Tolvaptan-Type Vasopressin Receptor Ligands - ACS Publications

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Tolvaptan-Type Vasopressin Receptor Ligands: Important Role of Axial Chirality in the Active Form Hidetsugu Tabata, Tetsuya Yoneda, Tetsuta Oshitari, Hideyo Takahashi, and Hideaki Natsugari* Faculty of Pharma Sciences, Teikyo University, 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605, Japan S Supporting Information *

ABSTRACT: The anti and syn isomers of tolvaptan-type compounds, N-benzoyl-5-hydroxy-1-benzazepines (5a−c), were prepared in a stereocontrolled manner by biasing the conformation with a methyl group at C9 and C6, respectively, and the enantiomeric forms were separated. Examination of the affinity at the human vasopressin receptors revealed that the axial chirality (aS) plays a more important role than the central chirality at C5 in receptor recognition, and the most preferable form was shown to be (E,aS,5S).

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INTRODUCTION The nonpeptide arginine vasopressin (VP) V2 receptor antagonists1 have been developed as the “vaptan” class of ligands for the treatment of hyponatremia, congestive heart failure, and other diseases associated with water homeostasis disturbance. Among the VP antagonists, tolvaptan2 and mozavaptan3 (Figure 1) are representative: both contain a

Figure 2. Stereoisomers [E/Z-amide rotamers and (aS)/(aR)-axial isomers] in 1−3. The Z-amide isomers shown in the brackets were negligible in the 1H NMR spectrum.

amide isomers, compounds 1−3 all exist predominantly in the E-form, and the Z-isomer was negligible in the 1H NMR spectrum.10 Thus, 1−3 exist only as a mixture of (aS)/(aR)axial isomers (Figure 2). The molecules with a substituent at the ortho-position (C9)11 on the benzene ring (e.g., CH3) were conformationally “frozen” and could be separated into the (aS)- and (aR)-axial isomers using chiral HPLC, although the conformational change in the molecules without the substituent at C9 was too rapid for isolation of the isomers (i.e., achiral form) at room temperature (rt). Those studies led to the finding of the importance of axial chirality in exerting the biological activity of the vaptan class of VP receptor ligands, exhibiting the active structure (eutomer) with the (E,aS)-form of the types 1 and 2 (Figure 2).5,6

Figure 1. Nonpeptide VP receptor antagonists.

preserved scaffold, i.e., 1-benzazepine linked through N-1 to a substituted benzoyl group (blue lines in the structures in Figure 1), and the characteristic feature is a substituent (OH and NMe2) added to the 1-benzazepine nucleus at C5. Interest in VP antagonists is still growing in the search for new ligands, their new indications, and in the biological role of the V1a and V1b subtype receptors.1 A topic of recent interest was the approval of tolvaptan as the first drug for autosomal-dominant polycystic kidney disease (ADPKD) in Japan.4 In our previous papers,5,6 atropisomerism in N-benzoyl-1,5benzodiazepines (1, X = NCH 3 ) 5 and N-benzoyl-1,5benzothiazepines (2, X = S; 3, X = S*=O)6 as VP receptor ligands was described (Figure 2), in which the E/Z-amide rotamers7 around the N−C(=O) bond and (aS)/(aR)-axial isomers8 based on the Ar−N(CO) (sp2−sp2) axis were considered9 (Figure 2). In general, however, for the E/Z© 2017 American Chemical Society

Received: March 17, 2017 Published: May 5, 2017 4503

DOI: 10.1021/acs.jmedchem.7b00422 J. Med. Chem. 2017, 60, 4503−4509

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prepared using conventional methods starting from 5-oxo-1benzazepine derivatives (6a−c), as shown in Scheme 1. First,

Whereas 1 and 2 possess no central chirality, tolvaptan and mozavaptan (Figure 1) possess central chirality at C5 bearing a substituent and are developed as racemates. Thus, in these two vaptans, four stereoisomers originating in two stereogenic elements at C5 and the axis are presumed to exist, i.e., (aR,5S)/ (aS,5R) and (aS,5S)/(aR,5R) (all adopt the E-amide conformation) (Figure 3). The former and latter pairs of

Scheme 1. Preparation of Tolvaptan-Type N-Benzoyl-5hydroxy-1-benzazepines (5a−c) and Separation into the Enantiomersa

a

Only the major anti (aR*,5S*) isomer of 5c was isolated in the pure form from the anti/syn mixture. Figure 3. Anti/syn diastereomers and the enantiomers in (E)-N-4-[(2methylbenzoyl)amino]benzoyl-1,5-benzothiazepine S-oxides (3) and (E)-5-hydroxy-N-(p-toluoyl)-1-benzazepines (4).

N-benzoylation of 6a−c using p-nitrobenzoyl chloride was followed by catalytic reduction using Pd/C to give the paminobenzoyl compounds 7a−c. Then, amidation of 7a−c with o-toluoyl chloride followed by reduction of the carbonyl group at C5 with NaBH4 furnished the racemic N-benzoyl-5-hydroxy1-benzazepine derivatives (5a−c).16 As for the E/Z-amide isomers,7 5a−c all exist predominantly in the E-form, and the Z-isomer was negligible in the 1H NMR spectrum. Thus, in these compounds, the anti (aR*,5S*) and syn (aS*,5S*) diastereomers with the E-amide form are presumed to exist. In 5a (R1 = R2 = H), the ring inversion via rotation around the Ar−N(C=O) axis readily occurs to form anti/syn diastereomers in solution at rt [1:0.31 as determined by the 1 H NMR spectrum in DMSO-d6)],17 which are inseparable by HPLC. Meanwhile, in 5c (R1 = H, R2 = CH3), the rotation around the axis is restricted at rt by the ortho substituent (C9CH3) to form separable anti/syn diastereomers with a ratio of 1:0.17, from which the major anti (aR*,5S*) isomer could be isolated in the pure form with HPLC using a nonchiral column. Attempted isolation of the minor syn (aS*,5S*) isomer in the pure form failed because the retention times of the anti/syn isomers are too close for separation (isolation) of the minor syn isomer. The anti/syn stereochemistry of 5a and 5c was confirmed by comparisons with the 1H NMR spectral data on the corresponding N-(p-toluoyl) derivatives of 5-hydroxy-1benzazepines (4) previously reported.14 On the other hand, in 5b (R1 = CH3, R2 = H), because of the steric hindrance caused by the C6 methyl group (allylic 1,3strain), the C5 hydroxy group is confined so as to adopt a sterically less hindered axial (ax) orientation, forming the syn (aS*,5S*) isomer exclusively (Figure 4). The syn stereochemistry of 5b was confirmed by comparisons with the analytical data on the 5-hydroxy-6-methyl-N-(p-toluoyl)-1benzazepine (4) previously reported.14 The racemic N-benzoyl-5-hydroxy-1-benzazepine derivatives (5a−c) thus obtained were successfully separated and isolated

stereoisomers correspond to the anti and syn diastereomers, respectively: the description “anti/syn” is used for the relative arrangement of the C5 substituent and the N-benzoyl group, i.e., anti and syn denote the arrangement on the opposite and the same side, respectively. To the best of our knowledge, however, the active stereochemistry of the vaptans (i.e., tolvaptan, mozavaptan) including the central chirality at C5 has not been discussed thoroughly to date.12,13 In our previous paper, 6 we prepared (E)-N-4-[(2methylbenzoyl)amino]benzoyl-1,5-benzothiazepine S-oxides (3), examined the in vitro affinities at the VP receptors, and proposed that the most active stereochemistry of 3 might be in the (aS,5S)-form (3b) (Figure 3). However, the conclusion was not definite because the affinities (Ki values) were weak (at micromolar level). On the other hand, recently we have revisited the basic stereochemistry of N-benzoyl-1-benzazepines with a substituent (CH3, OH, NMe2) at C5 (e.g., 4 in Figure 3),14 for which the stereochemistry in the solution state was previously ambiguously reported,15 to reveal new physicochemical aspects of these heterocycles including revision of the reported15 stereochemistry. In this paper, using the stereochemical information obtained in 4, we synthesized the anti and syn diastereomers of the tolvaptan-type compounds (5hydroxy-N-benzoyl-1-benzazepine derivatives) (5a−c) (Scheme 1) in stereocontrolled manners, and examination of the affinities at the human VP receptors for the enantiomers revealed that axial chirality present in the latent form in the molecule plays a more important role than the central chirality at C5 in receptor recognition. The results are presented here.

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CHEMISTRY Preparation of N-Benzoyl-5-hydroxy-1-benzazepine Derivatives (5a−c) and Separation to the Enantiomers. N-Benzoyl-5-hydroxy-1-benzazepine derivatives (5a−c) were 4504

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Figure 4. Syn (aS*,5S*) form of N-benzoyl-5-hydroxy-6-methyl-1benzazepine (5b): because of the steric hindrance (A1,3-strain), the 5OH group adopts a sterically less hindered ax orientation to form the syn isomer.

into the enantiomers with HPLC using a chiral column15 (Scheme 1). The chiroptical properties [optical rotation ([α]D) and circular dichroism (CD)] of the separated enantiomers were examined and the data are shown in Figure 5. The Figure 6. X-ray crystal structures of the optically active compounds (+)-(E,aS,5R)-5a (= anti) and (+)-(E,aS,5S)-5b (= syn).

thermodynamically unstable, with the 5-hydroxy group in the ax position in a chairlike form. As for the absolute stereochemistry of 5c (major diastereomer), although the Xray data could not be obtained, (+)-5c was deduced to be (E,aS,5R) by comparison with the chiroptical properties (CD and [α]D) of (+)-5a and (+)-5b.19

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RESULTS AND DISCUSSION Affinity at the V1a and V2 Receptors (Table 1). The in vitro affinities at the human VP V1a and V2 receptors of 5a−c are shown in Table 1. First, 5a−c in the racemic forms were evaluated. All compounds showed good potency in the binding experiment with the affinity (Ki) at the nanomolar to submicromolar level. Interestingly, 5b and 5c, which bear a methyl group in the benzene ring, have a tendency to bind Table 1. In Vitro Affinity for Human VP V1a and V2 Receptors of 5a−c

Figure 5. Chiroptical properties, [α]D20 (in CHCl3) and CD (in CH3OH) spectra of the separated enantiomers of 5a−c.

Ki (nM)a,b

chirality

enantiomers of 5a−c with the (+)-angle of [α]D exhibited similar CD spectra (blue lines) with positive bands around 250−320 nm, and those with the (−)-angle of [α]D exhibited similar CD spectra (red lines) with negative bands around 250−320 nm. Fortunately, the enantiomers (+)-5a and (+)-5b could be subjected to X-ray structural analysis,18 which revealed that the absolute stereochemistry of (+)-5a is (E,aS,5R) and that of (+)-5b is (E,aS,5S) (Figure 6). The X-ray structural analysis of (+)-5a indicated that, although (+)-5a exists as an inseparable equilibrium state of a mixture of anti and syn (1:0.31) in solution (NMR) as described above, in the solid state it takes the thermodynamically stable anti form with the 5-hydroxy group in the equatorial position in a chairlike form. In contrast, (+)-5b was shown to exist in both solid and solution state solely as the syn form, which is apparently

compd

[α]D rotation

C5

axis

hV1ac

hV2c

5a

(±) (−) (+) (±) (+) (−) (±) (−) (+)

S* S R S* S R S* S R

aR*/aS* (1:0.31) aR/aS (1:0.31) aS/aR (1:0.31) aS* aS aR aR* aR aS

1.58 1.85 1.03 6.75 2.06 78 6.26 4.68 2.84

7.56 4.87 8.30 160 46 1790 170 54 140

5b

5cd

Inhibition assays were performed for 2 h at 25 °C in 1% DMSO. bKi values shown are the means of duplicate measurements. cFor details, see the Experimental Section. dMajor anti [(aR,5S)/(aS,5R)] diastereomer. The minor syn [(aS,5S)/(aR,5R)] isomers could not be isolated because of the separation difficulty from the major anti isomer. a

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selectively to V1a (at the nanomolar level) rather than to V2 receptors (at the submicromolar level). The excellent V1aselective affinity may make them worthy candidates for further biological studies. Next, the separated enantiomers of 5a−c were subjected to the binding assay to examine the difference in potency between the enantiomers. However, contrary to our expectation, in 5a (R1 = R2 = H) and 5c (R1 = H, R2 = CH3), the enantiomers and the racemate exhibited similar levels of affinity (within a 3-fold difference) (Table 1). In contrast, 5b (R1 = CH3, R2 = H) showed a ca. 40-fold difference in potency between the enantiomers [active eutomer: (+)-(aS,5S) (syn)-5b and less active distomer: (−)-(aR,5R) (syn)-5b)] with median potency in the racemate (Table 1). Active Molecular Forms Recognized by the VP Receptors. The clear difference in potency between the enantiomers of 5b (a ca. 40-fold difference) indicates that the stereochemistry at the scaffold region, i.e., (E,aS,5S), is recognized by VP receptors. That the (aS)-form is important in exerting the activity is in accordance with the VP ligands of N-benzoyl-1,5-benzodiazepine (1) and N-benzoyl-1,5-benzothiazepine (2) (Figure 2) we previously reported.5,6 As for the stereochemistry at C5 of 5b, (5S) was shown to be the preferable form for binding with receptors, and it may be presumed that the (E,aS,5S)-form is the preferable and the (E,aR,5R)-form the least preferable among the four stereoisomers present in 5a−c. However, the results obtained in 5a and 5c, in which the enantiomers and the racemate exhibited similar levels of affinity, led to confusion in the analysis of the active stereochemistry. We considered the following to explain the results. In 5a, the ring inversion via rotation around the axis readily occurs at rt in the solution state to form inseparable anti/syn diastereomers in a ratio of 1:0.31 [i.e., (aR,5S)/(aS,5S) for (5S)-5a and (aS,5R)/(aR,5R) for (5R)-5a, each in a ratio of 1:0.31], meaning that the (aS)-form, which we presume is the active form, is present in the binding experiment in both the (5S)- and (5R)-forms of 5a. The observed similar level of affinity between (5S)-5a and (5R)-5a may imply that, although the (aS,5S)-form exists in (5S)-5a as the minor diastereomer in solution, it should contribute to exhibit the similar level of affinity as that of (5R)-5a because it possesses the strongest affinity among these four isomers. On the other hand, in the major anti (aS*,5R*)-5c, while the affinity of the enantiomer (aS,5R)-5c was envisioned to be reasonably estimated, the affinity of the other enantiomer (aR,5S)-5c was unexplainable because it does not take the (aS)form, which we presume is the active form. Thus, we speculated that the affinity observed in (aR,5S)-5c is ascribed to the minor syn diastereomer (aS,5S)-5c, which would be formed from the (aR,5S)-5c during the binding experiment (for 2 h at 25 °C). To confirm the conversion, the energy barrier between the two diastereomers (aR,5S) and (aS,5S) of 5c was examined. The time-dependent conversion rate [from (aR,5S)-5c to (aS,5S)5c] was estimated from chiral HPLC analysis of a solution of the enantiomer in toluene after allowing it to stand at 25 °C. The conversion reached an equilibrium state of (aR,5S)/ (aS,5S) = 1:0.17 after 4 h, and the ΔG⧧ value20 was determined to be 102 kJ/mol (Figure 7). The results indicate that the conversion from the (aR,5S)-form to (aS,5S)-form actually occurred during the binding experiment; if the amount of (aS,5S)-5c (the most potent isomer) formed from (aR,5S)-5c is 10−15%, the observed Ki value of (aR,5S)-5c is possible to be

Figure 7. Conversion of (aR,5S) (= anti)-5c to (aS,5S) (= syn)-5c. For the conversion profile, see Figure S1 in the Supporting Information.

at the similar level as that of the enantiomer (aS,5R)-5c.21 Meanwhile, as for the enantiomer (aS,5R)-5c, the conversion to (aR,5R)-form would give little effect on the observed Ki value because the affinity of the (aR,5R)-form may be too weak. All the above results support the assumption that the (E,aS)stereochemistry of the tolvaptan-type of N-benzoyl-5-hydroxy1-benzazepine derivatives (5a−c) is the preferable form and the (E,aS,5S)-form is the most preferable among the four stereoisomers for recognition by VP receptors (Figure 8): the

Figure 8. Four stereoisomers of the tolvaptan-type VP ligands (tolvaptan and 5a) with a C5-hydroxy group. For receptor recognition, the (E,aS,5R)-stereochemistry is preferable and the (E,aS,5S)-form is the most preferable. For the exact structure of tolvaptan, see Figure 1.

two X-ray crystal structures shown in Figure 6 are the desirable forms with the (aS)-stereochemistry. Thus, the axial chirality present in the latent form in the molecule plays a more important role than the central chirality at C5 in receptor recognition.

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CONCLUSION The anti and syn isomers of the tolvaptan-type compounds Nbenzoyl-5-hydroxy-1-benzazepines (5a−c) were successfully prepared in a stereocontrolled manner by biasing the conformation with a methyl group at C9 and C6, respectively, and the enantiomeric forms were separated. Examination of the affinity at the VP receptors revealed that the axial chirality (aS) plays a more important role than the central chirality at C5 in receptor recognition, and the (E,aS,5S)-form is the most preferable. Sometimes, and somewhat peculiarly, biologically active compounds bearing central chirality in their molecules exhibit similar levels of activity regardless of the chirality caused by the chiral center. In such cases, the activity may be due to the latent chirality caused by dynamic conformational changes at different sites of molecules, as was observed in 5a and 5c in this investigation. This paper suggests the importance of the detailed conformational analysis of the entire molecule, and the 4506

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Single-Crystal X-ray Analysis. The crystal structures of the optically active compounds [(+)-(aS,5R)-5a (= anti) and (+)-(aS,5S)5b (= syn)] were obtained by single-crystal X-ray analysis (Figure 6). Typical crystal data for (+)-(aS,5R)-5a and (+)-(aS,5S)-5b are shown in the Supporting Information. Stereochemical Stability of (−)-(aR,5S)-5c. The partial isomerization of (−)-(aR,5S)-5c (= anti) at 25 °C in toluene to the diastereomer (a S,5S)-5c (= syn) was examined as previously described.5 The conversion profile analyzed by a chiral HPLC is shown in Figure 7. The ΔG⧧ value20 was 102 kJ/mol. The method for determination of ΔG⧧ is shown in the Supporting Information. Procedure for in Vitro Binding Assay to Human V1a and V2 Receptors. All test compounds (shown in Table 1) had a purity of ≥99% as determined by HPLC analyses. The assays were performed according to the reported procedures.23,24 (1) Binding assay to hV1a:23 Human recombinant vasopressin V1a receptors expressed in HEK-293 cells were used in assay buffer (50 mM Tris-HCl, pH 7.4, 5 mM MgCl2, 0.1% BSA). A 0.26 μg aliquot was incubated with 0.03 nM [125I] phenylacetyl-D-Tyr(Me)-Phe-Gln-Asn-Arg-Pro-Arg-Tyr-NH2 for 2 h at 25 °C. Nonspecific binding was estimated in the presence of 1 μM (Arg8)-vasopressin. Receptors were filtered and washed, and the filters were then counted to determine [125I] phenylacetyl-D-Tyr(Me)Phe-Gln-Asn-Arg-Pro-Arg-Tyr-NH2 specifically bound. (2) Binding assay to hV2:24 CHO-K1 cells stably transfected with a plasmid encoding human vasopressin V2 receptors were used to prepare membranes in assay buffer (50 mM Tris-HCl, pH 7.4, 10 mM MgCl2, 0.1% BSA) using standard techniques. A 9 μg aliquot of membrane was incubated with 4 nM [3H] (Arg8)-vasopressin for 2 h at 25 °C. Nonspecific binding was estimated in the presence of 1 mM (Arg8)-vasopressin. Membranes were filtered and washed three times, and the filters were counted to determine [3H] (Arg8)-vasopressin specifically bound.

results may provide useful information for future drug design of biologically active compounds.

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EXPERIMENTAL SECTION22

5-Hydroxy-N-[4-[(2-methylbenzoyl)amino]benzoyl]-2,3,4,5tetrahydro-1-benzazepine (5a). Compound 5a (racemate) was prepared according to the procedure previously reported3 starting from 5-oxo-1-benzazepine (6a) via N-(p-aminobenzoyl)-5-oxo-1benzazepine (7a) as shown in Scheme 1. Because the physicochemical data of 5a including the 1H NMR spectral data were only briefly reported,3,12b in this paper the data were precisely examined to reveal the exact stereochemistry. The detailed 1H NMR spectral data in DMSO-d6 is shown in the Supporting Information, which indicates that 5a exists as an equilibrium mixture of anti/syn isomers with a 1:0.31 ratio in solution.17 5-Hydroxy-6-methyl-N-[4-[(2-methylbenzoyl)amino]benzoyl]-2,3,4,5-tetrahydro-1-benzazepine (5b). 6-Methyl compound (5b) was prepared from 6-methyl-5-oxo-1-benzazepine (6b)14 via 7b as shown in Scheme 1. The procedure is as follows. Step 1: To a stirred solution of 6b (275 mg, 1.57 mmol) in THF (10 mL) at 0 °C under argon, sodium hydride (60% in oil) (125 mg, 3.14 mmol) was added. The mixture was stirred at 25 °C for 30 min, cooled to 0 °C, and treated with 4-nitrobenzoyl chloride (873 mg, 4.71 mmol). After it was stirred at 25 °C for 15 h, the mixture was treated with H2O and extracted with ethyl acetate. The extract was washed with brine, dried over Na2SO4, and concentrated. The concentrate was purified by column chromatography (silica gel, ethyl acetate/hexane = 1:4) to afford 6-methyl-N-(4-nitrobenzoyl)-5-oxo-1-benzazepine (S1b) as white solids (506 mg, 1.56 mmol, 99%). To a stirred solution of S1b thus obtained (451 mg, 1.39 mmol) in EtOH (18 mL) was added palladium on carbon (10% Pd) (29 mg). After the mixture was stirred at 25 °C under a hydrogen atmosphere for 12 h, Pd was removed by filtration, and the filtrate was concentrated. The concentrate was purified by column chromatography (silica gel, ethyl acetate/hexane = 1:3) to afford N-(p-aminobenzoyl)-6-methyl-5-oxo-1-benzazepine (7b) as pale-yellow solids (373 mg, 1.27 mmol, 91%). Step 2: To a stirred solution of 7b (326 mg, 1.11 mmol) in CH2Cl2 (11 mL) was added EtN3 (185 mg, 1.83 mmol) and o-toluoyl chloride (205 mg, 1.33 mmol). After the mixture was stirred at 25 °C for 17 h, water was added to the mixture. The mixture was extracted with CH2Cl2. The extract was washed with 1 N HCl, saturated NaHCO3, and brine, dried over Na2SO4, and concentrated. The concentrate was purified by column chromatography (silica gel, ethyl acetate/hexane = 1:4) to afford 6-methyl-N-(o-methylbenzoyl)aminobenzoyl-5-oxo-1benzazepine (S2b) as white solids (454 mg, 1.10 mmol, 99%). To a solution of S2b thus obtained (171 mg, 0.414 mmol) in MeOH (6.0 mL) was added NaBH4 (24 mg, 0.621 mmol). After the mixture was stirred at 25 °C for 2 h, solvent was evaporated. The residue was diluted with CH2Cl2, washed with brine, dried over MgSO4, and concentrated. The concentrate was purified by column chromatography (silica gel, ethyl acetate/hexane = 1:2) to afford 5b (syn form) (racemate) as colorless crystals (144 mg, 0.347 mmol, 84%).16 5-Hydroxy-9-methyl-N-[4-[(2-methylbenzoyl)amino]benzoyl]-2,3,4,5-tetrahydro-1-benzazepine (5c). 9-Methyl derivative (5c) (racemate) was prepared from N-(p-aminobenzoyl)-9methyl-5-oxo-1-benzazepine (6c)14 via 7c according to a similar procedure described for the preparation of 5b from 6b via 7b. Compound 5c was formed as an anti/syn mixture with a ratio of ca. 1:0.17, from which only the major anti isomer could be isolated in the pure form with HPLC using a nonchiral column (YMC SIL-06).16 The physicochemical data of 5b, 5c, and the intermediates (S1b, S1c, 7b, 7c, S2b, and S2c) are shown in the Supporting Information. Separation of the N-Benzoyl-5-hydroxy-1-benzazepines (5a−c) into Enantiomers. The racemic compounds 5a−c were separated using chiral HPLC [Chiralpak IA (for 5a and 5b), and Chiral ART amylose-SA (for 5c)].16 As for 5a, the chiral synthesis of the enantiomers has been achieved.12a,b The separation conditions and physicochemical properties of the enantiomers are shown in the Supporting Information.

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ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jmedchem.7b00422.

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General experimental procedure, 1H-, 13C-, and 2DNMR spectra and physicochemical properties for new compounds and stereochemical stability of (−)-(aR,5S)5c (PDF) Molecular formula strings (CSV) X-ray crystal data for (+)-(aS,5R)-5a (CIF) X-ray crystal data for (+)-(aS,5S)-5b (CIF)

AUTHOR INFORMATION

Corresponding Author

*Phone: +81-42-685-3728. Fax: +81-42-685-3728. E-mail: [email protected]. ORCID

Hideaki Natsugari: 0000-0002-6442-1598 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was supported in part by a Grant-in-Aid for Scientific Research (C) (25460154) and a Grant-in-Aid for Young Scientists (B) (25860091) from the Japan Society for the Promotion of Science. H.T. thanks the MEXT-Supported Program for the Strategic Research Foundation at Private Universities (2013−2017) for financial support. 4507

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is consistent with the report that N-benzoyl-N-methylanilines exist in an E-structure: Azumaya, I.; Kagechika, H.; Fujiwara, Y.; Itoh, M.; Yamaguchi, K.; Shudo, K. Twisted intramolecular charge-transfer fluorescence of aromatic amides: conformation of the amide bonds in excited states. J. Am. Chem. Soc. 1991, 113, 2833−2838. (11) The numbering of the benzothiazepine rings of 2 and 3 used in the main text is different from that of the IUPAC nomenclature rule, i.e., the numbering of N and S in 2/3 are assigned as 1,4benzodiazepine (1) and 1-benzazepine (5), respectively, for convenience. (12) The tolvaptan-like compound (5a) in the racemic form was reported in ref 3 and in the chiral (5R and 5S)-forms appeared in the patent literature (ref 12a) and in the literature (ref 12b): (a) Ootsubo, K.; Yamashita, S.; Uchida, M.; Morita, K. Preparation of optically active 5-hydroxybenzoazepines as vasopressin antagonists. (Jpn Kokai Tokkyo Koho). JP 0680641, 1994. (b) Matsubara, J.; Kitano, K.; Otsubo, K.; Kawano, Y.; Ohtani, T.; Bando, M.; Kido, M.; Uchida, M.; Tabusa, F. Enantioselective synthesis of the metabolites of vasopressin V2 receptor antagonist OPC-31260 via lipase-catalyzed transesterification. Tetrahedron 2000, 56, 4667−4682. (13) The affinity at the rat VP receptors of 5a as the racemate was reported in ref 3, and that as the chiral (5R and 5S)-forms was briefly reported in the patent literature (ref 12a). The IC50 values (nM) for V1a (rat liver) and V2 (rat kidney) receptors reported are as follows: 5a (racemate), 140 for V1a and 29 for V2,3 (−)-(5S)-5a, 100 for V1a and 7.7 for V2,12a (+)-(5R)-5a, 140 for V1a and 32 for V2.12a In the literature, however, the detailed stereochemical analysis for exerting the activity was not performed. (14) Tabata, H.; Yoneda, T.; Tasaka, T.; Ito, S.; Oshitari, T.; Takahashi, H.; Natsugari, H. Stereochemistry of N-benzoyl-5substituted-1-benzazepines revisited: Synthesis of the conformationally biased derivatives and revision of the reported structure. J. Org. Chem. 2016, 81, 3136−3148. (15) Qadir, M.; Cobb, J.; Sheldrake, P. W.; Whittall, N.; White, A. J. P.; Hii, K. K.; Horton, P. N.; Hursthouse, M. B. Conformation analyses, dynamic behavior and amide bond distortions of mediumsized heterocycles. 1. Partially and fully reduced 1-benzazepines. J. Org. Chem. 2005, 70, 1545−1551. (16) As for 5a−c, workup after separation of the isomers (diastereomers and enantiomers) by column chromatography and/or HPLC was performed at low temperatures to prevent possible isomerization (e.g., evaporation of solvents at 0 °C and storage of the isolated compounds at −80 °C). (17) The ratio of anti/syn diastereomers of 5a observed in 1H NMR was almost the same (1:0.30−1:031) in other solvents (CDCl3, CD3OD, and CD3CN). (18) The absolute stereochemistry was determined based on the Flack parameter. (19) In particular, the (±)-angle of [α]D appears to be diagnostic for determination of the chirality at the axis from our amassed data for the related structures obtained thus far, i.e., compounds with (+)- and (−)-angles have (aS) and (aR) stereochemistry, respectively. In 5c, because the relative stereochemistry of the major anti isomer is (aR*,5S*), determination of the axial chirality leads inevitably to the determination of the central chirality at C5. (20) For determination of ΔG⧧ values, see Petit, M.; Lapierre, A. J. B.; Curran, D. P. Relaying asymmetry of transient atropisomers of oiodoanilides by radical cyclizations. J. Am. Chem. Soc. 2005, 127, 14994−14995. (21) If the differences of the intrinsic Ki values between (aS,5S)-5c and (aR,5S)-5c are 20-fold, and the formation of (aS,5S)-5c from (aR,5S)-5c is 10%, the observed Ki value of (aR,5S)-5c would be ca. half of the intrinsic value, which may be possible to be similar to that of the enantiomer (aS,5R)-5c. (22) For general experimental methods, see the Supporting Information. (23) Thibonnier, M.; Auzan, C.; Madhun, Z.; Wilkins, P.; BertiMattera, L.; Clauser, E. Molecular cloning, sequencing, and functional

ABBREVIATIONS USED ADPKD, autosomal-dominant polycystic kidney disease; VP, vasopressin; CD, circular dichroism; [α]D, optical rotation; ax, axial; eq, equatorial; ΔG‡, activation free-energy barrier to rotation

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REFERENCES

(1) For review articles on recent advances in VP receptor ligands, see: (a) Ryckmans, T. Advances in vasopressin receptor agonists and antagonists. Annu. Rep. Med. Chem. 2009, 44, 129−147. (b) Veeraveedu, P. T.; Palaniyandi, S. S.; Yamaguchi, K.; Komai, Y.; Thandavarayan, R. A.; Sukumaran, V.; Watanabe, K. Arginine vasopressin receptor antagonists (vaptans): pharmacological tools and potential therapeutic agents. Drug Discovery Today 2010, 15, 826− 841. (2) Kondo, K.; Ogawa, H.; Yamashita, H.; Miyamoto, H.; Tanaka, M.; Nakaya, K.; Kitano, K.; Yamamura, Y.; Nakamura, S.; Onogawa, T.; Mori, T.; Tominaga, M. 7-Chloro-5-hydroxy-1-[2-methyl-4-(2methylbenzoylamino)benzoyl]-2,3,4,5-tetrahydro-1H-1-benzazepine (OPC-41061): A potent, orally active nonpeptide arginine vasopressin V2 receptor antagonist. Bioorg. Med. Chem. 1999, 7, 1743−1754. (3) Ogawa, H.; Yamashita, H.; Kondo, K.; Yamamura, Y.; Miyamoto, H.; Kan, K.; Kitano, K.; Tanaka, M.; Nakaya, K.; Nakamura, S.; Mori, T.; Tominaga, M.; Yabuuchi, Y. Orally active, nonpeptide vasopressin V2 receptor antagonists: a novel series of 1-[4-(benzoylamino)benzoyl]-2,3,4,5-tetrahydro-1H-benzazepines and related compounds. J. Med. Chem. 1996, 39, 3547−3555. (4) Tolvaptan (SAMSCA/JINARC) was approved first as the drug for ADPKD in Japan in 2014, followed by the approval in Canada, Europe, and Korea in 2015. (5) Tabata, H.; Nakagomi, J.; Morizono, D.; Oshitari, T.; Takahashi, H.; Natsugari, H. Atropisomerism in the vaptan class of vasopressin receptor ligands: the active conformation recognized by the receptor. Angew. Chem., Int. Ed. 2011, 50, 3075−3079. (6) Yoneda, T.; Tabata, H.; Tasaka, T.; Oshitari, T.; Takahashi, H.; Natsugari, H. N-Benzoyl-1,5-benzothiazepine and its S-oxide as vasopressin receptor ligands: Insight into the active stereochemistry around the seven-membered ring. J. Med. Chem. 2015, 58, 3268−3273. (7) The description “E/Z” is used according to the IUPAC nomenclature around the N−C(=O) bond of the amide. The Eisomer has the cis relative arrangement of the two benzene rings of Nbenzoyl-1-benzazepines, and the Z-isomer has the trans arrangement. (8) The terms aS and aR are those of nomenclature based on the chiral axis, which correspond to P and M based on the helix nomenclature, respectively. The compounds may also be assigned using planar chirality nomenclature (pR/S), in which the −N1-Ar− plane is viewed as the chiral plane, and the carbon at −N(C=O) is designated as the pilot atom; thus, (aR) and (aS) correspond to (pS) and (pR), respectively. (9) For review articles on the relation between axial chirality and biological activity, see: (a) Kumarasamy, E.; Raghunathan, R.; Sibi, M. P.; Sivaguru, J. Nonbiaryl and heterobiaryl atropisomers: Molecular templates with promise for atropselective chemical transformations. Chem. Rev. 2015, 115, 11239−11300. (b) Ramig, K. Stereodynamic properties of medium-ring benzo-fused nitrogenous heterocycles: benzodiazepines, benzazepines, benzazocines, and benzazonines. Tetrahedron 2013, 69, 10783−10795. (c) Zask, A.; Murphy, J.; Ellestad, G. A. Biological stereoselectivity of atropisomeric natural products and drugs. Chirality 2013, 25, 265−274. (d) LaPlante, S. R.; Fader, L. D.; Fandrick, K. R.; Fandrick, D. R.; Hucke, O.; Kemper, R.; Miller, S. P. F.; Edwards, P. J. Assessing atropisomer axial chirality in drug discovery and development. J. Med. Chem. 2011, 54, 7005−7022. (e) Clayden, J.; Moran, W. J.; Edwards, P. J.; LaPlante, S. R. The challenge of atropisomerism in drug discovery. Angew. Chem., Int. Ed. 2009, 48, 6398−6401. (10) Although several very small peaks that may originate in the Zisomer were observed in the 1H NMR spectrum, the peaks were not sufficient to assign the structure. Formation of the E-structure in 5a−c 4508

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DOI: 10.1021/acs.jmedchem.7b00422 J. Med. Chem. 2017, 60, 4503−4509