highly effective strategy to treat painful conditions.4 NaV1.5 is the predominant sodium channel in cardiac myocytes and has been identified as an important off-target to avoid in NaV1.7 drug discovery. Inhibition of NaV1.5 has been demonstrated to lead to cardiac arrhythmias.18 As a Received: October 18, 2018 Published: November 30, 2018 908

DOI: 10.1021/acs.jmedchem.8b01621 J. Med. Chem. 2019, 62, 908−927

Journal of Medicinal Chemistry

Article

Figure 1. Structures of selected subtype-selective NaV1.7 inhibitors.

Figure 2. Acyl sulfonamides generally occupy a more favorable physicochemical property space (average cLogP of 3.0 vs 4.1 and MW of 400 vs 490 for acyl vs aryl sulfonamides) for achieving oral bioavailability.

sulfonamides.22a This report did not contain a characterization of the analogues in in vivo models of pain. We and others21b,22a found that for many series, large multiples of NaV1.7 halfmaximal inhibitory concentration (IC50) values are needed to achieve acceptable efficacy in in vivo models. We have recently reported in vivo studies on a class of acyl sulfonamides37 that showed efficacy at low NaV1.7 IC50 multiples. However, the design and optimization of compounds that are highly active in in vivo assays has so far been elusive, which prompted the current study on acyl sulfonamide optimization that focused on improving in vivo efficacy. An analysis of the patent landscape through 2013 for aryl and acyl sulfonamide NaV1.7 inhibitors27 guided our start of the program. We noticed that acyl sulfonamides occupy a more favorable physicochemical property space such as lower molecular weight (MW) and cLogP (Figure 2). For example, aryl sulfonamides displayed an average cLogP of 4.1 and an average MW of 490, while acyl sulfonamides showed an average cLogP of 3.0 and an average MW of 400. Given that optimization very often involves increases in molecular weight and cLogP, acyl sulfonamides represented an attractive series for progression within Lipinski property space, which was an

consequence, our optimization campaign focused on establishing selectivity for NaV1.7 over NaV1.5. Due to a high sequence homology among NaV isoforms, compounds that are highly specific for NaV1.7 have been difficult to identify despite considerable efforts by the pharmaceutical industry. Recently, subtype-selective small-molecule NaV1.7 inhibitors have been reported.19−25 An example is aryl sulfonamide 1 (PF05089771) (Figure 1), which selectively (>600-fold) blocks NaV1.7 over NaV1.5 and was advanced into clinical trials.20a,b,d Our earlier work described the discovery of two structurally distinct series of aryl sulfonamide NaV1.7 inhibitors exemplified by 221a and 3,21b as well as the X-ray structure of the binding site on hNaV1.7.26 We have also recently disclosed a novel class of triazolyl sulfonamide NaV1.7 inhibitors, exemplified by 4.21c Although potent on NaV1.7 and highly selective against NaV1.5, aryl sulfonamides such as 321b displayed poor absorption, distribution, metabolism, and excretion properties. Therefore, we sought to identify alternative chemical series to mitigate these liabilities. A literature survey on selective NaV1.7 inhibitors revealed a primary focus on aryl sulfonamides, with only a single paper on a companion series of acyl 909



Article

Table 1. In Vitro Activity and Pharmacokinetic Properties of Compound 7

[3H]GX-545 LBA IC50 (μM) hNaV1.7 EP IC50 (μM) hNaV1.5 EP IC50 (μM)/sel. h1.5/1.7 LM CLhep (ml/min/kg) (human/mouse) MDCKI Papp (A to B) PPB (%)h/m mouse IV PK (1 mg/kg) mouse PO PK (30 mg/kg)

0.07 0.006 1.9/340 19/28 13 × 10−6 cm/s 99.9/99.9 CL: 9.2 ml/min/kg, Vss: 1.2 l/kg, t1/2: 1.7 h AUC: 240 μMh, t1/2: 3.5 h, Cmax: 45 μM, F: 100%

Figure 3. Induced fit docking of 7 into VSD4 of the hNaV1.7 receptor, shown in cyan. The active site cavity and helices are from the induced fit version. Key residues and distances are shown. The cocrystal structure of GX-936 and VSD4 of NaV1.726 (PDB code 5EK0) is superimposed and shown in orange.

important criterion for us at the start of the program.28 Additionally, acyl sulfonamides represented a less explored chemical series compared to aryl sulfonamides,20a,22a providing an opportunity to deepen our understanding of this class of compounds. Known acyl sulfonamides such as 5 (PF-05241328)20a or 22a 6 have a highly planar architecture due to the presence of multiple aromatic rings. The number of aromatic rings in a molecule has been shown to negatively affect aqueous solubility and has been associated with an increased attrition risk in drug development,29 suggesting a need for the identification of acyl sulfonamides with improved druglike properties. Our strategy to increase favorable properties focused partly on increasing fsp3 (fraction of sp3 carbons relative to the total carbon count).30 These efforts culminated in 33, a potent, molecularly selective, metabolically stable NaV1.7 inhibitor that demonstrated efficacy in various models of pain.

Effects in cells were assessed by whole-cell patch voltage clamp electrophysiology (EP) measurements of sodium currents mediated by human hNaV1.7 and hNaV1.5 heterologously expressed in HEK293 cells.26 The LBA was used as a highthroughput assay to guide design and to establish binding to a preferred site in voltage sensing domain 4 (VSD4) of NaV1.7.26 The functional potency data obtained by EP are biologically more relevant as they track the movement of the channel in a cell but are associated with a larger measurement error.31 Given these assay characteristics, we were most interested in compounds that showed high potencies in both assays. Acyl sulfonamide 7,21c,32 possessing a pendant adamantyl group, was discovered in early efforts to explore ether-linked saturated rings. This compound inhibited hNaV1.7 with an IC50 of 0.07 μM in the LBA (Table 1). A voltage clamp assay confirmed the ligand binding result (IC50 = 0.006 μM) and indicated that 7 was highly selective against hNaV1.5 (IC50 = 1.9 μM, NaV1.5/NaV1.7 = 340-fold). Further profiling showed that 7 had good permeability, moderate solubility (31 μM), no significant cytochrome P450 (CYP) inhibition for isoforms tested (>10 μM for CYPs 3A4, 1A2, 2C19, 2C9, and 2D6), and no human ether-a-go-go-related gene liability (IC50 > 10 μM). However, 7 showed moderate and poor stabilities in mouse and human liver microsomes (MLMs = 32 ml/min/kg),

■

RESULTS AND DISCUSSION Compounds were evaluated in vitro in a radioligand binding assay (LBA) that measured the displacement of aryl sulfonamide [3H]GX-545 from human embryonic kidney (HEK) membranes that expressed hNaV1.7 including β1.26 910



Article

Figure 4. Small-molecule crystal structure of the sodium salt of 7 (crystals grown in 50% MeOH solution), with sodium ions complexed to the charged oxygen atom shown as orange spheres. Two molecules of methanol are also shown. For details, see the Supporting Information.

Table 2. Effects of Adamantane Replacements on NaV1.7 Potency and HLM Stability

a

Ligand binding assay using [3H]GX-545. IC50s are an average of at least two independent determinations. bIC50s are an average of two independent determinations. cPredicted hepatic clearance (CLhep) extrapolated from in vitro experiments in human liver microsomes (CLhep = 19 ml/min/kg correlates to a 100% extraction ratio);35 nd: not determined. dmLogD at pH 7.4. eLLE was calculated using hNaV1.7 EP IC50 and cLogP.

represented a good starting point for a lead optimization campaign. To better understand how the present acyl sulfonamides might interact with VSD4 of the NaV1.7 receptor, we modeled representative analogues into the crystal structure of VSD4 via induced fit docking. A proposed fit of 7 in VSD4 is shown in Figure 3, supported in part by mutagenesis and small-molecule X-ray data. For the aryl sulfonamide GX-936, a large loss of potency was seen for the R1608A and R1608K mutations (3000× and 400×, respectively), whereas the R1602A mutation caused little change in affinity.26 In contrast, mutagenesis studies with acyl sulfonamide 7 showed that both R1608 and R1602 were required for binding, with 300× and 50× decreases in potency observed for the R1608A and R1602A mutations, respectively. We therefore postulated that the acyl sulfonamide warhead anion is delocalized between the

HLMs = 19 ml/min/kg), respectively. Initial in vivo pharmacokinetic (PK) evaluation in mouse showed good oral bioavailability and exposure. The observed exposure, despite moderate mouse liver microsomal stability, was most likely a result of high plasma protein binding (PPB, >99.9% bound) that restricted the intrinsic clearance of the molecule. We were cognizant of the fact that high plasma protein binding would require exquisite potency to achieve efficacy in vivo without requiring very high exposure. In addition, optimization of metabolic stability was particularly important for these highly bound analogues as their low volume of distribution required good metabolic stability to achieve a half-life suitable for twice-a-day dosing. Hence, our optimization focused particularly on increasing potency and HLM stability. Given its potency, selectivity, and biopharmaceutical properties, 7 911



Article

Table 3. Effect of Substitutions on the Phenyl Core on NaV1.7 Potency and HLM Stability

a

Ligand binding assay using [3H]GX-545. IC50s are an average of at least two independent determinations. bIC50s are an average of two independent determinations. cPredicted hepatic clearance (CLhep) extrapolated from in vitro experiments in human liver microsomes (CLhep = 19 mL/(min kg) correlates to a 100% extraction ratio);35 nd: not determined. dLLE was calculated using hNaV1.7 EP IC50 and cLogP.

adamantane are main contributors to high metabolic clearance, so our initial SAR exploration focused on a diverse set of replacements aimed at understanding the impact on potency and metabolic stability (Table 2). Since metabolic stability is very often linked to lipophilicity,33 we also tracked measured LogD (mLogD) and cLogP as well as lipophilic ligand efficiency (LLE) (calculated using hNaV1.7 EP IC50 and cLogP) in the initial set of compounds to understand if there is a correlation. In addition, lowering lipophilicity is an established medicinal chemistry strategy to decrease the risk for drug promiscuity and unwanted pharmacology.34 The difference between cLogP and mLogD reflects the compound’s ability to exist as deprotonated species at pH 7.4. Cyclohexane 8 demonstrated a 7-fold loss in hNaV1.7 potency, but a slight increase in HLM stability compared to 7. Incorporation of an oxygen atom into the cyclohexyl ring (9) was not tolerated. Extending the linker by one carbon (10) only slightly increased the potency while negatively affecting metabolic stability. Shortening the linker (11) or reducing the ring size by one carbon (12) resulted in a further decrease in potency but enhanced metabolic stability. As expected, metabolic stability and lipophilicity of the left-hand portion of the molecule were generally correlated, with cyclopentyl ether 12 and ethylcyclopropyl ether 13 being the most stable in this subset. However, both displayed unacceptably low potency in the LBA. In this initial library, we found a relatively good correlation between mLogD and HLM stability. For example, 13 showed one of the lowest mLogD values (0.4) and was also the most stable in HLMs (HLM CLhep = 7 mL/(min/kg). On the other hand, the homologated cyclohexane 10, which showed one of the highest mLogD values compared to other analogues in this subset, had low stability in HLMs (CLhep = 19 mL/min/kg). However, in a larger data set, this tendency could not be confirmed. We suspect that structural features are overriding the HLM-lipophilicity trend.

carbonyl and sulfonyl moieties, making hydrogen bonding and ionic interactions with both R1608 and R1602. Sulfonyl oxygens are known to form hydrogen bonds with arginines, although the interaction is often weak. The combination of SO2 to R1608 and the amide carbonyl oxygen to R1602 interactions proposed for acyl sulfonamides rationalizes the roughly equipotent affinity between the acyl and aryl classes of inhibitors. A small-molecule X-ray structure of the sodium salt of 7 grown in aqueous media (Figure 4) shows a sodium ion complexed to an ionized amide carbonyl oxygen, which may partially exist as its imino tautomer, consistent with our hypothesis and binding to R1602 in VSD4. This proposed fit results in the linker phenyl ring being offset from the analogous aryl sulfonamide phenyl ring in the X-ray crystal structure of GX-93626 and is consistent with the lack of corresponding structure−activity relationship (SAR) between aryl and acyl sulfonamides. Specifically, the linker phenyl in 7 is predicted to be translated approximately 1.5−1.7 Å out of the active site, providing room for the adamantyl group and substituted cyclohexyl rings of the present series to fit into a mostly hydrophobic region flanked by W1538, V1541, F1583, M1582, and the side-chain carbon atoms of D1586. This fit would drive the chlorine of 7 into a different region than that occupied by the cyano in GX-936, essentially occupying the center of the channel. The X-ray structure of NaV1.7 VSD4 represents one conformation of a flexible system designed to enable outward movement of S4 during a depolarization. This flexibility and induced fit of ligands introduce uncertainty in the modeling calculations. Other states of this protein can no doubt exist. Nonetheless, the modeling experiments using induced fit docking have served to rationalize parts of the SAR such as phenyl ring and sulfonamide substitution and provided design guidance throughout the project. With these data in hand, we focused our attention on improving the potency and metabolic stability of compound 7. We were cognizant of the fact that large lipophilic groups like 912



Article

Table 4. Effects of Adamantane Modification on NaV1.7 Potency and HLM Stability

a

Ligand binding assay using [3H]GX-545. IC50s are an average of at least two independent determinations. bIC50s are an average of two independent determinations. cPredicted hepatic clearance CLhep extrapolated from in vitro experiments in human liver microsomes (CLhep = 19 ml/min/kg correlates to a 100% extraction ratio);35 nd: not determined. dLLE was calculated using hNaV1.7 EP IC50 and clogP.

and 23) resulted in negligible effect on HLM stability for 22 and markedly decreased HLM for 23. Both 22 and 23 displayed significant loss of hNaV1.7 potency compared to 7. Given 23 had relatively good potency and stability, we wanted to probe if replacing Cl with cyclopropyl would increase potency and retain HLM stability as seen for 20. While cyclopropyl analogue 24 did indeed show potency increases in ligand binding and EP, its HLM stability decreased significantly. Finally, substitution of each tertiary carbon on the adamantane with fluorine (25) substantially improved stability but again at a marked cost in hNaV1.7 potency and LLE relative to 7 or 20. Lessons taken from the SAR developed in Tables 2−4 influenced the design of the next round of analogues (Table 5). Specifically, our goal was to incorporate strategies used successfully to block metabolism of the adamantyl group on the synthetically less complex cyclohexyl ring while also taking advantage of potency improvements realized in the core. We also wanted to maintain the relatively high fsp3 count of the overall molecule. Replacing the 5-chloro in 8 with a cyclopropane (26) led to a significant boost in NaV1.7 potency, particularly in the functional assay, but unfortunately retained the HLM instability seen with previous analogues. MetID studies revealed three major metabolites resulting from oxidation of the cyclohexyl ring (Figure 5). Further improvements in stability were obtained through gem-difluorination at the 4-position of the cyclohexyl ring (27). This compound also retained the LLE shown by 26. However, it showed only moderate LBA potency compared to others in the series. We next turned to examining the effect of varying the acyl sulfonamide substitution on potency and stability. Interestingly, replacement of the N-methylsulfonyl substituent in 27

Considering that 7 displayed the highest potency in a functional assay and the highest LLE, we used this building block for further optimization to explore the SAR of substituents on the phenyl core (Table 3). A 2-fluoro substitution was essential for hNaV1.7 potency, as removal (14) or replacement with chlorine (15) resulted in significant drops in potency. Replacement of the 5-chlorine in 7 with fluorine (16) also significantly reduced potency. Likewise, replacement of the 5-chlorine with methyl (17), ethyl (18), or isopropyl (19) was not beneficial to potency. However, introducing a cyclopropyl group as a chloro replacement (20) turned out to be highly favorable, providing a greater than 10-fold potency enhancement with minimal increases in molecular weight or lipophilicity (cLogP = 3.5 and 3.6 for 7 and 20, respectively), resulting in a significant gain in LLE. Unfortunately, modification of the phenyl ring had no impact on HLM stability. The marked increase in potency of 20 over 7 may be a result of the cyclopropyl making additional hydrophobic contacts with the protein, or simply being more compatible with the adjacent hydrophobic membrane environment. Given the high potency and LLE for 20, we now explored ways of improving the metabolic stability of the adamantane itself. Metabolite identification (MetID) studies on 7 indicated that the adamantal skeleton was subject to oxidative metabolism. We therefore decided to investigate blocking potential sites of metabolism and reducing lipophilicity by incorporating polar substituents on this ring system. As shown in Table 4, introduction of a polar substituent (21) significantly enhanced HLM stability, but compromised potency on hNaV1.7 Other attempts to block the potential sites of oxidative metabolism via introduction of fluorine (22 913



Article

Table 5. Effects of Cyclohexane Modification on NaV1.7 Potency and HLM Stability

a

Ligand binding assay using [3H]GX-545. IC50s are an average of at least two independent determinations. bIC50s are an average of two independent determinations. cPredicted hepatic clearance CLhep extrapolated from in vitro experiments in human liver microsomes (CLhep = 19 ml/min/kg correlates to a 100% extraction ratio);35 nd: not determined. dLLE was calculated using hNaV1.7 EP IC50 and cLogP.

with a trifluoromethyl (32) retained potency relative to 30, but decreased stability. As an alternative to blocking metabolically labile sites by fluorine atoms, we also explored the incorporation of strained systems. An example is 33, which showed the highest functional activity against hNaV1.7 (IC50 = 0.6 nM) and retained moderate stability in human microsomes and hepatocytes (Table 6). Compounds 30 and 33 were selected for further characterization against a panel of human and rodent NaV isoforms as they showed distinct advantages over other analogues. Compound 33 showed the highest potency in EP, high LLE, and moderate stability, whereas 30 showed acceptable potency and HLM stability as well as the highest LLE in the cyclohexane series. Although 33 showed good selectivity over hNaV1.5, it was only about 10- and 2-fold selective against hNaV1.1 and hNaV1.2, respectively. Compounds 30 and 33 also did not inhibit (IC50 >10 μM) mNaV1.8 in tetrodotoxinresistant (TTX-R) channels in mouse dorsal root ganglion (DRG) neurons. Compound 30 showed high selectivity over hNaV 1.5 and greater selectivity against hNaV 1.1 than acyl sulfonamide 33. Compounds 30 and 33 were also tested against mouse and rat NaV1.7, where they showed potencies similar to those of the human isoform. The invariant human, rat, and mouse NaV1.7 potencies are a distinct characteristic that distinguish the present acyl sulfonamide series relative to other reported

Figure 5. Summary of the major metabolic pathway of 26.

with N-cyclopropylsulfonyl (28) or N-(2-methoxyethyl)sulfonyl (29) enhanced stability. Potency was retained in 28, while 29 was much less potent, presumably due to size limitations in this region of the protein. From this small SAR study of the warhead, we learned that our ability to modulate potency and stability via changes in acyl sulfonamide substitution was limited. We therefore turned our attention back to the cyclohexyl-ether appendage. We examined the effect of adding methyl groups36 by incorporation of a substituent on the tertiary carbon at the 1-position of the cyclohexyl ring. Compound 30 provided a moderate improvement in both potency and stability over 29. Increasing the size of the substituent at the tertiary position had a detrimental effect on stability (31). Therefore, the methyl group was incorporated as a preferred substituent for further cyclohexane explorations. Replacing the 4-difluoro group in 30 914



Article

Table 6. In Vitro Profiles of Compounds 30 and 33 compound

30

33

NaV1.7, IC50 (nM)a (human/mouse/rat) hNaV1.5, 1.1, 1.2, 1.6, IC50 (nM)a % inh. TTX-R at (10 μM) liver microsomes CLhep (ml/min/kg)b (human/mouse/rat/dog) hepatocytes CLhep (ml/min/kg)c (human/mouse/rat/dog) CYP 3A4/2C9/2C19/2D6/1A2, IC50 (μM) MDCK1, Papp (A to B) × 10−6 cm/s, ratio (BA/AB)

3/2.5/9 260/48/6/49 26 2/38/10/7 2.8/17/4.3/3.5 >10/>10/>10/>10/>10 22 0.55 11 1.4 0.4 56 99.6/99.8/99.8 32 1.4/6.8

0.6/2.2/3.7 50/6/2/38 38 12/65/22/nd 13/6.0/5.2/7.6 >10/6.1/>10/>10/>10 9 1.0 3.7 1.2 5.1 2.1 99.9/99.9/99.9 10 2.1/7.1

MDCK1-MDR1, Papp (A to B) × 10−6 cm/s, ratio (BA/AB) MDCK2-mBCRP, Papp (A to B) × 10−6 cm/s, ratio (BA/AB) PPB (%) (human/mouse/rat) kinetic solubility (μM) mLogD/LLE(EP/mLogD)d,e

a IC50 values are an average of two independent determinations using PatchXpress electrophysiology technique. bPredicted hepatic clearance (CLhep) extrapolated from in vitro human, mouse, and rat microsome experiments.35 cPredicted hepatic clearance (CLhep) extrapolated from in vitro human, mouse, rat, and dog hepatocyte experiments.35 dMeasured mLogD at pH 7.4. eLLE was calculated using hNaV1.7 EP IC50 and mLogD.

Table 7. Pharmacokinetic Profiles for Compounds 30 and 33 IV

PO

compd

species

t1/2 (h)

CL (ml/min/kg)

Vss (L/kg)

t1/2 (h)

Cmax (μM)

AUC (μM h)

F (%)

30 30 33 33 33

mousea ratb mousea ratb dogc

2.1 9.0 1.5 8.1 5.4

1.6 0.4 3.2 1.2 0.3

0.28 0.29 0.41 0.88 0.14

15.6 16.9 9.2 nc 6.0

69 25 22 13 9

350 160 113 78 83

49 66 31 49 68

a Average of three male FVB mice, dosed IV with 1 mg/kg of compound 30 or 33 in 5% dimethyl sulfoxide (DMSO)/35% poly(ethylene glycol)400 (PEG400)/60% phosphate-buffered saline (PBS) or dosed PO with 30 mg/kg of compound 30 or 33 as MCT suspension. bAverage of three male Sprague-Dawley rats, dosed IV with 1 mg/kg of compound 30 or 33 in 10% DMSO/50% PEG400/40% PBS or dosed PO with 5 mg/kg of compound 30 or 33 as MCT suspension. nc: not calculated. cAverage of three male beagle dogs, dosed IV with 33 in 0.5 mg/kg in 15% EtOH and 60% PEG400 in water and orally dosed with 33 at 1 mg/kg as MCT suspension.

LLE when calculated based on mLogD was high for both 30 and 33 and in line with low off-target activity described for 33. On the basis of their overall in vitro potencies and DMPK profiles, 30 and 33 were selected for further in vivo evaluation. Their PK profiles were determined in mouse and rat, and 33 was also evaluated in dog. The results are summarized in Table 7. Consistent with the predicted CLhep and high plasma protein binding, both compounds exhibited very low plasma clearance (CL). Both 30 and 33 displayed moderate half-lives (t1/2) in mice. Compound 30 and 33 showed increased half-lives in rat intravenous (IV) experiments, with 33 showing acceptable half-lives in dog IV and PO studies. They also displayed low volume of distribution (Vss) values in these animal species, presumably because of high plasma protein binding. In agreement with their high permeability and solubility, both compounds displayed good to moderate oral plasma exposures (area under curve (AUC)) and bioavailabilities (F). The oral bioavailabilities of 33 in mice, rats, and dogs were 31, 49, and 68%, respectively. To determine if the PK and potencies of our compounds were sufficient to demonstrate analgesia in vivo, 30 and 33 were evaluated in transgenic mice21c,37 that express a human variant of NaV1.7 that results in IEM (mutation I848T).38 Aconitine-induced pain in these mice provides a novel way to quantify engagement of hNaV1.7 in vivo and facilitates the

acyl and aryl sulfonamides (e.g., 4) that bind to the VSD4 site. We believe that the increased conformational flexibility between cycloalkylether and phenyl core allow our compounds to adopt a binding mode in rat NaV1.7 that is similar to the one shown in compound 7 in Figure 3, and adapt that binding mode to residue changes in the VSD between human, mouse, and rat. The less flexible, more deeply binding aryl sulfonamides such as 4 cannot readily adjust to the residue differences in the rNaV1.7 binding pocket and are less potent as a result. Gratifyingly, both 30 and 33 were selective for hNaV1.7 over hNaV1.5. No significant drug−drug interaction risk was observed, as 30 and 33 did not show notable CYP inhibition (Table 6). We tracked permeability in a mouse Madin−Darby canine kidney cell line (MDCK)2−breast cancer resistance protein (BCRP) cell line, which contains the main transporter found at the blood−brain barrier, to better understand the brain exposure of our compounds. Compound 33 had good permeability and low efflux, while 30 proved to be a mouse BCRP (mBCRP) substrate. Both compounds showed high protein binding in human, mouse, and rat (Table 6). We also evaluated 33 in a CEREP panel of 33 receptors at 10 μM concentration, where we saw >75% inhibition of only two targets: NaV ion channels (80%) and PPARγ (91%). The mLogD values at pH 7.4 were acceptable for both 30 (mLogD = 1.4) and 33 (mLogD = 2.1). Notably, 915



Article

were gratified to see this remarkable efficacy for 30 and 33 despite their high plasma protein binding. The significantly higher efficacy of 33 over 30 was rationalized by its 5-fold higher potency in the functional assay for the inhibition of hNaV1.7. In vivo effects of 33 were further assessed in a formalin-induced pain model in mice,39 an assay that has been used to evaluate analgesic effects for many sodium channel blockers.23,25b,40 This pain model utilizes formalin as a paininducing agent and generates a biphasic pain response. The initial response (phase I), occurring within 5−10 min after injection of formalin, indicates an acute localized irritation (partially due to activation of TRPA1 channels41), whereas a delayed response (phase II) that begins 10−15 min after injection indicates neurogenic inflammation.42 As illustrated in Figure 8, 33 produced significant response reduction in phase

characterization of selective inhibitors that, most often, show species specificity for rodent orthologs. This assay reports the number of nociceptor responses (shaking and licking) in response to the sodium channel activator aconitine. As illustrated in Figures 6 and 7, 30 and 33 demonstrated dose-

Figure 6. Effects of 30 on aconitine-induced pain in humanized IEM transgenic mice. Each bar represents the mean of at least four animals. The error bars represent standard errors of the mean. **** p < 0.0001 vs vehicle group. Statistical significance of results was calculated by two-way analysis of variance (ANOVA) using Prism version 7 (GraphPad software).

Figure 7. Effects of 33 on aconitine-induced pain in humanized IEM transgenic mice. Each bar represents the mean of at least four animals, except the 0.3 and 1 mg/kg groups (three animals for each group at these doses). The error bars represent standard errors of the mean. **** p < 0.0001 and *** p < 0.005 vs vehicle group for nociceptive events. Statistical significance of results was calculated by two-way ANOVA using Prism version 7 (GraphPad software).

Figure 8. Dose−response effect of 33 in the mouse formalin model. (A) Time course of inhibition of formalin-evoked pain behavior. Each point represents the mean number of times that a mouse (n = 4) shook or licked its injected paw over a 5 min period. (B) Quantification of the response to formalin binned into two different phases: phase I (0−10 min after formalin injection) and phase II (11−40 min after formalin injection). Each bar represents the mean of four animals. The error bars represent standard errors of the mean. *** p = 0.0021 vs vehicle group. Statistical significance of results was calculated by two-way ANOVA using prism version 7 software (GraphPad software).

related inhibition of aconitine-induced nociceptive behaviors, matched by dose-dependent increases in plasma concentrations. Consistent with its significantly higher functional potency against NaV1.7, 33 was much more potent in this study, demonstrating >75% analgesic efficacy starting at a plasma concentration of approximately 3.1 μM (which corresponds to an unbound plasma concentration of Cpu = 3 nM, approximately 5-fold the hNaV1.7 IC50) (0.3 mg/kg, oral dose). At these very low efficacious free plasma concentrations, we did not expect to see significant inhibition of other NaV isoforms. Plasma concentrations of 33 for the 3 and 10 mg/kg groups were measured as 25 and 79 μM, respectively. The brain concentrations were 1.9 and 6.2 μM, respectively, which provided a low brain-to-plasma ratio of approximately 0.08. We

II of the formalin assay at a PO dose of 10 mg/kg. The plasma concentration of 33 was measured at the end of the study (approximately 3 h after oral dosing) to be 50 μM. Despite the modest selectivity for the inhibition of hNaV1.7 over hNaV1.1 and hNaV1.2, 33 was well tolerated in mice, as there were no clinical observations over the course of this study. We also evaluated 30 and 33 for the inhibition of mNaV1.8 in TTX916



Article

Scheme 1. Syntheses of Compounds 7−14, 16, 21−23, and 25a

a

Reagents and conditions: (a) for 37 and 38: MeSO2NH2, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI), 4-dimethylaminopyridine (DMAP), CH2Cl2, room temperature (rt), 58−96%; for 39: MeSO2NH2, Et3N, CH3CN, rt, 54%; (b) R1OH, t-BuOK, DMSO, rt, 9−58%.

Scheme 2. Synthesis of Compound 15a

Reagents and conditions: (a) adamantan-1-ylmethyl methanesulfonate, K2CO3, dimethylformamide (DMF), 65 °C, 71%; (b) MeSO2NH2, Mo(CO)6, Pd(OAc)2, Xantphos, Et3N, 1,4-dioxane, 100 °C, microwave irradiation, 45 min, 10%.

a


Reagents and conditions: (a) 1-adamantane methanol, t-BuOK, DMSO, rt to 50 °C, 17%; (b) MeSO2NH2, EDCI, DMAP, CH2Cl2, rt, 27%.

a


Reagents and conditions: (a) di-tert-butyl dicarbonate, DMAP, tert-butanol, 50 °C, 99%; (b) (adamantan-1-yl)methanol, Cs2CO3, DMSO, 70 °C, 62%; (c) vinylboronic acid pinacol ester, tris(dibenzylideneacetone)dipalladium(0) (Pd2(dba)3), t-Bu3P·HBF4, Na2CO3, 1,4-dioxane, H2O, 100 °C, 92%; (d) TFA, CH2Cl2, rt, 97%; (e) Pd−C, H2, EtOAc, rt, 74%; (f) MeSO2NH2, EDCI, DMAP, CH2Cl2, rt, 69%.

a

fashion to prepare the desired molecules 7−14, 16, 21−23, and 25. Analogue 15 was synthesized in a two-step sequence from phenol 40, which was alkylated to give intermediate 41. Compound 15 was prepared from 41 via introduction of the acyl sulfonamide group by a palladium-catalyzed carbonylation.43 Compound 17 was prepared from 42 as detailed in Scheme 3, involving a nucleophilic aromatic substitution and an acyl sulfonamide formation in the presence of EDCI and DMAP. The synthesis of analogue 18 involved the esterification of 34, nucleophilic aromatic substitution of 44

resistant channels in mouse dorsal root ganglion neurons and found minimal inhibition (IC50 > 10 μM), indicating that their efficacy was not caused by the inhibition of this isoform.

■

CHEMISTRY The general synthetic routes to prepare acyl sulfonamides 7− 33 are outlined in scheme 1−8. For syntheses of analogues 7− 14, 16, 21−23, and 25 (Scheme 1), a simple and efficient nucleophilic aromatic substitution of intermediates 37, 38, and 39 with an appropriate alcohol was applied. This method was general, and a wide range of alcohols could be used in a library 917



Article


Reagents and conditions: (a) 2-isopropenyl-4,4,5,5-tetramethyl-1,3,2-dioxaborolane, Pd2(dba)3, t-Bu3P·HBF4, Na2CO3, 1,4-dioxane, H2O, 100 °C, 64%; (b) Pd−C, H2, EtOAc, rt, 94%; (c) TFA, CH2Cl2, rt, 84%; (d) MeSO2NH2, EDCI, DMAP, CH2Cl2, rt, 33%.

a

Scheme 6. Syntheses of Compounds 20 and 26−32a

a Reagents and conditions: (a) R1OH, Cs2CO3, DMSO, 70 °C, 43−95%; (b) cyclopropylboronic acid, Pd(OAc)2, Cy3P·HBF4, K3PO4, toluene, H2O, 100 °C, 86−99%; (c) TFA, CH2Cl2, rt, 50−86%; (d) R2SO2NH2, EDCI, DMAP, CH2Cl2, rt, 40−70%.

use of chloroiodomethane and diethylzinc45 to afford the key intermediate 73.

with (adamantan-1-yl)methanol yielding the predominantly para-regioisomer in 62% yield, Suzuki−Miyaura cross-coupling reaction44 of 45 with vinylboronic acid pinacol ester, hydrolysis of 46 with TFA, hydrogenation of 47, and finally acyl sulfonamide formation of acid 48 with methanesulfonamide (Scheme 4). For analogue 19, a method analogous to that used for 18 was applied. In this preparation, vinylboronic acid pinacol ester was replaced by 2-isopropenyl-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (Scheme 5). The syntheses of 20 and 26−32 started from an aromatic nucleophilic substitution of 44 with an appropriate alcohol in the same manner as described in the preparation of 45. Compounds 45 and 52−56 were then subjected to the Suzuki−Miyaura cross-coupling reaction44 with cyclopropylboronic acid to provide the corresponding cyclopropyl intermediates 57−62. After treatment with TFA, the resulting acids 63−68 were coupled with alkylsulfonamide to afford the desired acyl sulfonamides 20 and 26−32 (Scheme 6). Analogue 24 was directly prepared from 23 in a one-step transformation using the Suzuki−Miyaura cross-coupling reaction44 (Scheme 7). Compound 33 was synthesized in multiple steps from intermediate 44 (Scheme 8). In this preparation, the dioxolane protecting group in 70 was selectively removed with TFA in a mixture of THF and H2O without affecting the tert-butyl ester group. The Simmons− Smith cyclopropanation of 72 proceeded smoothly with the

■

CONCLUSIONS This paper details the discovery of 33, a highly potent and selective NaV1.7 inhibitor that displayed favorable PK in rodents, acceptable subtype selectivity over other Na V isoforms, and efficacy in in vivo models of pain as well as hNaV1.7 target engagement models. Starting with an analysis of the physicochemical property profiles of previously reported aryl- and acyl sulfonamides, followed by library syntheses, we identified lead molecule (7). Guided by induced fit docking into our recently published X-ray cocrystal structure of VSD4 of NaV1.7, we simplified the adamantane to a cyclohexane in compound 26 and conducted metabolite ID studies to guide further designs. Significant improvements in metabolic stability were achieved by identifying the primary metabolites of 26 and utilizing these data to systematically modify the structures to address proposed liabilities. These efforts culminated in the identification of 33, a highly potent and efficacious tool molecule. Finally, we also disclosed CEREP screening data and inhibition data against TTX-resistant currents in mouse DRGs, which provided clear evidence that the in vivo activity of compound 33 was primarily caused by its inhibition of NaV1.7 channels.

■


EXPERIMENTAL SECTION

General. Chemicals, reagents, and solvents were purchased from commercial sources and either used as supplied or purified using reported methods. Final compounds reported herein exhibited spectral data consistent with their proposed structure by nuclear magnetic resonance spectra (1H NMR and 13C NMR) and mass spectra data. NMR spectra were recorded on a Bruker Avance 300 spectrometer with chemical shifts (δ) reported in parts per million relative to the residual signal of the deuterated solvent. 1H NMR data are tabulated in the following order: multiplicity (s, singlet; d, doublet; t, triplet; m, multiplet; br, broad), coupling constants in hertz, and number of protons. Mass spectra were obtained using a Waters 2795/ZQ liquid chromatography (LC)/mass spectrometry (MS) system (Waters Corporation, Milford, MA). Final compounds

Reagents and conditions: (a) R1OH, Cs2CO3, DMSO, 70 °C, 43− 95%; (b) cyclopropylboronic acid, Pd(OAc)2, Cy3P·HBF4, K3PO4, toluene, H2O, 100 °C, 86−99%; (c) TFA, CH2Cl2, rt, 50−86%; (d) R2SO2NH2, EDCI, DMAP, CH2Cl2, rt, 40−70%. a

918



Article


a Reagents and conditions: (a) (8-methyl-1,4-dioxaspiro[4.5]decan-8-yl)methanol, Cs2CO3, DMSO, 70 °C, 54%; (b) cyclopropylboronic acid, Pd(OAc)2, Cy3P·HBF4, K3PO4, toluene, H2O, 100 °C, 83%; (c) trifluoroacetic acid (TFA), H2O, tetrahydrofuran (THF), rt, 90%; (e) MePPh3Br, LiHMDS, THF, −20 °C to rt, 82%; (e) ClCH2I, Et2Zn, ClCH2CH2Cl, 0 °C, 93%; (f) TFA, CH2Cl2, rt, 90%; (g) cyclopropanesulfonamide, EDCI, DMAP, CH2Cl2, rt, 41%.

(300 MHz, DMSO-d6) δ 12.10 (s br, 1H), 7.78 (d, J = 7.5 Hz, 1H), 7.26 (d, J = 12.5 Hz, 1H), 4.03 (d, J = 6.4 Hz, 2H), 3.91−3.86 (m, 2H), 3.38−3.30 (m, 5H), 2.10−2.00 (m, 1H), 1.70−1.65 (m, 2H), 1.43−1.29 (m, 2H). MS (ES−) m/z: 364.1, 366.1 (M − 1). 5-Chloro-4-(2-cyclohexylethoxy)-2-fluoro-N-(methylsulfonyl)benzamide (10). Using a method analogous to the preparation of compound 7, 10 was prepared from 37 and 2-cyclohexylethanol as a colorless solid (0.05 g, 14%). 1H NMR (300 MHz, DMSO-d6) δ 12.05 (s, 1H), 7.76 (d, J = 7.6 Hz, 1H), 7.26 (d, J = 12.4 Hz, 1H), 4.17 (t, J = 6.6 Hz, 2H), 3.29 (s, 3H), 1.76−1.41 (m, 8H), 1.27−0.90 (m, 5H). MS (ES−) m/z: 376.1, 378.1 (M − 1). 5-Chloro-4-(cyclohexyloxy)-2-fluoro-N-(methylsulfonyl)benzamide (11). Using a method analogous to the preparation of compound 7, 11 was prepared from 37 and cyclohexanol as a colorless solid (0.03 g, 9%). 1H NMR (300 MHz, DMSO-d6) δ 12.03 (s br, 1H), 7.73 (d, J = 7.6 Hz, 1H), 7.29 (d, J = 12.7 Hz, 1H), 4.64− 4.57 (m, 1H), 3.31 (s, 3H), 1.88−1.82 (m, 2H), 1.69−1.64 (m, 2H), 1.54−1.27 (m, 6H). MS (ES−) m/z: 348.1, 350.1 (M − 1). 5-Chloro-4-(cyclopentylmethoxy)-2-fluoro-N-(methylsulfonyl)benzamide (12). Using a method analogous to the preparation of compound 7, 12 was prepared from 37 and cyclopentylmethanol as a colorless solid (0.14 g, 40%). 1H NMR (300 MHz, DMSO-d6) δ 12.09 (s br, 1H), 7.77 (d, J = 7.5 Hz, 1H), 7.25 (d, J = 12.5 Hz, 1H), 4.04 (d, J = 6.8 Hz, 2H), 3.35 (s, 3H), 2.39−2.29 (m, 1H), 1.82−1.73 (m, 2H), 1.65−1.52 (m, 4H), 1.40−1.30 (m, 2H). MS (ES−) m/z: 348.1, 350.1 (M − 1). 5-Chloro-4-(2-cyclopropylethoxy)-2-fluoro-N-(methylsulfonyl)benzamide (13). Using a method analogous to the preparation of compound 7, 13 was prepared from 37 and 2-cyclopropylethan-1-ol as a colorless solid (0.26 g, 52%). 1H NMR (300 MHz, DMSO-d6) δ 12.13 (s br, 1H), 7.78 (d, J = 7.5 Hz, 1H), 7.26 (d, J = 12.5 Hz, 1H), 4.20 (t, J = 6.3 Hz, 2H), 3.35 (s, 3H), 1.70−1.63 (m, 2H), 0.87−0.79 (m, 1H), 0.47−0.41 (m, 2H), 0.17−0.12 (m, 2H). MS (ES−) m/z: 334.1, 336.1 (M − 1). 4-(Adamantan-1-ylmethoxy)-3-chloro-N-(methylsulfonyl)benzamide (14). Using a method analogous to the preparation of compound 7, 14 was prepared from 38 and (adamantan-1yl)methanol as a colorless solid (0.34 g, 52%). 1H NMR (300 MHz, DMSO-d6) δ 12.07 (s br, 1H), 8.04−7.94 (m, 2H), 7.25 (d, J = 8.5 Hz, 1H), 3.72 (s, 2H), 3.36 (s, 3H), 1.99 (s br, 3H), 1.71−1.66 (m, 12H). MS (ES−) m/z: 396.1, 398.1 (M − 1). 4-(Adamantan-1-ylmethoxy)-2,5-dichloro-N-(methylsulfonyl)benzamide (15). A mixture of 41 (0.16 g, 0.41 mmol), methanesulfonamide (0.12 g, 1.23 mmol), Mo(CO)6, (0.11 g, 0.41 mmol), and Et3N (0.23 mL, 1.64 mmol) in 1,4-dioxane was purged with nitrogen for 5 min, and Xantphos (0.35 g, 0.082 mmol) and Pd(OAc)2 (0.01 g, 0.041 mmol) were added. The reaction mixture

were higher than 95% pure as determined by analytical highperformance liquid chromatography (HPLC) on Agilent 1200 systems (Agilent Technologies, Santa Clara, CA) using an EMD Chromolith SpeedROD RP-18e column (4.6 mm i.d. × 50 mm length) (Merck KGaA, Darmstadt, Germany). The mobile phase consisted of a gradient of component “A” (0.1% v/v aqueous trifluoroacetic acid) and component “B” (acetonitrile) at a flow rate of 1 mL/min. The gradient program used was as follows: initial conditions 5% B, hold at 5% B for 1 min, linear ramp from 5 to 95% B over 5 min, 100% B for 3 min, return to initial conditions for 1 min. Peaks were detected at a wavelength of 254 nm with an Agilent photodiode array detector. Melting points were determined on a Fisher-Johns melting point apparatus and were uncorrected. Chemical names were generated using ChemBioDraw version 16.0 (CambridgeSoft, Cambridge, MA). 4-(Adamantan-1-ylmethoxy)-5-chloro-2-fluoro-N(methylsulfonyl)benzamide (7). To a mixture of adamantan-1ylmethanol (1.00 g, 6.0 mmol) and anhydrous DMSO (40 mL) was added potassium t-butoxide (1.68 g, 15.0 mmol) at room temperature. The resulting mixture was stirred at room temperature for 30 min followed by the addition of 37 (1.62 g, 6.0 mmol). The reaction mixture was again stirred at room temperature for 16 h, cooled to 0 °C, and quenched with hydrochloric acid (1 N, 30 mL), followed by extraction with ethyl acetate (200 mL). The organic layer was washed with water (2 × 40 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 10− 60% ethyl acetate (containing 2% acetic acid) in hexanes to afford 7 as a colorless solid (1.17 g, 46%). mp 194−196 °C (EtOAc/hexanes) 1H NMR (300 MHz, DMSO-d6) δ 12.10 (s br, 1H), 7.77 (d, J = 7.5 Hz, 1H), 7.23 (d, J = 12.5 Hz, 1H), 3.72 (s, 2H), 3.35 (s, 3H), 1.99 (s br, 3H), 1.75−1.64 (m, 12H). 13C NMR (75 MHz, DMSO-d6) δ 162.3 (d, J = 2.2 Hz), 159.8 (d, JC−F = 255 Hz), 158.3 (d, JC−F = 11 Hz), 130.6 (d, JC−F = 4 Hz), 116.8 (d, JC−F = 3 Hz), 113.6 (d, JC−F = 14 Hz), 102.7 (d, JC−F = 28 Hz), 79.1, 41.2, 38.5, 36.4, 33.4, 27.3. MS (electrospray (ES)−) m/z: 414.1, 416.1 (M − 1). 5-Chloro-4-(cyclohexylmethoxy)-2-fluoro-N-(methylsulfonyl)benzamide (8). Using a method analogous to the preparation of compound 7, 8 was prepared from 37 and cyclohexylmethanol as a colorless solid (0.11 g, 30%). 1H NMR (300 MHz, DMSO-d6) δ 12.04 (s br, 1H), 7.73 (d, J = 7.5 Hz, 1H), 7.21 (d, J = 12.6 Hz, 1H), 3.93 (d, J = 5.9 Hz, 2H), 3.31 (s, 3H), 1.79−1.60 (m, 6H), 1.29−0.97 (m, 5H). MS (ES−) m/z: 362.1, 364.1 (M − 1). 5-Chloro-2-fluoro-N-(methylsulfonyl)-4-((tetrahydro-2H-pyran4-yl)methoxy)benzamide (9). Using a method analogous to the preparation of compound 7, 9 was prepared from 37 and (tetrahydro2H-pyran-4-yl)methanol as a colorless solid (0.160 g, 15%). 1H NMR 919



Article

was heated at 100 °C for 45 min under microwave irradiation and then cooled to room temperature. The reaction was repeated twice on 0.82 mmol scale. The reaction mixtures were combined and concentrated in vacuo. The residue was diluted with EtOAc (40 mL) and saturated aqueous NH4Cl solution (10 mL). The organic layer was separated, and the aqueous layer was extracted with EtOAc (2 × 25 mL). The combined organic extract was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 10−40% EtOAc (containing 0.2% acetic acid) in hexanes to afford 15 as a colorless solid (0.09 g, 10%). 1H NMR (300 MHz, CDCl3) δ 8.96 (s br, 1H), 7.99 (s, 1H), 6.91 (s, 1H), 3.60 (s, 2H), 3.42 (s, 3H), 2.09−2.01 (m, 3H), 1.83−1.64 (m, 12H). MS (ES−) m/z: 430.2 (M −1), 432.2 (M −1). 4-(Adamantan-1-ylmethoxy)-2,5-difluoro-N-(methylsulfonyl)benzamide (16). Using a method analogous to the preparation of compound 7, 16 was prepared from 39 and adamantan-1-ylmethanol as a colorless solid (0.145 g, 36%): 1H NMR (300 MHz, DMSO-d6) δ 12.01 (s, 1H), 7.53 (dd, J = 6.8, 11.3 Hz, 1H), 7.23 (dd, J = 6.8, 12.3 Hz, 1H), 3.68 (s, 2H), 3.31 (s, 3H), 1.95 (s, 3H), 1.71−1.58 (m, 12H). MS (ES−) m/z: 398.1 (M − 1). 4-(Adamantan-1-ylmethoxy)-2-fluoro-5-methyl-N(methylsulfonyl)benzamide (17). To a mixture of 43 (0.16 g, 0.50 mmol) in anhydrous CH2Cl2 (25 mL) were added EDCI (0.22 g, 1.15 mmol), DMAP (0.14 g, 1.15 mmol), and MeSO2NH2 (11.44 g, 119.0 mmol) at room temperature. After stirring at room temperature for 16 h, the reaction mixture was quenched by the addition of a 1 M HCl solution (5 mL) at 0 °C. The organic layer was separated, and the aqueous layer was extracted with EtOAc (2 × 25 mL). The combined organic extract was washed with water and brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was crystallized from EtOAc and hexanes to afford 17 as a colorless solid (0.06 g, 27%). 1H NMR (300 MHz, DMSO-d6) δ 11.85 (s br, 1H), 7.48 (d, J = 8.3 Hz, 1H), 6.93 (d, J = 13.0 Hz, 1H), 3.62 (s, 2H), 3.33 (s, 3H), 2.15 (s, 3H), 1.99 (s br, 3H), 1.75−1.66 (m, 12H). MS (ES+) m/z: 396.2 (M + 1); MS (ES−) m/z: 394.2 (M − 1). 4-(Adamantan-1-ylmethoxy)-5-ethyl-2-fluoro-N(methylsulfonyl)benzamide (18). Using a method analogous to the preparation of 17, compound 18 was prepared from 48 and MeSO2NH2 as a colorless solid (0.17 g, 69%). 1H NMR (300 MHz, DMSO-d6) δ 11.87 (s br, 1H), 7.46 (d, J = 8.5 Hz, 1H), 6.94 (d, J = 13.1 Hz, 1H), 3.62 (s, 2H), 3.34 (s, 3H), 2.58 (q, J = 7.5 Hz, 2H), 1.99 (s br, 3H), 1.75−1.64 (m, 12H), 1.15 (t, J = 7.5 Hz, 3H). MS (ES+) m/z: 410.2 (M + 1); MS (ES−) m/z: 408.3 (M − 1). 4-(Adamantan-1 -ylmet ho xy)-2-fluo ro -5-isopr opyl-N(methylsulfonyl)benzamide (19). Using a method analogous to the preparation of 17, compound 19 was prepared from 51 and MeSO2NH2 as a colorless solid (0.16 g, 33%). 1H NMR (300 MHz, CDCl3) δ 8.73 (d, J = 16.5 Hz, 1H), 7.93 (d, J = 9.6 Hz, 1H), 6.57 (d, J = 14.7 Hz, 1H), 3.53 (s, 2H), 3.42 (s, 3H), 3.33−3.19 (m, 1H), 2.05 (s, 3H), 1.84−1.62 (m, 12H), 1.23 (d, J = 6.9 Hz). MS (ES−) m/z: 422.3 (M − 1). 4-(Adamantan-1-ylmethoxy)-5-cyclopropyl-2-fluoro-N(methylsulfonyl)benzamide (20). Using a method analogous to the preparation of 17, compound 20 was prepared from 63 and MeSO2NH2 as a colorless solid (0.45 g, 70%). mp 158−159 °C (EtOAc/hexanes). 1H NMR (300 MHz, CDCl3) δ 8.72 (d, J = 16.2 Hz, 1H), 7.58 (d, J = 9.3 Hz, 1H), 6.56 (d, J = 14.7 Hz, 1H), 3.56 (s, 2H), 3.41 (s, 3H), 2.09−2.00 (m, 4H), 1.81−1.58 (m, 12H), 0.99− 0.91 (m, 2H), 0.70−0.62 (m, 2H). 13C NMR (75 MHz, CDCl3) δ 163.9 (d, JC−F = 11 Hz), 160.5 (d, JC−F = 246 Hz), 161.5 (d, JC−F = 3 Hz), 129.9 (d, JC−F = 2 Hz), 128.2 (d, JC−F = 3 Hz), 108.6 (d, JC−F = 10 Hz), 98.7 (d, JC−F = 29 Hz), 78.9, 41.8, 39.3, 36.9, 33.8, 27.9, 9.3, 7.2. MS (ES+) m/z: 422.2 (M − 1); MS (ES−) m/z: 420.2 (M − 1). 5-Chloro-2-fluoro-4-((3-hydroxyadamantan-1-yl)methoxy)-N(methylsulfonyl)benzamide (21). Using a method analogous to the preparation of compound 7, 21 was prepared from 37 and 3(hydroxymethyl)adamantan-1-ol as a colorless solid (0.25 g, 29%). 1H

NMR (300 MHz, DMSO-d6) δ 12.11 (s br, 1H), 7.76 (d, J = 6.6 Hz, 1H), 7.23 (d, J = 12.3 Hz, 1H), 4.36 (s br, 1H), 3.78 (s, 2H), 3.35 (s, 3H), 2.14 (s br, 2H), 1.68−1.39 (m, 12H). MS (ES−) m/z: 430.1, 432.1 (M − 1). 5-Chloro-2-fluoro-4-((3-fluoroadamantan-1-yl)methoxy)-N(methylsulfonyl)benzamide (22). Using a method analogous to the preparation of compound 7, 22 was prepared from 37 and (3fluoroadamantan-1-yl)methanol as a colorless solid (0.37 g, 58%). 1H NMR (300 MHz, DMSO-d6) δ 12.12 (s br, 1H), 7.78 (d, J = 7.5 Hz, 1H), 7.24 (d, J = 12.4 Hz, 1H), 3.85 (s, 2H), 3.35 (s, 3H), 2.30 (s, 2H), 1.82−1.76 (m, 6H), 1.61−1.52 (m, 6H). MS (ES−) m/z: 432.1, 434.1 (M − 1). 5-Chloro-4-((4,4-difluoroadamantan-1-yl)methoxy)-2-fluoro-N(methylsulfonyl)benzamide (23). Using a method analogous to the preparation of compound 7, 23 was prepared from 37 and (4,4difluoroadamantan-1-yl)methanol as a colorless solid (0.47 g, 42%). 1 H NMR (300 MHz, CDCl3) δ 8.69 (s br, 1H), 8.06 (d, J = 8.4 Hz, 1H), 6.65 (d, J = 13.8 Hz, 1H), 3.61 (s, 2H), 3.39 (s, 3H), 2.28 (s br, 2H), 2.05−1.87 (m, 5H), 1.78−1.66 (m, 6H). MS (ES+) m/z: 452.1, 454.1 (M + 1); MS (ES−) m/z: 450.2, 452.2 (M − 1). 5-Cyclopropyl-4-((4,4-difluoroadamantan-1-yl)methoxy)-2-fluoro-N-(methylsulfonyl)benzamide (24). To a mixture of 23 (0.37 g, 0.81 mmol), cyclopropylboronic acid (0.21 g, 2.44 mmol), and K3PO4 (1.72 g, 8.10 mmol) in toluene (30 mL) and water (1.5 mL) under a nitrogen atmosphere were added Cy3P·HBF4 (0.06 g, 0.16 mmol) and Pd(OAc)2 (0.02 g, 0.9 mmol). The reaction mixture was heated to 100 °C for 18 h and then cooled to ambient temperature. Aqueous hydrochloric acid (5%, 20 mL) was added, and the mixture was extracted with EtOAc (100 mL × 3); the combined organics were washed with brine, dried over anhydrous sodium sulfate, and concentrated in vacuo. Purification of the residue by column chromatography (10−30% gradient ethyl acetate in hexanes) afforded the title compound as a colorless solid (0.32 g, 86%). 1H NMR (300 MHz, CDCl3) δ 8.69 (br s, 1H), 7.59 (d, J = 9.0 Hz, 1H), 6.53 (d, J = 14.1 Hz, 1H), 3.59 (s, 2H), 3.39 (s, 3H), 2.28 (br s, 2H), 2.08−1.92 (m, 6H), 1.53−1.51 (m, 6H), 0.97−0.89 (m, 2H), 0.66−0.60 (m, 2H). MS (ES−) m/z: 456.2 (M − 1). 5-Chloro-2-fluoro-N-(methylsulfonyl)-4-((3,5,7-trifluoroadamantan-1-yl)methoxy)benzamide (25). Using a method analogous to the preparation of compound 7, 25 was prepared from 37 and (3,5,7trifluoroadamantan-1-yl)methanol as a colorless solid (0.04 g, 28%). 1 H NMR (300 MHz, DMSO-d6) δ 12.10 (br s, 1H), 7.76 (d, J = 7.6 Hz, 1H), 7.20 (d, J = 12.28 Hz, 1H), 4.00 (s, 2H), 3.22 (s, 3H), 2.26−2.15 (m, 3H), 2.07−1.97 (m, 3H), 1.82−1.74 (m, 6H). MS (ES−) m/z 468.21, 470.20 (M − 1). 4-(Cyclohexylmethoxy)-5-cyclopropyl-2-fluoro-N(methylsulfonyl)benzamide (26). Using a method analogous to the preparation of 17, compound 26 was prepared from 64 and MeSO2NH2 as a colorless solid (0.19 g, 40%). 1H NMR (300 MHz, CDCl3) δ 8.71 (d, J = 16.5 Hz, 1H), 7.56 (d, J = 9.0 Hz, 1H), 6.56 (d, J = 14.7 Hz, 1H), 3.82 (d, J = 5.7 Hz, 2H), 3.41 (s, 3H), 2.11−2.01 (m, 1H), 1.96−1.68 (m, 6H), 1.39−1.04 (m, 5H), 0.98− 0.90 (m, 2H), 0.69−0.63 (m, 2H). MS (ES−) m/z: 368.3 (M − 1). 5-Cyclopropyl-4-((4,4-difluorocyclohexyl)methoxy)-2-fluoro-N(methylsulfonyl)benzamide (27). Using a method analogous to the preparation of 17, compound 27 was prepared from 65 and MeSO2NH2 as a colorless solid (0.04 g, 13%). 1H NMR (300 MHz, CDCl3) δ 8.69 (d, J = 15.0 Hz, 1H), 7.57 (d, J = 9.0 Hz, 1H), 6.55 (d, J = 14.4 Hz, 1H), 3.88 (d, J = 9.0 Hz, 2H), 3.39 (s, 3H), 2.25−2.11 (m, 2H), 2.04−1.66 (m, 6H), 1.54−1.41 (m, 2H), 0.97− 0.89 (m, 2H), 0.67−0.61 (m, 2H). MS (ES−) m/z: 404.1 (M − 1). 5-Cyclopropyl-N-(cyclopropylsulfonyl)-4-((4,4-difluorocyclohexyl)-methoxy)-2-fluorobenzamide (28). Using a method analogous to the preparation of 17, compound 28 was prepared from 65 and cyclopropanesulfonamide as a colorless solid (0.15 g, 46%). 1H NMR (300 MHz, CDCl3) δ 8.68 (d, J = 16.2 Hz, 1H), 7.59 (d, J = 9.0 Hz, 1H), 6.56 (d, J = 14.4 Hz, 1H), 3.89 (d, J = 6.0 Hz, 2H), 3.15−3.04 (m, 1H), 2.24−1.66 (m, 12H), 1.19−1.10 (m, 2H), 0.98−0.89 (m, 2H), 0.69−0.63 (m, 2H). MS (ES+) m/z: 432.0 (M + 1). 920



Article

5-Cyclopropyl-4-((4,4-difluorocyclohexyl)-methoxy)-2-fluoro-N((2-methoxyethyl)sulfonyl)benzamide (29). Using a method analogous to the preparation of 17, compound 29 was prepared from 65 and 2-methoxyethane-1-sulfonamide as a colorless solid (0.07 g, 26%). 1H NMR (300 MHz, CDCl3) δ 8.62 (d, J = 15.9 Hz, 1H), 7.56 (d, J = 9.0 Hz, 1H), 6.55 (d, J = 14.4 Hz, 1H), 3.91−3.75 (m, 6H), 3.30 (s, 3H), 2.24−1.42 (m, 10H), 0.97−0.88 (m, 2H), 0.71−0.60 (m, 2H). MS (ES+) m/z: 450.0 (M + 1). 5-Cyclopropyl-N-(cyclopropylsulfonyl)-4-((4,4-difluoro-1-methylcyclohexyl)-methoxy)-2-fluorobenzamide (30). Using a method analogous to the preparation of 17, compound 30 was prepared from 66 and cyclopropanesulfonamide as a colorless solid (0.17 g, 61%). mp 134−135 °C. 1H NMR (300 MHz, CDCl3) δ 8.6 6 (d, J = 16.2 Hz, 1H), 7.61 (d, J = 9.0 Hz, 1H), 6.55 (d, J = 14.4 Hz, 1H), 3.74 (s, 2H), 3.14−3.03 (m, 1H), 2.05−1.73 (m, 7H), 1.66−1.59 (m, 2H), 1.48−1.41 (m, 2H), 1.20−1.10 (m, 5H), 0.97−0.89 (m, 2H), 0.68−0.59 (m, 2H). 13C NMR (75 MHz, CDCl3) δ 163.3 (d, JC−F = 11 Hz), 161.1 (d, JC−F = 3 Hz), 160.4 (d, JC−F = 247 Hz), 129.9 (d, JC−F = 2 Hz), 128.8 (d, JC−F = 3 Hz), 123.3 (t, JC−F = 241 Hz), 109.4 (d, JC−F = 10 Hz), 98.9, 98.5, 33.7, 31.5, 30.7 (dd, JC−F = 6.93, 2.82 Hz), 29.6 (t, JC−F = 24.3 Hz), 21.2, 9.4, 7.1, 6.4. MS (ES+) m/z: 446.1 (M + 1). Anal. Calcd for C21H26F3NO4S: C, 56.62; H, 5.88; N, 3.14. Found: C, 56.80; H, 5.89; N, 3.15. 5-Cyclopropyl-N-(cyclopropylsulfonyl)-4-((1-ethyl-4,4difluorocyclohexyl)methoxy)-2-fluorobenzamide (31). Using a method analogous to the preparation of 17, compound 31 was prepared from 67 and cyclopropanesulfonamide as a colorless solid (0.17 g, 37%). 1H NMR (300 MHz, CDCl3) δ 8.68 (d, J = 16.2 Hz, 1H), 7.64 (d, J = 9.0 Hz, 1H), 6.59 (d, J = 14.4 Hz, 1H), 3.79 (s, 2H), 3.15−3.05 (m, 1H), 2.04−1.86 (m, 5H), 1.81−1.57 (m, 6H), 1.50− 1.41 (m, 2H), 1.19−1.10 (m, 2H), 0.98−0.84 (m, 5H), 0.69−0.61 (m, 2H). MS (ES−) m/z: 458.3 (M − 1). 5-Cyclopropyl-N-(cyclopropylsulfonyl)-2-fluoro-4-((1-methyl-4(trifluoromethyl)cyclohexyl)methoxy)benzamide (32). Using a method analogous to the preparation of 17, compound 32 was prepared from 68 and cyclopropanesulfonamide as a colorless solid (0.11 g, 61%). 1H NMR (300 MHz, DMSO-d6) δ 11.82 (s br, 1H), 7.15 (d, J = 8.3 Hz, 1H), 7.09 (d, J = 13.1 Hz, 1H), 3.93 (s, 2H), 3.12−3.03 (m, 1H), 2.30−2.20 (m, 1H), 2.05−1.96 (m, 1H), 1.87− 1.83 (m, 2H), 1.73−1.68 (m, 2H), 1.54−1.41 (m, 2H), 1.34−1.24 (m, 2H), 1.13−1.08 (m, 4H), 1.05 (s, 3H), 0.92−0.85 (m, 2H), 0.69−0.63 (m, 2H). MS (ES+) m/z: 478.1 (M + 1); MS (ES−) m/z: 476.2 (M − 1). 5-Cyclopropyl-N-(cyclopropylsulfonyl)-2-fluoro-4-((6methylspiro[2.5]octan-6-yl)methoxy)benzamide (33). Using a method analogous to the preparation of 17, compound 33 was prepared from 74 and cyclopropanesulfonamide as a colorless solid (0.05 g, 41%). mp 134−135 °C. 1H NMR (300 MHz, CDCl3) δ 8.70 (d, J = 16.3 Hz, 1H), 7.62 (d, J = 9.1 Hz, 1H), 6.59 (d, J = 14.5 Hz, 1H), 3.76 (s, 2H), 3.16−3.06 (m, 1H), 2.09−1.95 (m, 1H), 1.69− 1.40 (m, 7H), 1.28−1.02 (m, 7H), 0.98−0.79 (m, 3H), 0.69−0.62 (m, 2H), 0.32−0.18 (m, 4H). 13C NMR (75 MHz, CDCl3) δ 163.9 (d, JC−F = 11 Hz), 162.2, 161.2 (d, JC−F = 3 Hz), 158.9, 130.0 (d, JC−F = 3 Hz), 128.6 (d, JC−F = 3 Hz), 108.9 (d, JC−F = 10 Hz), 98.7 (d, JC−F = 29 Hz), 34.4, 33.5, 31.5, 30.9, 22.2, 18.7, 11.9, 11.9, 9.4, 7.1, 6.4. MS (ES+) m/z: 436.2 (M + 1); MS (ES−) m/z: 434.2 (M − 1). Anal. Calcd for C23H30FNO4S·0.2H2O: C, 62.91; H, 6.98; N, 3.19. Found: C, 62.73; H, 7.11; N, 3.20. 5-Chloro-2,4-difluoro-N-(methylsulfonyl)benzamide (37). To a solution of 5-chloro-2,4-difluorobenzoic acid (34) (15.0 g, 77.9 mmol) in anhydrous CH2Cl2 (250 mL) were added EDCI (22.6 g, 117.8 mmol), DMAP (21.6 g, 176.8 mmol), and MeSO2NH2 (11.4 g, 119.0 mmol) at room temperature. After stirring at room temperature for 16 h, the reaction mixture was quenched by the addition of 1 M HCl solution (50 mL). The organic layer was separated and washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was crystallized from EtOAc and hexanes (1:4, by volume) to give 37 as a colorless solid (12.3 g, 58%). 1H NMR (300 MHz, DMSO-d6) δ 12.40 (s br, 1H),

8.02−7.97 (m, 1H), 7.76−7.70 (m, 1H), 3.37 (s, 3H). MS (ES−) m/ z: 268.1, 270.1 (M − 1). 3-Chloro-4-fluoro-N-(methylsulfonyl)benzamide (38). Using a method analogous to the preparation of compound 37, 38 was prepared from 5-chloro-4-difluorobenzoic acid (35) and MeSO2NH2 as a colorless solid (13.8 g, 96%). 1H NMR (300 MHz, DMSO-d6) δ 12.27 (s br, 1H), 8.21−8.18 (m, 1H), 8.00−7.95 (m, 1H), 7.62−7.56 (m, 1H), 3.38 (s, 3H). 2,4,5-Trifluoro-N-(methylsulfonyl)benzamide (39). To a solution of MeSO2NH2 (0.49 g, 5.14 mmol) and Et3N (4.3 mL, 30.8 mmol) in anhydrous CH3CN (15 mL) was added 2,4,5-trifluorobenzoyl chloride (36) (1.00 g, 5.14 mmol). After stirring at room temperature for 16 h, the reaction mixture was quenched by the addition of 1 M HCl (20 mL) and extracted with EtOAc (2 × 50 mL). The combined organic extract was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was crystallized from EtOAc and hexanes to give 39 as a light-brown solid (0.7 g, 54%). 1H NMR (300 MHz, CDCl3) δ 8.76 (d, J = 14.2 Hz, 1H), 8.00−7.92 (m, 1H), 7.15−7.06 (m, 1H), 3.43 (s, 3H). MS (ES−) m/z: 252.0 (M − 1). 1-((4-Bromo-2,5-dichlorophenoxy)methyl)adamantine (41). To a mixture of 4-bromo-2,5-dichlorophenol (40) (1.35 g, 5.58 mmol) and K2CO3 (0.85 g, 6.14 mmol) in DMF (10 mL) was added adamantan-1-ylmethyl methanesulfonate (1.50 g, 6.14 mmol) at room temperature. After stirring at 65 and 125 °C for 16 and 48 h, respectively, the reaction mixture was cooled to room temperature, diluted with water (40 mL), and extracted with EtOAc (2 × 50 mL). The combined organic extract was washed with brine (50 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 5−10% EtOAc in hexanes to afford 41 as a colorless solid (1.70 g, 71%). 1H NMR (300 MHz, CDCl3) δ 7.57 (s, 1H), 6. 96 (s, 1H), 3.51 (s, 2H), 2.05−1.97 (m, 3H), 1.81−1.61 (m, 12H). 4-(Adamantan-1-ylmethoxy)-2-fluoro-5-methylbenzoic acid (43). To a solution of 1-adamantane methanol (2.40 g, 14.4 mmol) in anhydrous DMSO (20 mL) was added t-BuOK (4.86 g, 43.3 mmol). After stirring at room temperature for 30 min, 5-methyl-2,4difluorobenzoic acid (42) (2.50 g, 14.40 mmol) was added to the reaction mixture and stirred at 50 °C for 72 h. The reaction mixture was cooled to room temperature and acidified to pH 1 with an icecold 1 M HCl solution, followed by the addition of saturated NH4Cl solution (100 mL). The solid was collected by filtration and washed with H2O. The crude product was crystallized from EtOAc and hexanes to afford 43 as a colorless solid (0.77 g, 17%). 1H NMR (300 MHz, DMSO-d6) δ 12.70 (s br, 1H), 7.64 (d, J = 8.6 Hz, 1H), 6.86 (d, J = 13.2 Hz, 1H), 3.60 (s, 2H), 2.14 (s, 3H), 1.99 (s br, 3H), 1.74−1.63 (m, 12H). MS (ES+) m/z: 437.2 (M + 1). tert-Butyl-5-chloro-2,4-difluorobenzoate (44). To a solution of 34 (10.0 g, 51.93 mmol) and DMAP (0.02 g, 0.16 mmol) in t-BuOH (60 mL) was added di-tert-butyl dicarbonate (23.35 g, 106.98 mmol). The reaction mixture was heated at 50 °C for 16 h and concentrated in vacuo. The residue was diluted with EtOAc (200 mL); washed with 1 M HCl (50 mL), water, and brine; dried over anhydrous Na2SO4; and filtered. The filtrate was concentrated in vacuo to give 44 as a pale yellow oil (12.9 g, 99%). 1H NMR (300 MHz, CDCl3) δ 7.94 (t, J = 7.8 Hz, 1H), 6.95 (t, J = 9.4 Hz, 1H), 1.59 (s, 9H). tert-Butyl-4-(adamantan-1-ylmethoxy)-5-chloro-2-fluorobenzoate (45). To a mixture of 1-adamantane methanol (25.8 g, 0.155 mol) and 44 (50.0 g, 0.201 mol) in DMSO (250 mL) was added Cs2CO3 (109 g, 0.309 mol). The reaction mixture was heated at 70 °C for 20 h, then cooled to room temperature, filtered through a pad of celite, and washed with EtOAc (600 mL). The filtrate was washed with water and brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo. The residue was triturated in methanol (300 mL) and filtered to afford 45 as a pale yellow solid (37.8 g, 62%). 1H NMR (300 MHz, CDCl3) δ 7.84 (d, J = 7.8 Hz, 1H), 6.59 (d, J = 12.3 Hz, 1H), 3.53 (s, 2H), 2.01 (s, 3H), 1.78−1.61 (m, 12H), 1.55 (s, 9H). 921



Article

methanol as a pale yellow oil (5.9 g, 80%). 1H NMR (300 MHz, CDCl3) δ 7.88 (d, J = 7.8 Hz, 1H), 6.62 (d, J = 12.0 Hz, 1H), 3.88 (d, J = 6.0 Hz, 2H), 2.23−1.92 (m, 7H), 1.57 (m, 9H), 1.50−1.42 (m, 2H). tert-Butyl-5-chloro-4-((4,4-difluoro-1-methylcyclohexyl)methoxy)-2-fluorobenzoate (54). Using a method analogous to the preparation of 45, compound 54 was prepared from 44 and (4,4difluoro-1-methylcyclohexyl)methanol as a colorless oil (3.69 g, 51%). 1 H NMR (300 MHz, CDCl3) δ 7.88 (d, J = 7.5 Hz, 1H), 6.62 (d, J = 12.0 Hz, 1H), 3.74 (s, 2H), 2.06−1.55 (m, 17H), 1.16 (s, 3H). MS (ES+) m/z: 393.1 (M + 1). tert-Butyl-5-chloro-4-((1-ethyl-4,4-difluorocyclohexyl)methoxy)2-fluorobenzoate (55). Using a method analogous to the preparation of 45, compound 55 was prepared from 44 and (4,4-difluoro-1ethylcyclohexyl)methanol as a colorless oil (5.97 g, 77%). 1H NMR (300 MHz, CDCl3) δ 7.88 (d, J = 7.8 Hz, 1H), 6.63 (d, J = 11.7 Hz, 1H), 3.78 (s, 2H), 2.02−1.86 (m, 4H), 1.74−1.67 (m, 4H), 1.59− 1.57 (m, 11H), 0.87 (t, J = 7.51 Hz, 3H). MS (ES+) m/z: 351.2 (M − 56). tert-Butyl-5-chloro-2-fluoro-4-((1-methyl-4-(trifluoromethyl)cyclohexyl)methoxy)benzoate (56). Using a method analogous to the preparation of 45, compound 56 was prepared from 44 and (1methyl-4-(trifluoromethyl)cyclohexyl)methanol as a colorless oil (6.20 g, 95%). 1H NMR (300 MHz, CDCl3) δ 7.88 (d, J = 7.7 Hz, 1H), 6.65 (d, J = 12.0 Hz, 1H), 3.85 (s, 2H), 1.96−1.93 (m, 2H), 1.85−1.79 (m, 2H), 1.58 (s, 9H), 1.53−1.39 (m, 3H), 1.34−1.23 (m, 2H), 1.11 (s, 3H). tert-Butyl-4-(adamantan-1-ylmethoxy)-5-cyclopropyl-2-fluorobenzoate (57). To a mixture of 45 (15.8 g, 40.0 mmol), cyclopropylboronic acid (5.16 g, 60.0 mmol), and K3PO4 (38.2 g, 180.0 mmol) in toluene (160 mL) and H2O (8 mL) under a nitrogen atmosphere were added Cy3P·HBF4 (1.47 g, 3.99 mmol) and Pd(OAc)2 (0.45 g, 2.00 mmol). The reaction mixture was heated to 100 °C for 18 h and then cooled to room temperature. H2O (100 mL) was added, and the mixture was extracted with EtOAc (3 × 100 mL); the combined organic extract was washed with brine, dried over anhydrous Na2SO4, and filtrated. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with 5% ethyl acetate in hexanes to afford 57 as a colorless solid (13.8 g, 86%). 1H NMR (300 MHz, CDCl3) δ 7.37 (d, J = 8.4 Hz, 1H), 6.47 (d, J = 12.9 Hz, 1H), 3.49 (s, 2H), 2.05−1.95 (m, 4H), 1.78−1.61 (m, 12H), 1.55 (s, 9H), 0.91−0.84 (m, 2H), 0.64−0.58 (m, 2H). tert-Butyl-4-(cyclohexylmethoxy)-5-cyclopropyl-2-fluorobenzoate (58). Using a method analogous to the preparation of 57, compound 58 was prepared from 52 as a colorless oil (13.73 g, 88%). 1 H NMR (300 MHz, CDCl3) δ 7.36 (d, J = 8.4 Hz, 1H), 6.50 (d, J = 12.9 Hz, 1H), 3.78 (d, J = 5.7 Hz, 2H), 2.07−1.98 (m, 1H), 1.92− 1.69 (m, 6H), 1.56 (s, 9H), 1.39−1.02 (m, 5H), 0.93−0.85 (m, 2H), 0.67−0.61 (m, 2H). MS (ES+) m/z: 387.2 (M + 39). tert-Butyl-5-cyclopropyl-4-((4,4-difluorocyclohexyl)methoxy)-2fluorobenzoate (59). Using a method analogous to the preparation of 57, compound 59 was prepared from 53 and cyclopropylboronic acid as a pale yellow oil (5.37 g, 86%). 1H NMR (300 MHz, CDCl3) δ 7.39 (d, J = 8.4 Hz, 1H), 6.49 (d, J = 12.6 Hz, 1H), 3.85 (d, J = 5.7 Hz, 2H), 2.24−1.91 (m, 8H), 1.59−1.47 (m, 11H), 0.94−0.84 (m, 2H), 0.70−0.58 (m, 2H). MS (ES−) m/z: 383.0 (M − 1). tert-Butyl-5-cyclopropyl-4-((4,4-difluoro-1-methylcyclohexyl)methoxy)-2-fluorobenzoate (60). Using a method analogous to the preparation of 57, compound 60 was prepared from 54 and cyclopropylboronic acid as a colorless oil (3.69 g, 99%). 1H NMR (300 MHz,CDCl3) δ 7.42 (d, J = 8.4 Hz, 1H), 6.50 (d, J = 12.6 Hz, 1H), 3.72 (s, 2H), 2.06−1.73 (m, 8H), 1.65−1.62 (m, 1H), 1.57 (s, 9H), 1.15 (s, 3H), 0.94−0.85 (m, 2H), 0.66−0.59 (m, 2H). MS (ES +) m/z: 399.2 (M + 1). tert-Butyl-5-cyclopropyl-4-((1-ethyl-4,4-difluorocyclohexyl)methoxy)-2-fluorobenzoate (61). Using a method analogous to the preparation of 57, compound 61 was prepared from 55 and cyclopropylboronic acid as a colorless oil (5.66 g, 94%). 1H NMR (300 MHz, CDCl3) δ 7.41 (d, J = 8.4 Hz, 1H), 6.50 (d, J = 12.6 Hz,

tert-Butyl-4-(adamantan-1-ylmethoxy)-2-fluoro-5-vinylbenzoate (46). To a mixture of 45 (2.00 g, 5.06 mmol), vinylboronic acid pinacol ester (1.56 g, 10.1 mmol), and Na2CO3 (1.61 g, 15.2 mmol) in 1,4-dioxane (20 mL) and water (5 mL) were added t-Bu3P·HBF4 (0.15 g, 0.51 mmol) and Pd2(dba)3 (0.023 g, 0.025 mmol) under a nitrogen atmosphere. The reaction mixture was heated to 100 °C for 24 h and then cooled to room temperature. Water (50 mL) was added and the mixture was extracted with EtOAc (3 × 100 mL). The combined organic extract was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified over column chromatography eluted with 30% CH2Cl2 in hexanes to give 46 as a pale yellow solid (1.80 g, 92%). 1H NMR (300 MHz, CDCl3) δ 7.95 (d, J = 9.0 Hz, 1H), 6.98−6.86 (m, 1H), 6.52 (d, J = 12.0 Hz, 1H), 5.76 (d, J = 18.0 Hz, 1H), 5.26 (d, J = 9.0 Hz, 1H), 3.51 (s, 2H), 1.78−1.62 (m, 12H), 1.58−1.52 (m, 3H), 1.22 (s, 9H). MS (ES+) m/z: 387.2 (M + 1). 4-(Adamantan-1-ylmethoxy)-2-fluoro-5-vinylbenzoic Acid (47). To a solution of 46 (1.80 g, 4.65 mmol) in CH2Cl2 (10 mL) was added TFA (10 mL). The reaction mixture was stirred at room temperature for 16 h and then concentrated in vacuo. The residue was triturated in hexanes (50 mL), and the solid was collected by filtration to give 47 as a pale yellow solid (1.50 g, 97%). 1H NMR (300 MHz, DMSO-d6) δ 12.94 (s br, 1H), 7.92 (d, J = 8.6 Hz, 1H), 6.96−6.81 (m, 2H), 5.84−5.78 (m, 1H), 5.32−5.28 (m, 1H), 3.62 (s, 2H), 1.95 (s, 3H), 1.70−1.60 (m, 12H). MS (ES+) m/z: 331.2 (M + 1); MS (ES−) m/z: 329.2 (M − 1). 4-(Adamantan-1-ylmethoxy)-5-ethyl-2-fluorobenzoic Acid (48). A mixture of 47 (1.00 g, 3.03 mmol) in EtOAc (50 mL) was hydrogenated over 10% palladium on activated carbon (0.10 g) with atmospheric hydrogen at room temperature for 42 h. The mixture was filtered through a pad of celite and washed with EtOAc (50 mL). The filtrate was concentrated in vacuo, and the residue was recrystallized from EtOAc and hexanes to afford 48 as a colorless solid (0.75 g, 74%). 1H NMR (300 MHz, DMSO-d6) δ 12.80 (s br, 1H), 7.63 (d, J = 8.6 Hz, 1H), 6.87 (d, J = 13.2 Hz, 1H), 3.61 (s, 2H), 5.57 (d, J = 7.5 Hz, 2H), 1.99 (s, 3H), 1.75−1.63 (m, 12H), 1.14 (t, J = 7.5 Hz, 3H). MS (ES+) m/z: 333.2 (M + 1). tert-Butyl-4-(adamantan-1-ylmethoxy)-2-fluoro-5-(prop-1-en-2yl)benzoate (49). Using a method analogous to the preparation of 46, compound 49 was prepared from 45 and 2-isopropenyl-4,4,5,5tetramethyl-1,3,2-dioxaborolane as a colorless solid (1.94 g, 64%). 1H NMR (300 MHz, CDCl3) δ 7.69 (d, J = 8.7 Hz, 1H), 6.54 (d, J = 13.2 Hz, 1H), 5.17−5.13 (m, 1H), 5.10−5.07 (m, 1H), 3.50 (s, 2H), 2.15 (s, br, 3H), 2.02 (s br, 3H), 1.81−1.62 (m, 12H), 1.58 (s, 9H). MS (ES+) m/z: 401.2 (M + 1). tert-Butyl-4-(adamantan-1-ylmethoxy)-2-fluoro-5-isopropylbenzoate (50). A mixture of 49 (1.94 g, 4.84 mmol) in EtOAc (50 mL) was hydrogenated over 10% palladium on activated carbon (0.19 g) with atmospheric hydrogen at room temperature for 16 h, filtered through a pad of celite, and concentrated in vacuo to afford 50 as a pale yellow oil (1.84 g, 94%). 1H NMR (300 MHz, CDCl3) δ 7.69 (d, J = 8.7 Hz, 1H), 6.50 (d, J = 12.9 Hz, 1H), 3.49 (s, 2H), 3.30−3.18 (m, 1H), 2.04 (s br, 3H), 1.83−1.64 (m, 12H), 1.58 (s, 9H), 1.22 (d, J = 6.9 Hz, 1H). MS (ES+) m/z: 403.3 (M + 1). 4-(Adamantan-1-ylmethoxy)-2-fluoro-5-isopropylbenzoic Acid (51). Using a method analogous to the preparation of 47, compound 51 was prepared from 50 as a colorless solid (1.31 g, 84 %). 1H NMR (300 MHz, DMSO-d6) δ 12.82 (s br, 1H), 7.61 (d, J = 8.7 Hz, 1H), 6.84 (d, J = 13.2 Hz, 1H), 3.58 (s, 2H), 3.22−3.10 (m, 1H), 1.96 (s, 3H), 1.75−1.59 (m, 12H), 1.15 (d, J = 6.9 Hz, 6H). MS (ES+) m/z: 347.2 (M + 1). tert-Butyl-5-chloro-4-(cyclohexylmethoxy)-2-fluorobenzoate (52). Using a method analogous to the preparation of 45, compound 52 was prepared from 44 and cyclohexylmethanol as a colorless oil (15.37 g, 43%). 1H NMR (300 MHz, CDCl3) δ 7.86 (d, J = 7.5 Hz, 1H), 6.61 (d, J = 12.0 Hz, 1H), 3.81 (d, J = 6.0 Hz, 2H), 1.90−1.68 (m, 6H), 1.57 (s, 9H), 1.39−1.04 (m, 5H). tert-Butyl-5-cyclopropyl-4-((4,4-difluoro-cyclohexyl)methoxy)-2fluorobenzoate (53). Using a method analogous to the preparation of 45, compound 53 was prepared from 44 and (4,4-difluorocyclohexyl)922



Article

1H), 3.74 (s, 2H), 2.02−1.86 (m, 5H), 1.82−1.60 (m, 6H), 1.55 (s, 9H), 0.92−0.82 (m, 5H), 0.65−0.56 (m, 2H). MS (ES+) m/z: 413.1 (M + 1). tert-Butyl 5-cyclopropyl-2-fluoro-4-((1-methyl-4(trifluoromethyl)cyclohexyl)methoxy)benzoate (62). Using a method analogous to the preparation of 45, compound 62 was prepared from 56 and cyclopropylboronic acid as a pale yellow oil (1.66 g, 96%). 1H NMR (300 MHz, CDCl3) δ 7.42 (d, J = 8.4 Hz, 1H), 6.53 (d, J = 12.6 Hz, 1H), 3.81 (s, 2H), 2.00−1.95 (m, 3H), 1.84−1.79 (m, 2H), 1.57 (s, 9H), 1.55−1.41 (m, 3H), 1.33−1.22 (m, 2H), 1.11 (s, 3H), 0.91−0.85 (m, 2H), 0.64−0.59 (m, 2H). 4-(Adamantan-1-ylmethoxy)-5-cyclopropyl-2-fluorobenzoic Acid (63). Using a method analogous to the preparation of 47, compound 63 was prepared from 57 as a colorless solid (10.1 g, 85%). 1H NMR (300 MHz, DMSO-d6) δ 12.77 (s br, 1H), 7.29 (d, J = 8.4 Hz, 1H), 6.83 (d, J = 13.2 Hz, 1H), 3.59 (s, 2H), 2.04−1.92 (m, 4H), 1.71−1.58 (m, 12H), 0.91−0.83 (m, 2H), 0.59−0.52 (m, 2H). 4-(Cyclohexylmethoxy)-5-cyclopropyl-2-fluorobenzoic Acid (64). Using a method analogous to the preparation of 47, compound 64 was prepared from 58 as a colorless solid (6.64 g, 58%). 1H NMR (300 MHz, DMSO-d6) δ 12.79 (s br, 1H), 7.29 (d, J = 8.4 Hz, 1H), 6.88 (d, J = 13.2 Hz, 1H), 3.88 (d, J = 5.7 Hz, 2H), 2.06−1.96 (m, 1H), 1.86−1.62 (m, 6H), 1.34−1.02 (m, 5H), 0.93−0.85 (m, 2H), 0.62−0.55 (m, 2H). MS (ES−) m/z: 291.3 (M − 1). 5-Cyclopropyl-4-((4,4-difluorocyclohexyl)-methoxy)-2-fluorobenzoic Acid (65). Using a method analogous to the preparation of 47, compound 65 was prepared from 59 as a colorless solid (2.29 g, 50%). 1H NMR (300 MHz, DMSO-d6) δ 12.83 (s br, 1H), 7.30 (d, J = 8.4 Hz, 1H), 6.91 (d, J = 13.2 Hz, 1H), 3.96 (d, J = 6.0 Hz, 2H), 2.10−1.74 (m, 8H), 1.47−1.33 (m, 2H), 0.92−0.84 (m, 2H), 0.61− 0.54 (m, 2H). MS (ES+) m/z: 329.1 (M + 1). 5-Cyclopropyl-4-((4,4-difluoro-1-methylcyclohexyl)methoxy)-2fluorobenzoic Acid (66). Using a method analogous to the preparation of 47, compound 66 was prepared from 60 as a colorless solid (2.74 g, 86%). 1H NMR (300 MHz, CDCl3) δ 7.56 (d, J = 8.4 Hz, 1H), 6.57 (d, J = 12.9 Hz, 1H), 3.75 (s, 2H), 2.08−1.75 (m, 7H), 1.66−1.55 (m, 2H), 1.16 (s, 3H), 0.97−0.88 (m, 2H), 0.68−0.61 (m, 2H). MS (ES+) m/z: 343.2 (M + 1). 5-Cyclopropyl-4-((1-ethyl-4,4-difluorocyclohexyl)methoxy)-2-fluorobenzoic Acid (67). Using a method analogous to the preparation of 47, compound 67 was prepared from 61 as a colorless solid (3.497 g, 72%). 1H NMR (300 MHz, CDCl3) δ 9.31 (s br, 1H), 7.55 (d, J = 8.4 Hz, 1H), 6.57 (d, J = 12.6 Hz, 1H), 3.77 (s, 2H), 2.02−1.85 (m, 5H), 1.82−1.57 (m, 6H), 0.96−0.84 (m, 5H), 0.67−0.59 (m, 2H). MS (ES+) m/z: 357.2 (M + 1). 5-Cyclopropyl-2-fluoro-4-((1-methyl-4-(trifluoromethyl)cyclohexyl)methoxy)benzoic Acid (68). Using a method analogous to the preparation of 47, compound 68 was prepared from 62 as a colorless solid (1.30 g, 81%). 1H NMR (300 MHz, DMSO-d6) δ 12.84 (s br, 1H), 7.33 (d, J = 8.5 Hz, 1H), 7.02 (d, J = 13.2 Hz, 1H), 3.92 (s, 2H), 2.26−2.20 (m, 1H), 2.04−1.95 (m, 1H), 1.87−1.83 (m, 2H), 1.72−1.67 (m, 2H), 1.54−1.41 (m, 2H), 1.34−1.23 (m, 2H), 1.05 (s, 3H), 0.91−0.85 (m, 2H), 0.60−0.55 (m, 2H). MS (ES−) m/ z: 373.2 (M − 1). tert-Butyl-5-chloro-2-fluoro-4-((8-methyl-1,4-dioxaspiro[4.5]decan-8-yl)methoxy)benzoate (69). Using a method analogous to the preparation of 45, compound 69 was prepared from 44 and (8methyl-1,4-dioxaspiro[4.5]decan-8-yl)methanol as a colorless oil (5.13 g, 54%). 1H NMR (300 MHz, CDCl3) δ 7.87 (d, J = 7.9 Hz, 1H), 6.62 (d, J = 12.3 Hz, 1H), 3.95 (s, 4H), 3.74 (s, 2H), 1.76−1.60 (m, 8H), 1.58 (s, 9H), 1.15 (s, 3H). MS (ES+) m/z: 359.1 (M − 55). tert-Butyl-5-cyclopropyl-2-fluoro-4-((8-methyl-1,4-dioxaspiro[4.5]decan-8-yl)methoxy)benzoate (70). Using a method analogous to the preparation of 57, compound 70 was prepared from 69 and cyclopropylboronic acid as a colorless oil (2.59 g, 83%). 1H NMR (300 MHz, CDCl3) δ 7.41 (d, J = 8.4 Hz, 1H), 6.50 (d, J = 12.7 Hz, 1H), 3.95 (s, 4H), 3.71 (s, 2H), 2.05−1.94 (m, 1H), 1.77−1.60 (m, 8H), 1.57 (s, 9H), 1.14 (s, 3H), 0.93−0.85 (m, 2H), 0.65−0.59 (m, 2H). MS (ES+) m/z: 365.2 (M − 55). tert-Butyl-5-cyclopropyl-2-fluoro-4-((1-methyl-4-oxocyclohexyl)methoxy)benzoate (71). To a solution of 70 (2.59 g, 6.16 mmol) in

THF (6 mL) and H2O (4.3 mL) was added TFA (2.2 mL). After stirring at room temperature for 18 h, the reaction was quenched by the addition of a 2 M aqueous NaOH solution (15 mL) and then extracted with EtOAc (3 × 50 mL). The combined organic extract was washed with H2O and brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 0−15% EtOAc in hexanes to afford 71 as a colorless oil (2.09 g, 90%). 1 H NMR (300 MHz, CDCl3) δ 7.42 (d, J = 8.4 Hz, 1H), 6.53 (d, J = 12.5 Hz, 1H), 3.82 (s, 2H), 2.56−2.36 (m, 4H), 2.05−1.78 (m, 5H), 1.57 (s, 9H), 1.30 (s, 3H), 0.92−0.86 (m, 2H), 0.65−0.60 (m, 2H). MS (ES+) m/z: 321.2 (M − 55). tert-Butyl-5-cyclopropyl-2-fluoro-4-((1-methyl-4methylenecyclohexyl)methoxy)benzoate (72). To a cooled (−20 °C) suspension of methyltriphenylphosphonium bromide (1.44 g, 4.02 mmol) in THF (12 mL) was added LiHMDS (1.0 M solution in THF, 4.0 mL, 4.00 mmol). After stirring at −20 °C for 90 min, a solution of 71 (1.00 g, 2.68 mmol) in THF (6 mL) was added. The reaction mixture was slowly warmed to room temperature, stirred for 12 h, quenched by the addition of saturated aqueous NH4Cl solution (20 mL), and diluted with ethyl acetate (100 mL). The organic layer was separated, and the aqueous layer was extracted with EtOAc (3 × 30 mL). The combined organic extract was washed with brine, dried over Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 0−5% EtOAc in hexanes to afford 72 as a colorless oil (0.83 g, 82%). 1H NMR (300 MHz, CDCl3) δ 7.41 (d, J = 8.4 Hz, 1H), 6.51 (d, J = 12.7 Hz, 1H), 4.65 (s, 2H), 3.71 (s, 2H), 2.30−2.15 (m, 4H), 2.05−1.93 (m, 1H), 1.68−1.50 (m, 13H), 1.15 (s, 3H), 0.93−0.86 (m, 2H), 0.66−0.60 (m, 2H). MS (ES+) m/z: 319.1 (M − 55). tert-Butyl-5-cyclopropyl-2-fluoro-4-((6-methylspiro[2.5]octan-6yl)methoxy)benzoate (73). To a cooled (0 °C) solution of 72 (0.30 g, 0.80 mmol) and chloroiodomethane (0.21 mL, 2.65 mmol) in 1,2dichloroethane (2 mL) was added diethylzinc (1.0 M solution in hexanes, 1.33 mL, 1.33 mmol). After stirring at 0 °C for 2 h, the reaction was quenched by the addition of a 1 N HCl solution (5 mL) and extracted with CH2Cl2 (3 × 30 mL). The combined organic extract was dried over Na2SO4 and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography eluted with a gradient of 0−5% EtOAc in hexanes to afford 73 as a colorless oil (0.29 g, 93%). 1H NMR (300 MHz, CDCl3) δ 7.41 (d, J = 8.5 Hz, 1H), 6.52 (d, J = 12.8 Hz, 1H), 3.72 (s, 2H), 2.09−1.94 (m, 1H), 1.73−1.42 (m, 14H), 1.19−1.03 (m, 5H), 0.97−0.84 (m, 3H), 0.70−0.59 (m, 2H), 0.33−0.16 (m, 4H). MS (ES+) m/z: 333.2 (M − 55). 5-Cyclopropyl-2-fluoro-4-((6-methylspiro[2.5]octan-6-yl)methoxy)benzoic Acid (74). To a cooled (0 °C) solution of 73 (0.28 g, 0.71 mmol) and anisole (0.11 mL, 1.07 mmol) in CH2Cl2 (3 mL) was added TFA (0.75 mL, 9.61 mmol). After stirring for 4 h at 0 °C, the reaction mixture was diluted with CH2Cl2 (20 mL) and washed with H2O (5 × 10 mL) until the last wash was neutral as monitored by pH paper. The organic layer was dried over anhydrous Na2SO4 sulfate and filtered. The filtrate was concentrated in vacuo to afford 74 as a colorless solid (0.21 g, 90%). 1H NMR (300 MHz, CDCl3) δ 7.48 (d, J = 8.5 Hz, 1H), 6.55 (d, J = 13.0 Hz, 1H), 3.71 (s, 2H), 2.08− 1.90 (m, 1H), 1.78−1.38 (m, 6H), 1.17−1.02 (m, 4H), 0.95−0.80 (m, 3H), 0.68−0.56 (m, 2H), 0.33−0.13 (m, 4H). MS (ES−) m/z: 331.3 (M − 1). Molecular Modeling. Induced fit docking of compound 7 to the hNaV1.7 VSD4 active site: using the X-ray structure of NaV1.726 as the source, chain B of the protein was first isolated and prepped using the Protein Preparation Wizard.46 Compound 7 was docked into the VSD4 active site via induced fit docking46b,47 using default settings, which optimized side chains within 5 Å of the ligand in addition to docking. The centroid of the cocrystal structure ligand GX-936 was used as the center of the grid box. A single receptor-ligand model was chosen from the resulting set through visual inspection. Compound 7 was then minimized in the active site. 923


Journal of Medicinal Chemistry Use of Animal Subjects. All studies involving animals were either approved by the Animal Care Committee of Xenon Pharmaceuticals (Burnaby, BC, Canada) and performed in compliance with the Canadian Council on Animal Care guidelines (mouse efficacy and mouse PK studies) or approved by and conformed to the guidelines and principles set by the Institutional Animal Care and Use Committee of Genentech.

■

ABBREVIATIONS

■

REFERENCES

AUC, area under curve; BCRP, breast cancer resistance protein; CIP, congenital insensitivity to pain; Cmax, maximal concentration; CL, plasma clearance; CLhep, hepatic clearance; cLogP, calculated logarithm of partition coefficient; CYP, cytochromes P450; DMAP, 4-dimethylaminopyridine; DMF, dimethylformamide; DMSO, dimethyl sulfoxide; DRG, dorsal root ganglion; EDCI, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide; EP, electrophysiology; F, oral bioavailability; HEK, human embryonic kidney; HLM, human liver microsome; IEM, inherited erythromelalgia; LBA, ligand binding assay; LLE, lipophilic ligand efficiency; MCT, 0.5% w/w methylcellulose and 0.2% v/v Tween-80 in water; MDCKI, Madin−Darby canine kidney cell line I; MDRI, multidrugresistant protein I, also refer to P-glycoprotein; MetID, metabolite identification; mLogD, measured distribution coefficient at pH 7.2; NaV1.7, voltage-gated sodium channel type 7; NaV1.5, voltage-gated sodium channel type 5; Pd2(dba)3, tris(dibenzylideneacetone)dipalladium(0); PK, pharmacokinetic; PPB, plasma protein binding; SCN9A, gene encoding the sodium channel NaV1.7; SAR, structure−activity relationship; TFA, trifluoroacetic acid; t1/2, terminal half-life; VSD4, voltage sensing domain 4; Vss, volume of distribution

ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jmedchem.8b01621. Description of in vitro biological assays, in vivo rodent pain assays, acute dissociation of mouse DRG neuron and patch clamp recording, CEREP screening data for compound 33, 1H NMR spectra of compounds 7−33, HPLC purity data for compounds 7, 20, 26, 30, and 33, and experimental procedure and small-molecule X-ray coordinates for compound 7 (PDF)

■

■

Article

Molecular formula strings (CSV)

AUTHOR INFORMATION

Corresponding Authors

*E-mail: cdehnhardt@xenon-pharma. Phone 604-679-9736 (C.M.D.). *E-mail: [email protected]. Phone 650-225-2749 (D.F.O.).

(1) Mulroy, M. F. Systemic toxicity and cardiotoxicity from local anesthetics: incidence and preventive measures. Reg. Anesth. Pain Med. 2002, 27, 556−561. (2) Walia, K. S.; Khan, E. A.; Ko, D. H.; Raza, S. S.; Khan, Y. N. Side effects of antiepileptics − a review. Pain Pract. 2004, 4, 194−203. (3) Bhattachacharya, A.; Wickenden, A. D.; Chaplan, S. R. Sodium channel blockers for the treatment of neuropathic pain. Neurotherapeutics 2009, 6, 663−678. (4) (a) Dib-Hajj, S. D.; Cummins, T. R.; Black, J. A.; Waxman, S. G. From genes to pain: NaV1.7 and human pain disorders. Trends Neurosci. 2007, 30, 555−563. (b) Dib-Hajj, S. D.; Yang, Y.; Black, J. A.; Waxman, S. G. The NaV1.7 sodium channel: from molecule to man. Nat. Rev. Neurosci. 2013, 14, 49−62. (c) Waxman, S. G.; Zamponi, G. W. Regulating excitability of peripheral afferents: emerging ion channel targets. Nat. Neurosci. 2014, 17, 153−163. (5) Nagasako, E. M.; Oaklander, A. L.; Dworkin, R. H. Congenital insensitivity to pain: an update. Pain 2003, 101, 213−219. (6) Cox, J. J.; Reimann, F.; Nicolas, A. K.; Thornton, G.; Roberts, E.; Springell, K.; Karbani, G.; Jafri, H.; Mannan, J.; Raashid, Y.; Al-Gazali, L.; Hamamy, H.; Valente, E. M.; Gorman, S.; Williams, R.; McHale, D. P.; Wood, J. N.; Gribble, F. M.; Woods, C. G. An SCN9A channelopathy causes congenital inability to experience pain. Nature 2006, 444, 894−898. (7) Goldberg, Y. P.; MacFarlane, J.; MacDonald, M. L.; Thompson, J.; Dube, M.-P.; Mattice, M.; Fraser, R.; Young, C.; Hossain, S.; Pape, T.; Payne, B.; Radomski, C.; Donaldson, G.; Oves, E.; Cox, J.; Younghusband, H. B.; Green, R.; Duff, A.; Boltshauser, E.; Grinspan, G. A.; Dimon, J. H.; Sibley, B. G.; Andria, G.; Toscano, E.; Kerdraon, J.; Bowsher, D.; Pimstone, S. N.; Samuels, M. E.; Sherrington, R.; Hayden, M. R. Loss-of-function mutations in the NaV1.7 gene underlie congenital indifference to pain in multiple human populations. Clin. Genet. 2007, 71, 311−319. (8) Ahmad, S.; Dahllund, L.; Eriksson, A. B.; Hellgren, D.; Karlsson, U.; Lund, P.-E.; Meijer, I. A.; Meury, L.; Mills, T.; Moody, A.; Morinville, A.; Morten, J.; O’Donnell, D.; Raynoschek, C.; Salter, H.; Rouleau, G. A.; Krupp, J. J. A stop codon mutation in SCN9A causes lack of pain sensation. Hum. Mol. Genet. 2007, 16, 2114−2121. (9) Nilsen, K. B.; Nicholas, A. K.; Woods, C. G.; Mellgren, S. I.; Nebuchennykh, M.; Aasly, J. Two novel SCN9A mutations causing insensitivity to pain. Pain 2009, 143, 155−158.

ORCID

Thilo Focken: 0000-0003-1993-2476 Benjamin D. Sellers: 0000-0002-4194-1240 Brian S. Safina: 0000-0001-8134-9949 Christoph M. Dehnhardt: 0000-0001-5213-6354 Present Addresses ∥

Syrachem Sciences Inc., 7600 Glover Road, Langley, British Columbia V2Y 1Y1, Canada (S.C.). ⊥ Inception Sciences Canada, 887 Great Northern Way, Suite 210, Vancouver, British Columbia V5T 4T5, Canada (T.S.). # WuXi AppTec Co., Ltd., 288 FuTe Zhong Road, Shanghai 200131, P. R. China (C.-A.C.). ∇ College of Pharmacy, University of Michigan, 428 Church Street, Ann Arbor, Michigan 48109-1065, United States (A.W.). Author Contributions

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS The authors thank Eric Kam for assistance in the preparation of compounds 9, 12, and 13; Noah Stuart and Sam Goodchild for additional EP data; and Rhena Yoo for assistance in ligand binding assay test. They also thank Bruce Roth, Tarek S. Mansour, Renata M. Oballa, and the Xenon-Genentech JRC for their leadership and support. In addition, they thank James Empfield and Steven Wesolowski for helpful revisions of this manuscript. 924



Article

for use as intravenous agents and chemical probes. Bioorg. Med. Chem. Lett. 2017, 27, 4805−4811. (d) McDonnell, A.; Collins, S.; Ali, Z.; Iavarone, L.; Surujbally, R.; Kirby, S.; Butt, R. P. Efficacy of the NaV1.7 blocker PF-05089771 in a randomized, placebo-controlled, double-blind clinical study in subjects with painful diabetic peripheral neuropathy. Pain 2018, 159, 1465−1476. (21) (a) Sun, S.; Jia, Q.; Zenova, A. Y.; Chafeev, M.; Zhang, Z.; Lin, S.; Kwan, R.; Grimwood, M. E.; Chowdhury, S.; Young, C.; Cohen, C. J.; Oballa, R. M. The discovery of benzenesulfonamide-based potent and selective inhibitors of voltage-gated sodium channel NaV1.7. Bioorg. Med. Chem. Lett. 2014, 24, 4397−4401. (b) Focken, T.; Liu, S.; Chahal, N.; Dauphinais, M.; Grimwood, M. E.; Chowdhury, S.; Hemeon, I.; Bichler, P.; Bogucki, D.; Waldbrook, M.; Bankar, G.; Sojo, L. E.; Young, C.; Lin, S.; Shuart, N.; Kwan, R.; Pang, J.; Chang, J. H.; Safina, B. S.; Sutherlin, D. P.; Johnson, J. P., Jr.; Dehnhardt, C. M.; Mansour, T. S.; Oballa, R. M.; Cohen, C. J.; Robinette, C. L. Discovery of aryl sulfonamides as isoform-selective inhibitors of NaV1.7 with efficacy in rodent pain models. ACS Med. Chem. Lett. 2016, 7, 77−82. (c) Focken, T.; Chowdhury, S.; Zenova, A.; Grimwood, M.; Chabot, C.; Sheng, T.; Hemeon, I.; Decker, S.; Wilson, M.; Bichler, P.; Jia, Q.; Sun, S.; Young, C.; Lin, S.; Goodchild, S. J.; Shuart, N. G.; Chang, E.; Xie, Z.; Li, B.; Khakh, K.; Bankar, G.; Waldbrook, M.; Kwan, R.; Nelkenbrecher, K.; Tari, P. K.; Chahal, N.; Sojo, L.; Robinette, L.; White, A.; Chen, C.; Zhang, Y.; Pang, J.; Chang, J. H.; Hackos, D.; Johnson, J. P., Jr.; Cohen, C. J.; Ortwine, D.; Sutherlin, D.; Dehnhardt, C.; Safina, B. Design of conformationally constrained acyl sulfonamide isosteres: Identification of N([1,2,4]triazolo[4,3-a]pyridin-3-yl)methane-sulfonamides as potent and selective hNaV1.7 inhibitors for the treatment of pain. J. Med. Chem. 2018, 61, 4810−4831. (22) (a) DiMauro, E. F.; Altmann, S.; Berry, L. M.; Bregman, H.; Chakka, N.; Chu-Moyer, M.; Bojic, E. F.; Foti, R. S.; Fremeau, R.; Gao, H.; Gunaydin, H.; Guzman-Perez, A.; Hall, B. E.; Huang, H.; Jarosh, M.; Kornecook, T.; Lee, J.; Ligutti, J.; Liu, D.; Moyer, B. D.; Ortuno, D.; Rose, P. E.; Schenkel, L. B.; Taborn, K.; Wang, J.; Wang, Y.; Yu, V.; Weiss, M. M. Application of a parallel synthetic strategy in the discovery of biaryl acyl sulfonamides as efficient and selective NaV1.7 inhibitors. J. Med. Chem. 2016, 59, 7818−7839. (b) Marx, I. E.; Dineen, T. A.; Able, J.; Bode, C.; Bregman, H.; Chu-Moyer, M.; DiMauro, E. F.; Du, B.; Foti, R. S.; Fremeau, R. T., Jr.; Gao, H.; Gunaydin, H.; Hall, B. E.; Huang, L.; Kornecook, T.; Kreiman, C. R.; La, D. S.; Ligutti, J.; Lin, M. J.; Liu, D.; McDermott, J. S.; Moyer, B. D.; Peterson, E. A.; Roberts, J. T.; Rose, P.; Wang, J.; Youngblood, B. D.; Yu, V.; Weiss, M. M. Sulfonamides as selective NaV1.7 inhibitors: Optimizing potency and pharmacokinetics to enable in vivo target engagement. ACS Med. Chem. Lett. 2016, 7, 1062−1067. (c) Weiss, M. M.; Dineen, T. A.; Marx, I. E.; Altmann, S.; Boezio, A. A.; Bregman, H.; Chu-Moyer, M.; DiMauro, E. F.; Bojic, E. F.; Foti, R. S.; Gao, H.; Graceffa, R. F.; Gunaydin, H.; Guzman-Perez, A.; Huang, H.; Huang, L.; Jarosh, M.; Kornecook, T.; Kreiman, C. R.; Ligutti, J.; La, D. S.; Lin, M. J.; Liu, D.; Moyer, B. D.; Nguyen, H. N.; Peterson, E. A.; Rose, P. E.; Taborn, K.; Youngblood, B. D.; Yu, V.; Fremeau, R. T., Jr. Sulfonamides as selective NaV1.7 inhibitors: Optimizing potency and pharmacokinetics while mitigating metabolic liabilities. J. Med. Chem. 2017, 60, 5969−5989. (d) Graceffa, R. F.; Boezio, A. A.; Able, J.; Altmann, S.; Berry, L. M.; Boezio, C.; Butler, J. R.; ChuMoyer, M.; Cooke, M.; DiMauro, E. F.; Dineen, T. A.; Bojic, E. F.; Foti, R. S.; Fremeau, R. T., Jr.; Guzman-Perez, A.; Gao, H.; Gunaydin, H.; Huang, H.; Huang, L.; Ilch, C.; Jarosh, M.; Kornecook, T.; Kreiman, C. R.; La, D. S.; Ligutti, J.; Milgram, B. C.; Lin, M. J.; Marx, I. E.; Nguyen, H. N.; Peterson, E. A.; Rescourio, G.; Roberts, J.; Schenkel, L.; Shimanovich, R.; Sparling, B. A.; Stellwagen, J.; Taborn, K.; Vaida, K. R.; Wang, J.; Yeoman, J.; Yu, V.; Zhu, D.; Moyer, B. D.; Weiss, M. M. Sulfonamides as selective NaV1.7 inhibitors: Optimizing potency, pharmacokinetics, and metabolic properties to obtain atropisomeric quinolinone (AM-0466) that affords robust in vivo Activity. J. Med. Chem. 2017, 60, 5990−6017. (e) La, D. S.; Peterson, E. A.; Bode, C.; Boezio, A. A.; Bregman, H.; Chu-Moyer, M. Y.; Coats, J.; DiMauro, E. F.; Dineen, T. A.; Du, B.; Gao, H.; Graceffa, R.;

(10) Cox, J. J.; Sheynin, J.; Shorer, Z.; Reimann, F.; Nicholasm, A. K.; Zubovic, L.; Baralle, M.; Wraige, E.; Manor, E.; Levy, J.; Woods, C. G.; Parvari, R. Congenital insensitivity to pain: novel SCN9A missense and in-frame deletion mutations. Hum. Mutat. 2010, 31, E1670−E1686. (11) Bogdanova-Mihaylova, P.; Alexander, M. D.; Murphy, R. P. J.; Murphy, S. M. SCN9A-associated congenital insensitivity to pain and anosmia in an Irish patient. J. Peripher. Nerv. Syst. 2015, 20, 86−87. (12) Drenth, J. P.; Finley, W. H.; Breedveld, G. L.; Testers, L.; Michiels, J. J.; Guillet, G.; Taieb, A.; Kirby, R. L.; Heutink, P. The primary erythemyalgia-susceptibility gene is located on chromosome 2q31-32. Am. J. Hum. Genet. 2001, 68, 1277−1282. (13) Yang, Y.; Wang, Y.; Li, S.; Xe, Z.; Li, H.; Ma, L.; Fan, J.; Bu, D.; Liu, B.; Fan, Z.; Wu, G.; Jin, J.; Ding, B.; Zhu, X.; Shen, Y. Mutations in SCN9A, encoding a sodium channel alpha subunit, in patients with primary erythermalgia. J. Med. Genet. 2004, 41, 171−174. (14) Dib-Hajj, S. D.; Rush, A. M.; Cummins, T. R.; Hisama, F. M.; Novella, S.; Tyrrell, L.; Marshall, L.; Waxman, S. G. Gain-of-function mutation in NaV1.7 in familial erythromelalgia induces bursting of sensory neurons. Brain 2005, 128, 1847−1854. (15) Dib-Hajj, S. D.; Rush, A. M.; Cummins, T. R.; Waxman, S. G. Mutation in the sodium channel NaV1.7 underlie inherited erythromelalgia. Drug Discovery Today: Dis. Mech. 2006, 3, 343−350. (16) Fertleman, C. R.; Baker, M. D.; Parker, K. A.; Moffatt, S.; Elmslie, F. V.; Abrahamsen, B.; Ostman, J.; Klugbauer, N.; Wood, J. N.; Gardiner, R. M.; Rees, M. SCN9A mutations in paroxysmal extreme pain disorder: allelic variants underlie distinct channel defects and phenotypes. Neuron 2006, 52, 767−774. (17) Faber, C. G.; Hoeijmakers, J. G.; Ahn, H. S.; Cheng, X.; Han, C.; Choi, J. S.; Estacion, M.; Lauria, G.; Vanhoutte, E. K.; Gerrits, M. M.; Dib-Hajj, S.; Drenth, J. P.; Waxman, S. G.; Merkies, I. S. Gain of function Naν1.7 mutations in idiopathic small fiber neuropathy. Ann. Neurol. 2012, 71, 26−39. (18) Amin, A. S.; Asghari-Roodsari, A.; Tan, H. L. Cardiac sodium channelopathies. Pflugers Arch. 2010, 460, 223−237. (19) (a) Nardi, A.; Damann, N.; Hertrampf, T.; Kless, A. Advances in targeting voltage-gated sodium channels with small molecules. ChemMedChem 2012, 7, 1712−1740. (b) Bagal, S. K.; Chapman, M. L.; Marron, B. E.; Prime, R.; Storer, R. I.; Swain, N. A. Recent progress in sodium channel modulators for pain. Bioorg. Med. Chem. Lett. 2014, 24, 3690−3699. (c) Sun, S.; Cohen, C. J.; Dehnhardt, C. M. Inhibitors of voltage-gated sodium channel NaV1.7: patent applications since 2010. Pharm. Pat. Anal. 2014, 3, 509−521. (d) de Lera Ruiz, M.; Kraus, R. L. Voltage-gated sodium channels: Structure, function, pharmacology, and clinical indications. J. Med. Chem. 2015, 58, 7093−7118. (20) (a) Jones, H. M.; Butt, R. P.; Webster, R. W.; Gurrell, I.; Dzygiel, P.; Flanagan, N.; Fraier, D.; Hay, T.; Iavarone, L. E.; Luckwell, J.; Pearce, H.; Phipps, A.; Segelbacher, J.; Speed, B.; Beaumont, K. Clinical micro-dose studies to explore the human pharmacokinetics of four selective inhibitors of human NaV1.7 voltage-dependent sodium channels. Clin. Pharmacokinet. 2016, 55, 875−887. (b) Swain, N. A.; Batchelor, D.; Beaudoin, S.; Bechle, B. M.; Bradley, P. A.; Brown, A. D.; Brown, B.; Butcher, K. J.; Butt, R. P.; Chapman, M. L.; Denton, S.; Ellis, D.; Galan, S. R. G.; Gaulier, S. M.; Greener, B. S.; de Groot, M. J.; Glossop, M. S.; Gurrell, I. K.; Hannam, J.; Johnson, M. S.; Lin, Z.; Markworth, C. J.; Marron, B. E.; Millan, D. S.; Nakagawa, S.; Pike, A.; Printzenhoff, D.; Rawson, D. J.; Ransley, S. J.; Reister, S. M.; Sasaki, K.; Storer, R. I.; Stupple, P. A.; West, C. W. Discovery of clinical candidate 4-[2-(5-amino-1Hpyrazol-4-yl)-4-chlorophenoxy]-5-chloro-2-fluoro-N-1,3-thiazol-4-ylbenzenesulfonamide (PF-05089771): design and optimization of diary ether aryl sulfonamides as selective inhibitors of NaV1.7. J. Med. Chem. 2017, 60, 7029−7042. (c) Storer, R. I.; Pike, A.; Swain, N. A.; Alexandrou, A. J.; Bechle, B. M.; Blakemore, D. C.; Brown, A. D.; Castle, N. A.; Corbett, M. S.; Flanagan, N. J.; Fengas, D.; Johnson, M. S.; Jones, L. H.; Marron, B. E.; Payne, C. E.; Printzenhoff, D.; Rawson, D. J.; Rose, C. R.; Ryckmans, T.; Sun, J.; Theile, J. W.; Torella, R.; Tseng, E.; Warmus, J. S. Highly potent and selective NaV1.7 inhibitors 925



Article

Gunaydin, H.; Guzman-Perez, A.; Fremeau, R., Jr.; Huang, X.; Ilch, C.; Kornecook, T. J.; Kreiman, C.; Ligutti, J.; Lin, M. J.; McDermott, J. S.; Marx, I.; Matson, D. J.; McDonough, S. I.; Moyer, B. D.; Nguyen, H. N.; Taborn, K.; Yu, V.; Weiss, M. M. The discovery of benzoxazine sulfonamide inhibitors of NaV1.7: Tools that bridge efficacy and target engagement. Bioorg. Med. Chem. Lett. 2017, 27, 3477−3485. (23) (a) Wu, Y.; Guernon, J.; Shi, J.; Ditta, J.; Robbins, K. J.; Rajamani, R.; Easton, A.; Newton, A.; Bourin, C.; Mosure, K.; Soars, M. G.; Knox, R. J.; Matchett, M.; Pieschl, R. L.; Post-Munson, D. J.; Wang, S.; Herrington, J.; Graef, J.; Newberry, K.; Bristow, L. J.; Meanwell, N. A.; Olson, R.; Thompson, L. A.; Dzierba, C. Development of new benzenesulfonamides as potent and selective NaV1.7 inhibitors for the treatment of pain. J. Med. Chem. 2017, 60, 2513−2525. (b) Wu, Y.; Guernon, J.; McClure, A.; Luo, G.; Rajamani, R.; Ng, A.; Easton, A.; Newton, A.; Bourin, C.; Parker, D.; Mosure, K.; Barnaby, O.; Soars, M. G.; Knox, R. J.; Matchett, M.; Pieschl, R.; Herrington, J.; Chen, P.; Sivarao, D. V.; Bristow, L. J.; Meanwell, N. A.; Bronson, J.; Olson, R.; Thompson, L. A.; Dzierba, C. Discovery of non-zwitterionic aryl sulfonamides as NaV1.7 inhibitors with efficacy in preclinical behavioral models and translational measures of nociceptive neuron activation. Bioorg. Med. Chem. 2017, 25, 5490−5505. (24) (a) Roecker, A. J.; Egbertson, M.; Jones, K. L. G.; Gomez, R.; Kraus, R. L.; Li, Y.; Koser, A. J.; Urban, M. O.; Klein, R.; Clements, M.; Panigel, J.; Daley, C.; Wang, J.; Finger, E. N.; Majercak, J.; Santarelli, V.; Gregan, I.; Cato, M.; Filzen, T.; Jovanovska, A.; Wang, Y.; Wang, D.; Joyce, L. A.; Sherer, E. C.; Peng, X.; Wang, X.; Sun, H.; Coleman, P. J.; Houghton, A. K.; Layton, M. E. Discovery of selective, orally bioavailable, N-linked arylsulfonamide NaV1.7 inhibitors with pain efficacy in mice. Bioorg. Med. Chem. Lett. 2017, 27, 2087−2093. (b) Pero, J. E.; Rossi, M. A.; Lehman, H. D. G. F.; Kelly, M. J., III; Mulhearn, J. J.; Wolkenberg, S. E.; Cato, M. J.; Clements, M. K.; Daley, C. J.; Filzen, T.; Finger, E. N.; Gregan, Y.; Henze, D. A.; Jovanovska, A.; Klein, R.; Kraus, R. L.; Li, Y.; Liang, A.; Majercak, J. M.; Panigel, J.; Urban, M. O.; Wang, J.; Wang, Y.; Houghton, A. K.; Layton, M. E. Benzoxazolinone aryl sulfonamides as potent, selective NaV1.7 inhibitors with in vivo efficacy in a preclinical pain model. Bioorg. Med. Chem. Lett. 2017, 27, 2683−2688. (25) (a) Wu, W.; Li, Z.; Yang, G.; Teng, M.; Qin, J.; Hu, Z.; Hou, L.; Shen, L.; Dong, H.; Zhang, Y.; Li, J.; Chen, S.; Tian, J.; Zhang, J.; Ye, L. The discovery of tetrahydropyridine analogs as hNaV1.7 selective inhibitors for analgesia. Bioorg. Med. Chem. Lett. 2017, 27, 2210− 2215. (b) Teng, M.; Wu, W.; Li, Z.; Yang, G.; Qin, J.; Wang, Y.; Hu, Z.; Dong, H.; Hou, L.; Hu, G.; Shen, L.; Zhang, Y.; Li, J.; Chen, S.; Tian, J.; Ye, L.; Zhang, J.; Wang, H. Discovery of aminocyclohexene analogues as selective and orally bioavailable hNaV1.7 inhibitors for analgesia. Bioorg. Med. Chem. Lett. 2017, 4979−4984. (26) Ahuja, S.; Mukundl, S.; Deng, L.; Khakh, K.; Chang, E.; Ho, H.; Shriverl, S.; Young, C.; Lin, S.; Johnson, J. P., Jr.; Wu, P.; Li, J.; Coons, M.; Tam, C.; Brillantes, B.; Sampang, H.; Mortara, K.; Bowman, K. K.; Clark, K. R.; Estevez, A.; Xie, Z.; Verschoof, H.; Grimwood, M.; Dehnhardt, C.; Andrez, J.-C.; Focken, T.; Sutherlin, D. P.; Safina, B. S.; Starovasnik, M. A.; Ortwine, D. F.; Franke, Y.; Cohen, C. J.; Hackos, D. H.; Koth, C. M.; Payandeh, J. Structural basis of NaV1.7 inhibition by an isoform-selective small molecule antagonist. Science 2015, 350, No. aac5464. (27) (a) Beaudoin, S.; Laufer-Sweiler, M. C.; Markworth, C. J.; Marron, B. E.; Millan, D. S.; Rawson, D. J.; Reister, S. M.; Sasaki, K.; Storer, R. I.; Stupple, P. A.; Swain, N. A.; West, C. W.; Zhou, S. Sulfonamide Derivatives. PCT International Application WO20100794432010. (b) Markworth, C. J.; Marron, B. E.; Rawson, D. J.; Storer, R. I.; Swain, N. A.; West, C. W.; Zhou, S. Chemical Compounds. PCT International Application WO20120047062012. (c) Greener, B.; Marron, B. E.; Millan, D. S.; Rawson, D. J.; Storer, R. I.; Swain, N. A. Chemical Compounds. PCT International Application WO20120047142012. (d) Brown, A. D.; de Groot, M. J.; Marron, B. E.; Rawson, D. J.; Ryckmans, T.; Storer, R. I.; Stupple, P. A.; Swain, N. A.; West, C. W. Benzenesulfonamides Useful

As Sodium Channel Inhibitors. PCT International Application WO20120047432012. (e) Bell, A. S.; Brown, A. D.; de Groot, M. J.; Gaulier, S. M.; Lewthwaite, R. A.; Marsh, I. R.; Millan, D. S.; Perez, P. M.; Rawson, D. J.; Sciammetta, N.; Storer, R. I.; Swain, N. A. NSulfonamide Derivatives Useful as Voltage Gated Sodium Channel Inhibitors. PCT International Application WO20120078612012. (f) Brown, A. D.; Rawson, D. J.; Storer, R. I.; Swain, N. A. Chemical Compounds. PCT International Application WO20120078682012. (g) Brown, A. D.; Rawson, D. J.; Storer, R. I.; Swain, N. A. Chemical Compounds. PCT International Application WO20120078692012. (h) Bell, A. S.; Brown, A. D.; Lewthwaite, R. A.; Perez, P. M.; Rawson, D. J.; Storer, R. I.; Swain, N. A. Chemical Compounds. PCT International Application WO20120078772012. (i) Bell, A. S.; Brown, A. D.; de Groot, M. J.; Lewthwaite, R. A.; Marsh, I. R.; Millan, D. S.; Perez, P. M.; Rawson, D. J.; Sciammetta, N.; Storer, R. I.; Stupple, P. A.; Swain, N. A. Sulfonamide Derivatives as NaV1.7 Inhibitors for the Treatment of Pain. PCT International Application WO20120078832012. (j) Bell, A. S.; de Groot, M. J.; Lewthwaite, R. A.; Marsh, I. R.; Sciammetta, N.; Storer, R. I.; Swain, N. A. Indazole Derivatives as Sodium Channel Inhibitors. PCT International Application WO20120957812012. (k) Rawson, D. J.; Storer, R. I.; Swain, N. A. Sulfonamide Derivatives. PCT International Application WO20130883152013. (l) Storer, R. I.; Swain, N. A. Sulfonamide Derivatives and use Thereof as VGSC Inhibitors. WO20130936882013. (m) Brown, A. D.; Galan, S. R. G.; Millan, D. S.; Rawson, D. J.; Storer, R. I.; Stupple, P. A.; Swain, N. A. NAminosulfonyl Benzamides. PCT International Application WO20131028262013. (28) Both series displayed the same average LLE of 2.5. (29) (a) Lovering, F.; Bikker, J.; Humblet, C. Escape from flatland: increasing saturation as an approach to improving clinical success. J. Med. Chem. 2009, 52, 6752−6756. (b) Ritchie, T. J.; Macdonald, S. J. F. The impact of aromatic ring count on compound developability− are too many aromatic rings a liability in drug design? Drug Discovery Today 2009, 14, 1011. (30) Yan, A.; Gasteiger, J. Prediction of aqueous solubility of organic compounds by topological descriptors. QSAR Comb. Sci. 2003, 22, 821−829. (31) The holding potential was chosen to produce optimal potent block. Drug effects at physiological relevant voltages would be half as potent. (32) Andrez, J.-C; Chowdhury, S.; Decker, S.; Dehnhardt, C. M.; Focken, T.; Grimwood, M. E.; Hemeon, I. W.; Jia, Q.; Li, J.; Ortwine, D. F.; Safina, B. S.; Sheng, T.; Sun, S.; Sutherlin, D. P.; Wilson, M. S.; Zenova, A. Y. N-Substituted Benzamides and Their Use in the Treatment of Pain. PCT International Application WO2013177224A12013. (33) Johnson, T. W.; Dress, K. R.; Edwards, M. Using golden triangle to optimize clearance and oral absorption. Bioorg. Med. Chem. Lett. 2009, 19, 5560−5564. (34) (a) Edwards, M. P.; Price, D. A. Role of physicochemical properties and ligand lipophilicity efficiency in addressing drug safety risks. Annu. Rep. Med. Chem. 2010, 45, 380−391. (b) Leeson, P. D.; Springthorpe, B. The influence of drug-like concepts on decisionmaking in medicinal chemistry. Nat. Rev. Drug Discovery 2007, 6, 881−890. (35) Obach, R. S.; Baxter, J. G.; Liston, T. E.; Silber, B. M.; Jones, B. C.; MacIntyre, F.; Rance, D. J.; Wastall, P. The prediction of human pharmacokinetic parameters from preclinical and in vitro metabolism data. J. Pharmacol. Exp. Ther. 1997, 283, 46−58. (36) (a) Barreiro, E. J.; Kümmerle, A. E.; Fraga, C. A. M. The methylation effect in medicinal chemistry. Chem. Rev. 2011, 111, 5215−5246. (b) Schönherr, H.; Cernak, T. Profound methyl effects in drug discovery and a call for new C−H methylation reactions. Angew. Chem., Int. Ed. 2013, 52, 12256−12267. (37) Bankar, G.; Howard, S.; Nelkenbrecher, K.; Waldbrook, M.; Dourado, M.; Goodchild, S. J.; Shuart, N.; Lin, S.; Young, C.; Xie, Z.; Khakh, K.; Chang, E.; Sojo, L. E.; Lindgren, A.; Chowdhury, S.; Decker, S.; Grimwood, M.; Andrez, J. C.; Dehnhardt, C. M.; Pang, J.; 926



Article

receptor induced fit effects. J. Med. Chem. 2006, 49, 534−553. (c) Sherman, W.; Beard, H. S.; Farid, R. Use of an induced fit receptor structure in virtual screening. Chem. Biol. Drug Des. 2006, 67, 83−84.

Chang, J. H.; Safina, B. S.; Sutherlin, D. P.; Johnson, J. P.; Hackos, D. H.; Robinette, C. L.; Cohen, C. J. Selective NaV1.7 antagonists with long residence time show improved efficacy against inflammatory and neuropathic pain. Cell Rep. 2018, 24, 3133−3145. (38) Osoegawa, K.; Woon, P. Y.; Zhao, B.; Frengen, E.; Tateno, M.; Catanese, J. J.; de Jong, P. An improved approach for construction of bacterial artificial chromosome libraries. Genomics 1998, 52, 1−8. (39) (a) Dubuisson, D.; Dennis, S. G. The formalin test: a quantitative study of the analgesic effects of morphine, meperidine, and brain stem stimulation in rats and cats. Pain 1977, 4, 161−177. (b) Yaksh, T. L.; Ozaki, G.; McCumber, D.; Rathbun, M.; Svensson, C.; Malkmus, S.; Yaksh, M. C. An automated flinch detecting system for use in the formalin nociceptive. bioassay. J. Appl. Physiol. 2001, 90, 2386−2402. (c) Abbadie, C.; Taylor, B. K.; Peterson, M. A.; Basbaum, A. I. differential contribution of the two phases of the formalin test to the pattern of c-fos expression in the rat spinal cord: studies with remifentanil and lidocain. Pain 1997, 69, 101−110. (40) (a) Priest, B. T.; Garcia, M. L.; Middletion, R. E.; Brochu, R. M.; Clark, S.; Dai, G.; Dick, I. E.; Felix, J. P.; Liu, C. J.; Reiseter, B.; Schmalhofer, W. A.; Shao, P.; Tang, Y. S.; Chou, M. Z.; Kohler, M. G.; Smith, M. M.; Warren, V. A.; Williams, B. S.; Cohen, C. J.; Martin, W. J.; Meinke, P. T.; Parsons, W. H.; Wafford, K. A.; Kaczorowski, G. J. A disubstituted succinamide is a Potent sodium channel blocker with efficacy in a rat pain model. Biochemistry 2004, 43, 9866−9876. (b) Bregman, H.; Berry, L.; Buchanan, J. L.; Chen, A.; Bingfan Du, B.; Feric, E.; Hierl, M.; Huang, L.; Immke, D.; Janosky, B.; Johnson, D.; Li, X.; Ligutti, J.; Liu, D.; Malmberg, A.; Matson, D.; McDermott, J.; Miu, P.; Nguyen, H. N.; Patel, V. F.; Waldon, D.; Wilenkin, B.; Zheng, X.; Zou, A.; McDonough, S. I.; DiMauro, E. F. Identification of potent, state-dependent inhibitors of NaV1.7 with oral efficacy in the formalin model of persistent pain. J. Med. Chem. 2011, 54, 4427− 4445. (c) Macsari, I.; Besidski, Y.; Csjernyik, G.; Nilsson, L. I.; Sandberg, L.; Yngve, U.; Åhlin, K.; Bueters, T.; Eriksson, A. B.; Lund, P.-E.; Venyike, E.; Oerther, S.; Blakeman, K. H.; Luo, L.; Arvidsson, P. I. 3-Oxoisoindoline-1-carboxamides: Potent, state-dependent blockers of voltage-gated sodium channel NaV1.7 with efficacy in rat pain models. J. Med. Chem. 2012, 55, 6866−6880. (d) Kers, I.; Csjernyik, G.; Macsari, I.; Nylöf, M.; Sandberg, L.; Skogholm, K.; Bueters, T.; Eriksson, A. B.; Oerther, S.; Lund, P.; Venyike, E.; Nyström, J.; Besidski, Y. Structure and activity relationship in the (S)-N-chroman3-ylcarboxamide series of voltage-gated sodium channel blockers. Bioorg. Med. Chem. Lett. 2012, 22, 5618−5624. (41) McNamara, C. R.; Mandel-Brehm, J.; Bautista, D. M.; Siemens, J.; Deranian, K. L.; Zhao, M. TRPA1 mediates formalin-induced pain. Proc. Natl. Acad. Sci. U.S.A. 2007, 104, 13525−13530. (42) Tjølsen, A.; Berge, O. G.; Hunskaar, S.; Rosland, J. H.; Hole, K. The formalin test: an evaluation of the method. Pain 1992, 51, 5−17. (43) Wu, X.; Ronn, R.; Gossas, T.; Larhed, M. Easy-to-execute carbonylations: microwave synthesis of acyl sulfonamides using Mo(CO)6 as a solid carbon monoxide source. J. Org. Chem. 2005, 70, 3094−3098. (44) Miyaura, N.; Suzuki, A. palladium-catalyzed cross-coupling reactions of organoboron compounds. Chem. Rev. 1995, 95, 2457− 2483. (45) Denmark, S. E.; Edwards, J. P. A comparison of (chloromethyl)- and (iodomethyl)zinc cyclopropanation reagents. J. Org. Chem. 1991, 56, 6974−6981. (46) (a) Sastry, G. M.; Adzhigirey, M.; Day, T.; Annabhimoju, R.; Sherman, W. Protein and ligand preparation: Parameters, protocols, and influence on virtual screening enrichments. J. Comput.-Aided Mol. Des. 2013, 27, 221−234. (b) Schrö dinger Release 2014-1: Schrö dinger Suite 2017-4. Protein Preparation Wizard; Epik, Schrödinger, LLC, New York, NY, 2016; Impact, Schrödinger, LLC, New York, NY, 2016; Prime, Schrödinger, LLC, New York, NY, 2017. (47) (a) Farid, R.; Day, T.; Friesner, R. A.; Pearlstein, R. A. New insights about HERG blockade obtained from protein modeling, potential energy mapping, and docking studies. Bioorg. Med. Chem. 2006, 14, 3160−3173. (b) Sherman, W.; Day, T.; Jacobson, M. P.; Friesner, R. A.; Farid, R. Novel procedure for modeling ligand/ 927


Identification of Selective Acyl Sulfonamide ... - ACS Publications

Recommend Documents