Design, Synthesis and Structure-Activity Relationship Studies of (4-Alkoxyphenyl)glycinamides and Bioisosteric 1,3,4-Oxadiazoles as GPR88 Agonists

Abstract

Increasing evidence implicates the orphan G protein-coupled receptor 88 (GPR88) in a number of striatal-associated disorders. In this study, we report the design and synthesis of a series of novel (4-alkoxyphenyl)glycinamides (e.g., 31) and the corresponding 1,3,4-oxadiazole bioisosteres derived from the 2-AMPP scaffold (1) as GPR88 agonists. The 5-amino-1,3,4-oxadiazole derivatives (84, 88–90) had significantly improved potency and lower lipophilicity compared to 2-AMPP. Compound 84 had an EC₅₀ of 59 nM in the GPR88 overexpressing cell-based cAMP assay. In addition, 84 had an EC₅₀ of 942 nM in the [³⁵S]GTPγS binding assay using mouse striatal membranes but was inactive in membranes from GPR88 knockout mice, even at a concentration of 100 μM. In vivo pharmacokinetic testing of 90 in rats revealed that the 5-amino-1,3,4-oxadiazole analogues may have limited brain permeability. Taken together, these results provide the basis for further optimization to develop a suitable agonist to probe GPR88 functions in the brain.

Graphical Abstract

INTRODUCTION

The orphan G protein-coupled receptor 88 (GPR88) has recently attracted considerable interest in studying its biological functions, mainly through genetic interference. GPR88 is highly expressed in the striatum of the brain and is involved in both the striatonigral and striatopallidal pathways, suggesting the receptor may play a role in regulating striatal functions.^1-5 Genetic knockout^6-15 and transcriptional profiling studies^{3, 16-20} in rodents have suggested that GPR88 plays an important role in regulating the dopaminergic system and is implicated in a number of disorders such as Parkinson’s Disease, schizophrenia, anxiety, and drug addiction. Additionally, human genetic studies have demonstrated positive associations between the Gpr88 gene and schizophrenia²¹ and evidence that a Gpr88 variant was linked to childhood speech delay, learning disabilities, and chorea, indicating the relevance of GPR88 in the genetic risk for these diseases.²² Taken together, both animal and human data suggest that GPR88 is a potential novel drug target.

To date, the endogenous ligand for GPR88 has not been discovered. GPR88 is most closely related to the biogenic amine receptors, and has the highest sequence homology with the 5-HT_1d receptor and the β₃ adrenergic receptor (27% and 21% identity, respectively).¹ Chemogenomic analysis, based on the alignment of 30 critical residues predicted to line the binding cavity of GPCRs, clustered GPR88 with metabotropic glutamate and GABA_B receptors.²³ In order to characterize GPR88 signaling mechanisms and biological functions, our laboratory, as well as others, has carried out a medicinal chemistry campaign to develop GPR88 synthetic agonist probes.^24-32 We have previously reported that a synthetic small-molecule, 2-PCCA [(1R,2R)-2-(pyridin-2-yl)cyclopropane carboxylic acid ((2S,3S)-2-amino-3-methylpentyl)-(4′-propylbiphenyl-4-yl)amide, Figure 1], was able to activate GPR88 through a Gα_i-coupled signaling pathway in our TR-FRET-based Lance cAMP assay in GPR88 overexpressing CHO cells.²⁶ Recently, we have demonstrated that a potent, selective, and brain-penetrant GPR88 agonist RTI-13951-33 (Figure 1), derived from the 2-PCCA scaffold, significantly reduced alcohol self-administration and alcohol intake in a dose-dependent manner in rats when administered intraperitoneally and at doses that did not affect locomotor activity and sucrose self-administration.²⁹ These findings support the development and pharmacological validation of GPR88 agonists as a potential therapeutic to treat alcohol addiction.

Figure 1.

2-PCCA scaffold-based GPR88 agonists.

2-AMPP [(2S)-N-((1R)-2-amino-1-(4-(2-methylpentyloxy)-phenyl)ethyl)-2-phenylpropanamide (1, Figure 2)] is another promising GPR88 agonist scaffold for medicinal chemistry optimization.^{14, 28, 31} Early structure-activity relationship (SAR) studies of 1 have provided a preliminary understanding of receptor tolerances at three distinct sites for agonist activity (Figure 2).^{28, 31, 32} For example, the lipophilicity of the alkoxy group on site A is favorable for potency. The amino group on site B can be replaced by other functionalities (e.g., hydroxyl 2, ester 3, and amide 4), all of which have comparable or slightly improved EC₅₀ values relative to 1. Site C, on the other hand, has limited space for structural modifications, possibly involving a sterically defined aromatic stacking interaction with the GPR88 receptor.

Figure 2.

Structures of 2-AMPP (1) and 2–4, and preliminary SAR.

2-AMPP was moderately potent with an EC₅₀ of 414 nM in our TR-FRET-based Lance cAMP assay²⁸ and was reported to have a poor brain penetration due to its high lipophilicity (clog P = 4.53, calculated using Instant JChem 5.4.0, ChemAxon Ltd.).³¹ Recently, 2-AMPP (referring to compound 19 in the literature³³) was shown to have an EC₅₀ of 634 nM in the BRET-based cAMP assay in GPR88 overexpressing HEK293 cells, which is in line with our EC₅₀ value of this compound. We faced two major challenges for the development of 2-AMPP based agonists as in vivo probes: (a) potency and (b) brain bioavailability. To address these questions, we planned to further explore the SAR on sites A and B and reasoned that both potency and brain permeability can be improved by fine-tuning the lipophilicity on site A and modifying the functionality on site B. Herein, we report the design, synthesis, and pharmacological evaluation of a series of (4-alkoxyphenyl)glycinols, (4-alkoxyphenyl)glycinamides, and the corresponding bioisosteric 1,3,4-oxadiazoles as GPR88 agonists.

RESULTS AND DISCUSSSION

Chemistry.

The overall synthetic approach followed methods detailed in our earlier publication.²⁸ All the synthesized target compounds were characterized by ¹H NMR, ¹³C NMR, and HRMS, and determined to be >95% pure by HPLC analyses. The characterization data are in agreement with the assigned structures. The reaction yield is presented in the Experimental Section. Compounds 12–28 were synthesized following procedures depicted in Scheme 1. Boc-protection of the amino group in (R)-2-phenylglycine methyl ester (5) afforded 6. O-Alkylation of 6 with an appropriate alcohol under Mitsunobu conditions or via S_N2 substitution with an alkyl p-toluenesulfonate gave ethers 7. Reduction of the methyl ester with sodium borohydride in the presence of lithium chloride followed by the Boc group deprotection with HCl led to amino alcohols 8. Boc-removal of 7, followed by HBTU-mediated coupling with (S)-2-phenylpropionic acid provided amides 11. Alternatively, TIPS-protection of phenol 6 afforded 9, which was subjected to Boc-deprotection, followed by HBTU-mediated coupling with (S)-2-phenylpropionic acid and TIPS-deprotection to furnish phenol 10. Subsequent O-alkylation of the phenol group also provided the common intermediate 11. Finally, coupling of 8 with (S)-2-phenylpropionic acid using HBTU or reduction of the ester function in 11 furnished the target alcohols 12–28.

Scheme 1.

Synthesis of target compounds 12–28. Reagents and conditions: (a) Boc₂O, DIPEA, DCM, rt, overnight; (b) PPh₃, DEAD, alcohol, THF, rt, overnight; (c) NaBH₄, LiCl, THF-EtOH (1:1), rt, 3 h; (d) 4 M HCl in dioxane, DCM, rt, 16 h; or TFA: DCM, rt, 6 h; (e) (S)-2-phenylpropionic acid, HBTU, TEA, MeCN, rt, 5 h; (f) TIPSCl, imidazole, DCM, rt, overnight; (g) TBAF, THF, 0 °C, 3 h; (h) alkyl p-toluenesulfonate, K₂CO₃ MeCN, 65 °C, overnight.

Synthesis of amides 30–69 is outlined in Scheme 2. Briefly, acids 29, synthesized according to the procedure in our previous publication,²⁸ were coupled with an appropriate amine using the standard amide coupling reagents, such as Boc anhydride, EDC/HOBt, or by forming an acid chloride to give the target amides 30–69.

Scheme 2.

Synthesis of target compounds 30–69. Reagents and conditions: Method A (a) pyridine, dioxane, NH₄HCO₃, Boc₂O, rt, overnight; Method B (a) EDC hydrochloride, HOBt, DIPEA, amine, DMF, rt, overnight; Method C (a) oxalyl chloride, DMF (cat.), DCM, rt; then amine, Et₃N, rt, overnight.

Synthesis of 5-alkyl- and 5-amino-1,3,4-oxadiazoles is outlined in Schemes 3 and 4. As depicted in Scheme 3, reaction of ester 3 or 11h,²⁸ derived from 4-hydroxyphenylacetic acid, with hydrazine hydrate in refluxing ethanol afforded the corresponding hydrazides 70a,b. Coupling of 70a,b with trimethyl orthoformate or trimethyl orthoacetate in the presence of catalytic acid provided target 1,3,4-oxadiazoles 71–74. On the other hand, oxadiazoles derived from 4-hydroxyphenylpropanoic acid were synthesized according to the procedure shown in Scheme 4. Reaction of acid 75 with acetyl chloride in methanol at 65 °C afforded the methyl ester 76, which was protected with Boc anhydride to provide the Boc-protected amine 77. Mitsunobu reaction of 77 with the appropriate alcohol furnished the corresponding alkyl ethers 78a–f. Removal of the Boc group from the amine followed by HBTU-mediated coupling with (S)-2-phenylpropionic acid provided amides 80a–f. The ester function in 80 was converted to the corresponding hydrazide by heating the ester with hydrazine hydrate in ethanol. Finally, hydrazides 81a–f were condensed with trimethyl orthoformate, trimethyl orthoacetate or cyanogen bromide to furnish the target 1,3,4-oxadiazoles 82–91.

Scheme 3.

Synthesis of target compounds 71–74. Reagents and conditions: (a) hydrazine monohydrate, EtOH, reflux, 3 h; (b) CH(OMe)₃, PTSA, 85 °C, 2 h or CH₃C(OMe)₃, HOAc, m-xylene, reflux, 6 h.

Scheme 4.

Synthesis of target compounds 82–91. Reagents and conditions: (a) acetyl chloride, MeOH, reflux, overnight; (b) Boc₂O, DIPEA, DCM, rt, overnight; (c) PPh₃, DEAD, alcohol, rt, overnight; or alkyl p-toluenesulfonate, K₂CO₃, MeCN, 65 °C, overnight; (d) 4 M HCl in dioxane, DCM, rt, overnight; (e) (S)-2-phenylpropionic acid, HBTU, TEA, MeCN, rt, 5 h; (f) hydrazine monohydrate, EtOH, reflux, 3 h; (g) CH(OMe)₃, PTSA, 85 °C, 2 h; or CH₃C(OMe)₃, HOAc, m-xylene, reflux, 6 h; or CNBr, MeOH, reflux, 3 h.

Pharmacological Evaluation and SAR Study.

All synthesized compounds in this study were evaluated for GPR88 agonist activity in our previously established in vitro GPR88 Lance TR-FRET cAMP assay.²⁶ The TR-FRET signal (665 nm) was converted to fmol cAMP by interpolating from the standard cAMP curve. Fmol cAMP was plotted against the log of compound concentration and data were fit to a three-parameter logistic curve to generate maximum response (E_max) and EC₅₀ values. In our assay, 2-PCCA had an E_max of 100 ± 2 (mean ± S.E.M.) and RTI-13951-33 had an E_max of 103 ± 2 relative to 2-PCCA. Collectively, all of the active compounds in this study had E_max values comparable to 2-PCCA and RTI-13951-33, except for compounds 28 and 43, which had E_max values of 84% and 83%, respectively. These values might indicate that the compounds are partial agonists; however, it is important to note that E_max was calculated against synthetic agonists because the endogenous ligand for GPR88 has not yet been discovered. Figure 3 displays the concentration-response curves of representative compounds (RTI-13951-33, 84, and 90). Given the importance of an alkoxy substitution on site A in the 2-AMPP scaffold for GPR88 activity reported earlier, we first examined a series of ether analogues 12–28 by varying the length, shape, and steric and electronic properties, with the aim of identifying a side chain that can lower the lipophilicity while maintaining the potency of 2-{4-[(2-methylpentyl)oxy]phenyl)}glycinol 2 (clog P = 4.64, EC₅₀ = 195 nM). As can be seen from Table 1, the GPR88 agonist activity of this series was sensitive to the branching and length of the alkoxy side chain. First, the position of methyl branching was important for activity, as both 1-methylpentyl 12 (EC₅₀ = 380 nM) and 4-methylpentyl 13 (EC₅₀ = 282 nM) were less potent than the 2-methylpentyl analogue 2. Second, the branched alkyl was more potent than the linear alkyl group, as exemplified by 16 (EC₅₀ = 174 nM) relative to 14 (EC₅₀ = 295 nM). Third, the length of the alkyl group was important as potency decreased from n-pentyl to n-propyl (14, 18, and 20). Among the three cyclic alkyl analogues (17, 19, and 21), only the cyclobutylmethyl 19 was favored with an EC₅₀ of 234 nM. Attempts to add an additional oxygen atom into the side chain to reduce clog P resulted in a 10-fold loss of activity (22, 23). In general, the GPR88 agonist activity was correlated with lipophilicity of the compounds. The potency decreased as the clog P of compounds 12–23 decreased. Compounds 16 and 19 with an (S)-2-methylbutyl and a cyclobutylmethyl group, respectively, provided the best balance between potency and lipophilicity. To further explore SAR of the cyclobutylmethyl 19, we synthesized and tested a series of substituted analogues 24–28. Unfortunately, all of these compounds suffered from loss of potency; in particular 24, 26, and 27 were completely inactive, suggesting that there is a limited steric tolerance in this side chain position.

Figure 3.

Concentration-response curves of RTI-13951-33, 84, and 90 in the GPR88 Lance TR-FRET cAMP assay. The TR-FRET signal (665 nm) was converted to fmol cAMP by interpolating from the standard cAMP curve. Percent inhibition of forskolin-induced (300 nM) cAMP levels was plotted against the log of compound concentration and data were fit to a three-parameter logistic curve to generate EC₅₀ values. Each data point is the mean ± S.D. of at least three independent experiments performed in duplicate.

Table 1.

Biological data of 4-alkoxyphenylglycinols

Compound	R	clog Pa	cAMP pEC50 (EC50, nM)b	Emaxc
2-PCCA		6.19	7.14 ± 0.02 (73)	100 ± 2
RTI-13951-33		3.34	7.33 ± 0.05 (47)	103 ± 2
2	2-Methylpentyl	4.64	6.71 ± 0.09 (195)	100 ± 4
12	1-Methylpentyl	4.69	6.42 ± 0.03 (380)	96 ± 6d
13	4-Methylpentyl	4.56	6.55 ± 0.05 (282)	96 ± 1
14	n-Pentyl	4.28	6.53 ± 0.08 (295)	98 ± 3
15	2-Methylbutyl	4.20	6.60 ± 0.10 (251)	103 ± 2d
16	(S)-2-Methylbutyl	4.20	6.76 ± 0.08 (174)	97 ± 3d
17	Cyclopentylmethyl	4.18	6.59 ± 0.01 (257)	104 ± 1d
18	n-Butyl	3.83	6.42 ± 0.07 (380)	105 ± 13
19	Cyclobutylmethyl	3.73	6.63 ± 0.05 (234)	91 ± 8
20	n-Propyl	3.39	6.25 ± 0.04 (562)	98 ± 3
21	Cyclopropylmethyl	3.29	6.20 ± 0.04 (631)	96 ± 5
22	4-Methoxybutyl	3.04	5.33 ± 0.08 (4677)	95 ± 1
23	3-Methoxypropyl	2.52	5.23 ± 0.06 (5888)	94 ± 3
24		4.11	<5.00	N.D.
25		4.02	6.51 ± 0.04 (309)	97 ± 4d
26		4.32	<5.00	N.D.
27		3.86	<5.00	N.D.
28		3.22	5.89 ± 0.05 (1288)	84 ± 3d

^aclog P was calculated using Instant JChem 5.4.0 (ChemAxon Ltd.).

^bpEC₅₀ values are means ± standard error of at least three independent experiments performed in duplicate.

^cE_max value is % of 2-PCCA (mean ± standard error).

^dE_max value is % of RTI-13951-33 (mean ± standard error).

N.D., Not determined.

We next investigated site B with a rationale that modification of the amide functionality on this site would improve the potency of 4 (EC₅₀ = 616 nM, Figure 2). In addition, amide formation with a variety of readily available amines can rapidly expand structural diversity for SAR. To this end, we performed an in-depth examination of the substitution effects on the amide nitrogen by varying the size, lipophilicity, polarity, and electronic properties. The study began with the aliphatic substitutions on the amide nitrogen, as shown in Table 2. The primary amide 30 was equipotent to the tertiary N-dimethyl 4, whereas a monomethyl group (31) improved potency by 2-fold. The agonist activity increased further with an ethyl group (32), decreased with an n-propyl group (33), then maintained a moderate potency (EC₅₀ = 200–300 nM) with small to large-sized alkyl substitutions (34–42) except for cyclohexyl (43). The N-ethyl analogue 32 (EC₅₀ = 120 nM) emerged as the most potent compound in the amide series. Further modifications by adding polar functionalities at the terminal end of N-ethyl, such as ester (44), hydroxyl (45), ether (46, 47), carbamate (48, 50), and amine (49), led to a significant loss of potency. Interestingly, compounds (49, 51–54) with a protonatable nitrogen had the least activity. This SAR trend was also observed in the amide analogues with aromatic substitutions (Table 3). Phenyl and benzyl groups (55, 56) were favourable with an EC₅₀ of 245 nM and 219 nM, respectively, whereas pyridine rings (57–59), capable of forming a salt to improve aqueous solubility, were less active. Other polar five-membered heterocycles (60–69) were also poorly tolerated. It appeared that the activity deteriorated with heterocycles containing more heteroatoms. It should be noted that although two different side chains (2-methylpentyl and cyclobutylmethyl) on site B were used in Table 3, there was little difference in potency contributions between the two groups (60 vs. 61). Overall, the SAR suggested that compounds with a lipophilic alkyl substitution on the amide nitrogen tend to have better potency.

Table 2.

Biological data of 4-alkoxyphenylglycinamides

Compound	R	cAMP pEC50 (EC50, nM)a	Emaxb
4		6.21 ± 0.08 (616)	97 ± 4
30	H	6.24 ± 0.07 (575)	97 ± 7
31	Methyl	6.53 ± 0.08 (295)	116 ± 4
32	Ethyl	6.92 ± 0.11 (120)	99 ± 4c
33	n-Propyl	6.40 ± 0.04 (398)	113 ± 8
34	i-Propyl	6.74 ± 0.04 (182)	102 ± 3
35	t-Butyl	6.50 ± 0.01 (316)	109 ± 10c
36	Butan-2-yl	6.61 ± 0.04 (245)	96 ± 3
37	3-Methylbutan-2-yl	6.67 ± 0.07 (214)	93 ± 6c
38	Cyclopropyl	6.62 ± 0.04 (240)	98 ± 4
39	Cyclopropylmethyl	6.68 ± 0.09 (209)	94 ± 4c
40	Cyclobutyl	6.55 ± 0.04 (282)	98 ± 2
41	1-Methylcyclobutyl	6.63 ± 0.04 (234)	91 ± 4c
42	Cyclopentyl	6.68 ± 0.01 (209)	107 ± 4c
43	Cyclohexyl	6.25 ± 0.07 (562)	83 ± 2c
44	MeOOCCH2	6.33 ± 0.05 (468)	103 ± 8
45	HOCH2CH2	6.37 ± 0.12 (427)	99 ± 4
46	MeOCH2CH2	6.27 ± 0.02 (537)	107 ± 11
47		6.33 ± 0.04 (468)	110 ± 1c
48	BocNHCH2CH2	6.05 ± 0.07 (891)	97 ± 7c
49	NH2CH2CH2	5.10 ± 0.05 (7943)	129 ± 4c
50	MeN(Boc)CH2CH2	6.29 ± 0.03 (513)	111 ± 1c
51	MeNHCH2CH2	5.26 ± 0.06 (5495)	122 ± 4c
52	Me2NCH2CH2	5.26 ± 0.06 (5495)	103 ± 4c
53		5.47 ± 0.07 (3388)	120 ± 5c
54		<5.00	N.D.

^apEC₅₀ values are means ± standard error of at least three independent experiments performed in duplicate.

^bE_max value is % of 2-PCCA (mean ± standard error).

^cE_max value is % of RTI-13951-33 (mean ± standard error).

N.D., Not determined.

Table 3.

Biological data of 4-alkoxyphenylglycinamides containing aromatics

Compound	Structure	R	cAMP pEC50 (EC50, nM)a	Emaxb
55	A		6.61 ± 0.02 (245)	90 ± 9
56	A		6.66 ± 0.08 (219)	93 ± 4
57	A		6.21 ± 0.02 (617)	95 ± 5
58	A		6.23 ± 0.08 (589)	103 ± 4
59	A		6.06 ± 0.05 (871)	104 ± 2
60	A		6.57 ± 0.09 (269)	98 ± 4
61	B		6.26 ± 0.06 (550)	90 ± 2
62	B		6.14 ± 0.03 (724)	103 ± 0
63	B		6.34 ± 0.03 (457)	104 ± 3
64	B		6.17 ± 0.05 (676)	94 ± 5
65	B		6.13 ± 0.04 (741)	103 ± 3
66	B		6.00 ± 0.05 (1000)	116 ± 9
67	B		5.44 ± 0.04 (3631)	111 ± 0
68	B		5.31 ± 0.04 (4898)	114 ± 6
69	B		5.29 ± 0.08 (5129)	108 ± 4

^apEC₅₀ values are means ± standard error of at least three independent experiments performed in duplicate.

^bE_max value is % of RTI-13951-33 (mean ± standard error).

Bioisosteric replacement is an essential tool in the SAR study to improve potency, selectivity, and pharmacokinetics.³⁴ The oxadiazole moiety is stable to chemical and enzymatic degradation and capable of forming a hydrogen bond; therefore, it has been broadly used as a nonclassical bioisostere for ester and amide functionalities.³⁵ Because the 1,3,4-oxadiazoles have a lower lipophilicity (in general, an order of magnitude of clog P) compared to its 1,2,4-isomers, we selected 1,3,4-oxadiazole as our initial testing set. Replacement of the amide group in 4 with a 1,3,4-oxadiazole or a 5-methyl-1,3,4-oxadiazole moiety gave analogues 71 and 72, respectively, which were equipotent to 4 with EC₅₀ values in the 500–600 nM range (Table 4). There was no difference in potency between the 2-methylpentyl and cyclobutylmethyl side chains (73, 74 vs. 71, 72). Interestingly, the addition of a methylene linker between the oxadiazole moiety and the benzylic carbon led to a 5-fold increase in potency (82, 83 vs. 71, 72). Further modification by attaching an amino group to the 5-position gave the most potent compound 84 (EC₅₀ = 59 nM) in the series. Attempts to lower the lipophilicity by exchanging the 2-methylpentyl side chain in 82–84 with the cyclobutylmethyl group, unfortunately, resulted in less active compounds 85–87. After identifying a favorable 1,3,4-oxadiazole pharmacophore on site B, we turned our attention back to the side chain on site A, in which we reasoned that the chiral center of the methyl branching might have an impact on the potency. The (S)-isomer 88 and the (R)-isomer 89 had equivalent GPR88 activity with an EC₅₀ value of 78 nM and 74 nM, respectively, in line with the EC₅₀ value of the racemic mixture 84. However, in the case of the 2-methylbutyl side chain, (S)-90 was approximately 2-fold more potent than (R)-91, which is consistent with the observation in the corresponding hydroxyl (on site B) analogues that (S)-2-methylbutyl 16 is slightly more potent than racemic 15 (Table 1).

Table 4.

Biological data of bioisosteric 1,3,4-oxadiazoles

Compound	Structure	R	clog Pa	cAMP pEC50 (EC50, nM)b	Emaxc
71	A		4.26	6.27 ± 0.01 (537)	103 ± 5d
72	A		4.39	6.28 ± 0.06 (525)	101 ± 5d
73	B		3.35	6.32 ± 0.09 (479)	94 ± 0
74	B		3.48	6.26 ± 0.06 (550)	96 ± 4
82	A		4.35	6.95 ± 0.10 (112)	96 ± 4
83	A		4.47	6.83 ± 0.08 (148)	95 ± 3
84e	A		4.20	7.23 ± 0.03 (59)	99 ± 0
85	B		3.56	6.53 ± 0.03 (295)	104 ± 3
86	B		3.44	6.43 ± 0.01 (372)	98 ± 3
87e	B		3.29	6.86 ± 0.01 (138)	92 ± 9
88e	C		4.20	7.11 ± 0.02 (78)	97 ± 3
89e	D		4.20	7.13 ± 0.08 (74)	103 ± 3
90e	E		3.76	7.15 ± 0.08 (71)	101 ± 1
91e	F		3.76	6.86 ± 0.04 (138)	107 ± 2

^aclog P was calculated using Instant JChem 5.4.0 (ChemAxon Ltd.).

^bpEC₅₀ values are means ± standard error of at least three independent experiments performed in duplicate.

^cE_max value is % of RTI-13951-33 (mean ± standard error).

^dE_max value is % of 2-PCCA (mean ± standard error).

^eCompounds were tested as the HCl salt.

To further characterize the GPR88 agonist activity, we tested our best compound in the [³⁵S]GTPγS binding assay using mouse striatal membrane preparations. RTI-13951-33 (cAMP: EC₅₀ = 25 nM) increased [³⁵S]GTPγS binding with an EC₅₀ = 535 nM (E_max = 200%).²⁹ The E_max is expressed as percentage of activation above the basal binding, which is set as 100%, and the basal binding refers to binding in the absence of the agonist. Compound 84 also exhibited strong enhancement of [³⁵S]GTPγS binding activity (EC₅₀ = 942 nM, E_max = 229%) in mouse striatal membranes (Figure 4). Importantly, the compound was inactive in membranes prepared from GPR88 KO mice at concentrations tested up to 100 μM, indicating that it had a GPR88-specific agonist signaling activity in the striatum. It is worth noting that although the GPR88 agonist activity in the [³⁵S]GTPγS binding assay using a native tissue system is approximately 10- to 20-fold less potent than the cAMP assay in a GPR88 overexpressing cell line, the rank order of the potency of compounds is consistent between the two assay systems.

Figure 4.

[³⁵S]GTPγS binding of compound 84 in wild-type (WT) mouse striatal membranes vs GPR88 KO mouse striatal membranes. The data are the means of triplicate measurements with standard deviation shown as error bars.

Solubility and Preliminary PK Studies.

One of the major challenges for development of GPR88 probes is their ability to cross the blood–brain barrier (BBB) and have sufficient brain exposure to modulate receptor functions. Calculated physicochemical properties, such as lipophilicity (clog P) and topological polar surface area (TPSA), are useful indicators of a successful CNS drug. In general, a balance between clog P (2–4)³⁶ and TPSA (< 76 Ų)³⁷ would lead to good solubility and BBB permeability. Therefore, we tested the kinetic aqueous solubility (at pH = 7.4) and pharmacokinetic (PK) properties of select compounds to determine their drug-likeness. PK data for the in vivo effective agonist RTI-13951-33 are also presented for a comparison.²⁹ As shown in Table 5, both compounds 1 and 84 have a poor solubility of <1 μM, which is expected for compounds with a high clog P (4.53 and 4.20, respectively). Compound 90 (clog P = 3.76) has an increased solubility of 2.9 ± 0.3 μM, which confirms that while solubility is a challenge we still face, lowering lipophilicity does improve solubility. Compound 90 was further evaluated in a preliminary PK study to assess whether this compound has sufficient brain exposure. Following an intraperitoneal (i.p.) dose of 10 mg/kg in rats, 90 reached the peak plasma concentration of 270 ng/mL at 30 min (the first time point tested). The brain concentration also peaked at 30 min with a C_max of 39 ng/mL (92 nM), which is slightly above its EC₅₀ of 71 nM in the cAMP functional assay. The overall brain to plasma AUC ratio (B/P), as determined by AUC_0–inf ratio, was 0.1, indicating that 90 has limited brain penetration. As a comparison, RTI-13951-33 has a brain C_max of 287 ng/mL (539 nM) and a B/P ratio of 0.5 in rats (i.p., 10 mg/kg dose).²⁹ Compound 90 has a clog P of 3.76 but a high TPSA of 103 Ų, which likely limits its brain permeability. Further optimization of the 1,3,4-oxadiazole analogues is required to improve brain permeability.

Table 5.

Physicochemical, solubility and PK properties of compounds 1, 84, and 90.

Compounda	cAMP EC50 (nM)	clog Pb	TPSAb	Kinetic solubility at pH 7.4 (μM)	rat PK (i.p., 10 mg/kg)
					Plasma		Brain		B/P
					Cmax (ng/mL)	AUC0-inf (ng/mL • h)	Cmax (ng/mL)	AUC0-inf (ng/mL • h)
1	414	4.53	64.3	<1
84	59	4.20	103.3	<1
90	71	3.76	103.3	2.9 ± 0.3	270	1001	39	95	0.1
RTI-13951-33c	25	3.34	77.7		874	1510	287	825	0.5

^aAll compounds were tested as the HCl salt.

^bclog P and TPSA were calculated using Instant JChem 5.4.0 (ChemAxon Ltd.).

^cData were obtained from reference 29.

CONCLUSIONS

The orphan receptor GPR88 plays important roles in mediation of dopaminergic activity and striatal functions. To explore the therapeutic potential of this novel drug target, our group has carried out a medicinal chemistry campaign to develop GPR88 small-molecule agonist probes based on the 2-PCCA and 2-AMPP scaffolds.^25-29 The present study describes a series of novel (4-alkoxyphenyl)glycinols, (4-alkoxyphenyl)glycinamides, and the corresponding bioisosteric 1,3,4-oxadiazoles derived from 2-AMPP and explores their SAR requirements for high potency at the GPR88 receptor. Notably, 5-amino-1,3,4-oxadiazoles 84 (EC₅₀ = 59 nM) and 90 (EC₅₀ = 71 nM) emerged as the most potent compounds in this study. Compound 84 exhibited a significant [³⁵S]GTPγS binding activity (EC₅₀ = 942 nM) using the native tissue sample from mouse striatum but was inactive in GPR88 KO mice striatal membranes, even at a concentration of 100 μM, demonstrating that this type of compound has GPR88-specific agonist activity in the striatum. However, a preliminary PK study of 90 indicates limited brain permeability. Chemical modifications of 2-AMPP on site B (Figure 2) with other 5-membered heterocycles, as well as on site C, to further improve potency and ADME properties are currently underway. These studies will facilitate the identification of highly potent and brain-penetrant agonists to probe GPR88 functions in the brain.

EXPERIMENTAL SECTION

Chemistry.

General Methods.

All solvents and chemicals were reagent grade. Unless otherwise mentioned, all reagents and solvents were purchased from commercial vendors and used as received. Flash column chromatography was carried out on a Teledyne ISCO CombiFlash Rf system using prepacked columns. Solvents used include hexane, ethyl acetate (EtOAc), dichloromethane, and methanol. Purity and characterization of compounds were established by a combination of NMR, mass spectrometry, TLC, and HPLC analyses. ¹H and ¹³C NMR spectra were recorded on a Bruker Avance DPX-300 (300 MHz) spectrometer and were determined in CDCl₃, DMSO-d₆, or CD₃OD with tetramethylsilane (TMS) (0.00 ppm) or solvent peaks as the internal reference. Chemical shifts are reported in ppm relative to the reference signal and coupling constant (J) values are reported in hertz (Hz). Nominal mass spectra were obtained using an Agilent InfinityLab MSD single quadrupole mass spectrometer system (ESI). High resolution mass spectra (HRMS) were obtained using Agilent 1290 Infinity UHPLC-6230 TOF mass spectrometer (ESI). Thin layer chromatography (TLC) was performed on EMD precoated silica gel 60 F254 plates, and spots were visualized with UV light or iodine staining. CMA80 for column chromatography is a mixture of 80:18:2 chloroform/MeOH/NH₄OH. All final compounds were greater than 95% pure as determined by HPLC on a Waters 2695 Separation Module equipped with a Waters 2996 Photodiode Array Detector and a Phenomenex Synergi 4 mm Hydro-RP 80A C18 250 x 4.6 mm column using a flow rate of 1 mL/min starting with 1 min at 5% solvent B, followed by a 15 min gradient of 5-95% solvent B, followed by 9 min at 95% solvent B (solvent A, water with 0.1% TFA; solvent B, acetonitrile with 0.1% TFA and 5% water; absorbance monitored at 280 nm). All the synthesized target compounds were characterized by ¹H NMR, ¹³C NMR, and HRMS, and determined to be >95% pure by HPLC analyses.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-(4-hydroxyphenyl)acetate (6)

To a solution of (R)-2-phenylglycine methyl ester hydrochloride (5 g, 23 mmol) in DCM (175 mL) at 0 °C under nitrogen was added DIPEA (12 mL, 69 mmol), followed by di-tert-butyldicarbonate (5.2 g, 23 mmol). The reaction mixture was stirred at room temperature overnight and concentrated under reduced pressure. The residue was dissolved in EtOAc (200 mL), washed with 10% citric acid (3 x 50 mL), brine (3 x 50 mL), and dried (Na₂SO₄). Removal of the solvent under reduced pressure afforded crude 6 (6.45 g, 100% yield) as a colorless oil. ¹H NMR (300 MHz; CDCl₃) δ 7.16 (d, J = 9.0 Hz, 2H), 7.13 (d, J = 9.0 Hz, 2H), 6.44 (s, 1H), 5.60 (br d, J = 6.0 Hz, 1H), 5.22 (d, J = 6.0 Hz, 1H), 3.71 (s, 3H), 1.44 (s, 9H); ¹³C NMR (75 MHz; CDCl₃) δ 172.0, 156.3, 155.0, 128.5, 128.4, 115.8, 80.5, 57.1, 52.7, 28.3; MS (ESI) [M + H]⁺ m/z 282.3.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-{4-[(4-methylpentyl)oxy]phenyl}acetate (7b).

To a solution of 6 (200 mg, 0.71 mmol), 4-methylpentanol (177 μL, 1.42 mmol), and PPh₃ (316 mg, 1.21 mmol) in THF (10 mL) at room temperature under nitrogen was slowly added DEAD (0.19 mL, 1.21 mmol) dropwise, while keeping the reaction temperature below 35 °C. After addition, the reaction mixture was stirred at room temperature overnight and quenched with H₂O (10 mL). The layers were separated, and the aqueous layer was extracted with EtOAc (3 x 30 mL). The combined organic layers were washed with brine (3 x 30 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to chromatography on silica gel using 0–20% EtOAc in hexanes afforded 7b (130 mg, 50% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.85 (d, J = 9.0 Hz, 2H), 5.50 (br d, J = 6.0 Hz, 1H), 5.24 (d, J = 6.0 Hz, 1H), 3.92 (t, J = 6.0 Hz, 2H), 3.71 (s, 3H), 1.86–1.71 (m, 2H), 1.67–1.53 (m, 1H), 1.43 (s, 9H), 1.40–1.25 (m, 2H), 0.91 (d, J = 6.0 Hz, 6H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.3, 154.8, 128.7, 128.3, 114.8, 80.0, 68.4, 57.1, 52.5, 35.1, 28.3, 27.8, 27.1, 22.5; MS (ESI) m/z 366.6 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(pentyloxy)phenyl]acetate (7c).

The procedure for the synthesis of 7b was followed starting with 6 and n-pentanol to give 7c (50% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 5.50 (br d, J = 6.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.93 (t, J = 7.5 Hz, 2H), 3.71 (s, 3H), 1.82–1.71 (m, 2H), 1.43 (s, 9H), 1.42–1.25 (m, 4H), 0.92 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.3, 154.8, 128.7, 128.3, 114.8, 80.0, 68.0, 57.1, 52.5, 28.9, 28.3, 28.2, 22.4, 14.0; MS (ESI) m/z 352.3 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(butoxy)phenyl]acetate (7g).

The procedure for the synthesis of 7b was followed starting with 6 and n-butanol to give 7g (57% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 5.53 (br d, J = 9.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.94 (t, J = 7.5 Hz, 2H), 3.70 (s, 3H), 1.81–1.68 (m, 2H), 1.52–1.44 (m, 2H), 1.43 (s, 9H), 0.96 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.3, 154.8, 128.7, 128.3, 114.8, 80.0, 67.7, 57.1, 52.5, 31.3, 28.3, 19.2, 13.8; MS (ESI) m/z 338.6 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(cyclobutylmethoxy)phenyl]acetate (7h).

The procedure for the synthesis of 7b was followed starting with 6 and cyclobutylmethanol to give 7h (71% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.87 (d, J = 9.0 Hz, 2H), 5.50 (br d, J = 6.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.90 (d, J = 6.0 Hz, 2H), 3.70 (s, 3H), 2.84–1.68 (m, 1H), 2.19–2.06 (m, 2H), 2.01–1.80 (m, 4H), 1.43 (s, 9H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.4, 154.8, 128.8, 128.3, 114.9, 80.0, 72.1, 57.0, 52.6, 34.6, 28.3, 24.8, 18.6; MS (ESI) m/z 350.2 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-(4-propoxyphenyl)acetate (7i).

The procedure for the synthesis of 7b was followed starting with 6 and n-propanol to give 7i (57% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 5.54 (br d, J = 9.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.89 (t, J = 7.5 Hz, 2H), 3.70 (s, 3H), 1.85–1.72 (m, 2H), 1.43 (s, 9H), 1.02 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.3, 154.8, 128.7, 128.3, 114.8, 80.0, 69.5, 57.1, 52.5, 28.3, 22.5, 10.5; MS (ESI) m/z 324.3 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(cyclopropylmethoxy)phenyl]acetate (7j).

The procedure for the synthesis of 7b was followed starting with 6 and cyclopropylmethanol to give 7j (76% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.87 (d, J = 9.0 Hz, 2H), 5.54 (br d, J = 9.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.78 (d, J = 9.0 Hz, 2H), 3.70 (s, 3H), 1.43 (s, 9H), 1.32–1.16 (m, 1H), 0.67–0.58 (m, 2H), 0.39–0.28 (m, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.1, 154.8, 128.9, 128.4, 114.9, 80.0, 72.8, 57.0, 52.5, 28.3, 10.2, 3.2; MS (ESI) m/z 336.3 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(4-methoxybutoxy)phenyl]acetate (7k).

The procedure for the synthesis of 7b was followed starting with 6 and 4-methoxybutan-1-ol to give 7k (63% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 5.56 (br d, J = 6.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 3.96 (t, J = 6.0 Hz, 2H), 3.70 (s, 3H), 3.43 (t, J = 6.0 Hz, 2H), 3.33 (s, 3H), 1.91–1.68 (m, 4H), 1.43 (s, 9H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.1, 154.8, 128.8, 128.3, 114.8, 80.0, 72.3, 67.7, 58.5, 57.0, 52.5, 28.3, 26.2, 26.0; MS (ESI) m/z 368.4 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-[4-(4-methoxypropoxy)phenyl]acetate (7l).

The procedure for the synthesis of 7b was followed starting with 6 and 3-methoxypropan-1-ol to give 7l (68% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.26 (d, J = 9.0 Hz, 2H), 6.87 (d, J = 9.0 Hz, 2H), 5.55(br d, J = 6.0 Hz, 1H), 5.25 (d, J = 9.0 Hz, 1H), 4.03 (t, J = 6.0 Hz, 2H), 3.70 (s, 3H), 3.54 (t, J = 6.0 Hz, 2H), 3.34 (s, 3H), 2.08–1.97 (m, 2H), 1.43 (s, 9H); ¹³C NMR (75 MHz, CDCl₃) δ 171.9, 159.1, 154.8, 128.9, 128.3, 114.8, 80.0, 69.1, 64.9, 58.6, 57.0, 52.5, 29.5, 28.3; MS (ESI) m/z 354.5 [M + H]⁺.

(2R)-2-Amino-2-{4-[(4-methylpentyl)oxy]phenyl}ethan-1-ol Hydrochloride (8b).

To a suspension of NaBH₄ (35 mg, 0.93 mmol) in EtOH (1.5 mL) at 0 °C under nitrogen was added LiCl (39 mg, 0.93 mmol). After stirring at 0 °C for 10 min, a solution of 7b (130 mg, 0.36 mmol) in THF (1.5 mL) was added. The reaction mixture was stirred at room temperature for 3 h and quenched with saturated NH₄Cl solution (5 mL), followed by addition of H₂O (5 mL). The mixture was extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (20 mL), dried (Na₂SO₄), and concentrated under reduced pressure to give 125 mg of the crude intermediate alcohol. The Boc group was then deprotected with 4 M HCl in dioxane (2 mL) and DCM (5 mL). The reaction mixture was stirred at room temperature overnight and concentrated to give crude 8b (93 mg, 100% over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.21 (d, J = 9.0 Hz, 2H), 6.87 (d, J = 9.0 Hz, 2H), 4.01–3.84 (m, 3H), 3.74–3.60 (m, 1H), 3.51 (t, J = 9.0 Hz, 1H), 1.84–1.70 (m, 2H), 1.66–1.51 (m, 1H), 1.38–1.21 (m, 3H), 0.92 (d, J = 9.0 Hz, 6H); MS (ESI) free base m/z 238.3 [M + H]⁺.

(2R)-2-Amino-2-[4-(pentyloxy)phenyl]ethan-1-ol Hydrochloride (8c).

The procedure for the synthesis of 8b was followed starting with 7c to give crude 8c (100% yield over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.22 (d, J = 9.0 Hz, 2H), 6.87 (d, J = 9.0 Hz, 2H), 4.05–3.84 (m, 3H), 3.75–3.61 (m, 1H), 3.51 (t, J = 9.0 Hz, 1H), 1.84–1.70 (m, 2H), 1.50–1.29 (m, 5H), 0.93 (t, J = 7.5 Hz, 3H); MS (ESI) free base m/z 224.3 [M + H]⁺.

(2R)-2-Amino-2-(4-butoxyphenyl)ethan-1-ol Hydrochloride (8g).

The procedure for the synthesis of 8b was followed starting with 7g to give crude 8g (100% yield over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CD₃OD) δ 7.35 (d, J = 9.0 Hz, 2H), 6.98 (d, J = 9.0 Hz, 2H), 4.32–4.21 (m, 1H), 3.99 (t, J = 6.0 Hz, 2H), 3.87–3.72 (m, 2H), 1.82–1.67 (m, 2H), 1.58–1.42 (m, 2H), 0.98 (t, J = 7.5 Hz, 3H); MS (ESI) free base m/z 210.3 [M + H]⁺.

(2R)-2-Amino-2-(4-propoxyphenyl)ethan-1-ol Hydrochloride (8i)

The procedure for the synthesis of 8b was followed starting with 7i to give crude 8i (100% yield over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CD₃OD) δ 7.33 (d, J = 9.0 Hz, 2H), 6.95 (d, J = 9.0 Hz, 2H), 4.30–4.14 (m, 1H), 3.97–3.69 (m, 4H), 1.86–1.66 (m, 2H), 1.03 (t, J = 7.5 Hz, 3H); MS (ESI) free base m/z 196.2 [M + H]⁺.

(2R)-2-Amino-2-[4-(4-methoxybutoxy)phenyl]ethan-1-ol Hydrochloride (8k).

The procedure for the synthesis of 8b was followed starting with 7k to give crude 8k (100% yield over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39 (d, J = 9.0 Hz, 2H), 6.83 (d, J = 9.0 Hz, 2H), 5.54–5.28 (m, 1H), 4.53–4.30 (m, 1H), 4.00–3.60 (m, 4H), 3.49–3.36 (m, 2H), 3.32 (s, 3H), 1.86–1.57 (m, 4H); MS (ESI) free base m/z 240.4 [M + H]⁺.

(2R)-2-Amino-2-[4-(3-methoxypropoxy)phenyl]ethan-1-ol Hydrochloride (8l)

The procedure for the synthesis of 8b was followed starting with 7l to give crude 8l (100% yield over two steps) as an off-white foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39 (d, J = 9.0 Hz, 2H), 6.83 (d, J = 9.0 Hz, 2H), 5.47–5.36 (m, 1H), 4.47–4.33 (m, 1H), 3.98–3.84 (m, 3H), 3.80–3.68 (m, 1H), 3.49 (t, J = 6.0 Hz, 2H), 3.31 (s, 3H), 2.05–1.91(m, 2H); MS (ESI) free base m/z 226.2 [M + H]⁺.

Methyl (2R)-2-{[(tert-Butoxy)carbonyl]amino}-2-(4-{[tris(propan-2-yl)silyl]oxy}phenyl)acetate (9).

To a solution of 6 (1 g, 3.55 mmol) in dry DCM (20 mL) was added imidazole (532 mg, 7.82 mmol) and TIPS-Cl (837 mg, 3.91 mmol) at 0 °C. After stirring at room temperature for 16 h, the reaction was quenched with H₂O (2 mL) and the layers were separated. The aqueous layer was extracted with additional DCM (3 x 20 mL) and the combined organic layers were washed with brine (3 x 30 mL) and dried (Na₂SO₄). The solvent was removed under reduced pressure and the residue was subjected to chromatography on silica gel using 0–30% EtOAc in hexanes to furnish 9 (1.47 g, 95% yield) as a thick colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.5 Hz, 2H), 6.84 (d, J = 8.6 Hz, 2H), 5.42 (d, J = 6.3 Hz, 1H), 5.23 (d, J = 7.3 Hz, 1H), 3.71 (s, 3H), 1.43 (s, 9H), 1.31–1.16 (m, 3H), 1.12–1.05 (m, 18H); ¹³C NMR (75 MHz, CDCl₃) δ 172.0, 156.3, 155.0, 129.1, 128.3, 120.2, 80.1, 57.1, 52.5, 28.3, 17.9, 12.6; MS (ESI) m/z 460.0 [M + Na]⁺.

Methyl (2R)-2-(4-Hydroxyphenyl)-2-[(2S)-2-phenylpropanamido]acetate (10).

To a solution of 9 (968 mg, 2.25 mmol) in DCM (25 mL) was added TFA (2 mL) at 0 °C. After stirring for 16 h, the reaction was quenched with saturated NaHCO₃ (20 mL) and the layers were separated. The aqueous layer was extracted with additional DCM (3 x 20 mL) and the combined organic layers were washed with brine (3 x 20 mL), dried (Na₂SO₄), and concentrated. The residue was subjected to chromatography on silica gel using 0-30% EtOAc in hexanes (containing 1.5% Et₃N) to afford the free amine (650 mg, 85% yield) as a foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.22 (d, J = 8.6 Hz, 2H), 6.84 (d, J = 8.6 Hz, 2H), 4.58 (s, 1H), 3.69 (s, 3H), 2.33 (br s, 2H), 1.35–1.15 (m, 3H), 1.15–0.97 (m, 18H); ¹³C NMR (75 MHz, CDCl₃) δ 174.4, 156.0, 132.2, 127.9, 120.1, 58.1, 52.3, 17.9, 12.6; MS (ESI) m/z 322.0 [M + H⁺ - NH₃]⁺. To a solution of the amine intermediate (620 mg, 1.84 mmol) in MeCN (20 mL) at room temperature was added TEA (0.77 mL, 5.52 mmol), (S)-2-phenylpropionic acid (0.36 g, 2.39 mmol) and HBTU (1.05 g, 2.76 mmol). After stirring for 5 h, the reaction was quenched by H₂O (10 mL), followed by addition of EtOAc (50 mL). The layers were separated. The organic layer was washed with saturated NaHCO₃ (10 mL), brine (2 x 20 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to chromatography on silica gel using 0–30% EtOAc in hexanes to give the corresponding amide as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.16 (m, 5H), 7.02 (d, J = 8.6 Hz, 2H), 6.76 (d, J = 8.6 Hz, 2H), 6.32 (d, J = 7.0 Hz, 1H), 5.46 (d, J = 7.1 Hz, 1H), 3.73–3.52 (m, 4H), 1.49 (d, J = 7.1 Hz, 3H), 1.30–1.14 (m, 3H), 1.11–1.04 (m, 18H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.6, 156.2, 141.1, 128.8, 128.6, 128.2, 127.6, 127.2, 120.1, 55.9, 52.6, 46.7, 18.4, 17.9, 12.6; MS (ESI) m/z 470.0 [M + H]⁺. To a solution of the amide (864 mg, 1.84 mmol) in THF (20 mL) at 0 °C was added TBAF (1.0 M in THF, 2.76 mL, 2.76 mmol) via a syringe. After stirring at 0 °C for 4 h, the reaction was quenched by H₂O (15 mL) and diluted with EtOAc (20 mL). The layers were separated, and the aqueous layer was extracted with additional EtOAc (2 x 20 mL). The combined organic layers were washed with brine (2 x 2 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to chromatography on silica gel using 0–100% EtOAc in hexanes to furnish 10 (576 mg, 85% over two steps) as a yellowish waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.48–7.16 (m, 5H), 6.96 (d, J = 8.5 Hz, 2H), 6.78 (br s, 1H), 6.60 (d, J = 8.6 Hz, 2H), 6.48 (d, J = 6.6 Hz, 1H), 5.40 (d, J = 6.7 Hz, 1H), 3.73–3.51 (m, 4H), 1.51 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.4, 169.1, 153.9, 138.2, 126.4, 125.7, 125.1, 124.9, 113.3, 53.6, 50.3, 44.3, 15.7; MS (ESI) m/z 314.0 [M + H]⁺.

Methyl (2R)-2-[4-(Hexan-2-yloxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetate (11a)

The procedure for the synthesis of 7b was followed starting with 10 and 2-hexanol to give 11a (46% yield) as a sticky solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.17 (m, 5H), 7.11–7.00 (m, 2H), 6.83–6.68 (m, 2H), 6.33 (d, J = 6.9 Hz, 1H), 5.44 (d, J = 7.0 Hz, 1H), 4.39–4.23 (m, 1H), 3.67 (s, 3H), 3.66 – 3.55 (m, 1H), 1.80–1.63 (m, 1H), 1.63–1.52 (m, 1H), 1.50 (d, J = 7.2 Hz, 3H), 1.43–1.28 (m, 4H), 1.25 (d, J = 6.1 Hz, 3H), 0.89 (m, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.6, 158.3, 141.1, 128.8, 128.2, 127.9, 127.6, 127.2, 115.9, 73.9, 56.0, 53.0, 46.8, 36.1, 27.7, 22.6, 19.7, 18.4, 14.0; MS (ESI) m/z 398.0 [M + H]⁺.

Methyl (2R)-2-[4-(2-Methylbutoxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetate (11d).

The procedure for the synthesis of 7b was followed starting with 10 and 2-methyl-1-butanol to give 11d (29% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.20 (m, 5H), 7.19–7.01 (m, 2H), 6.86–6.75 (m, 2H), 6.31 (d, J = 7.0 Hz, 1H), 5.44 (d, J = 6.9 Hz, 1H), 3.82–3.53 (m, 6H), 1.89–1.75 (m, 1H), 1.63–1.42 (m, 4H), 1.37–1.10 (m, 1H), 0.99 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.6, 159.4, 128.8, 128.2, 127.6, 127.2, 114.8, 72.9, 56.0, 52.6, 46.8, 34.7, 26.1, 18.4, 16.5, 11.2; MS (ESI) m/z 384.0 [M + H]⁺.

Methyl (2R)-2-{4-[(2S)-2-Methylbutoxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetate (11e).

The procedure for the synthesis of 7b was followed with 10 and (S)-2-methyl-1-butanol to give 11e (41% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.18 (m, 5H), 7.07 (t, J = 8.7 Hz, 2H), 6..78 (d, J = 8.7 Hz, 2H), 6.34 (d, J = 6.6 Hz, 1H), 5.44 (d, J = 6.9 Hz, 1H), 3.84–3.52 (m, 6H), 1.92–1.70 (m, 1H), 1.64–1.44 (m, 4H), 1.35–1.11 (m, 1H), 0.98 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.6, 159.4, 141.1, 128.8, 128.2, 128.1, 127.6, 127.2, 114.8, 72.9, 56.0, 53.0, 46.8, 34.7, 26.1, 18.4, 16.5, 11.2; MS (ESI) m/z 384.0 [M + H]⁺.

Methyl (2R)-2-[4-(Cyclopentylmethoxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetate (11f).

The procedure for the synthesis of 7b was followed with 11 and (S)-2-methyl-1-butanol to furnish 9f (27% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.28 (ddd, J = 9.7, 7.3, 3.1 Hz, 5H), 7.08 (d, J = 8.7 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 6.35 (d, J = 6.9 Hz, 1H), 5.44 (d, J = 6.9 Hz, 1H), 3.77 (d, J = 6.9 Hz, 1H), 3.72–3.52 (m, 4H), 2.41–2.22 (m, 1H), 1.91–1.70 (m, 2H), 1.70–1.51 (m, 4H), 1.49 (d, J = 7.2 Hz, 3H), 1.43–1.14 (m, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.6, 159.4, 141.1, 128.8, 128.2, 127.6, 127.2, 114.8, 72.2, 56.0, 52.6, 46.8, 39.0, 29.4, 25.4, 18.4; MS (ESI) m/z 396.0 [M + H]⁺

Methyl (2R)-2-[4-(Cyclobutylmethoxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetate (11h).

To a solution of 7h (175 mg, 0.5 mmol) in DCM (5 mL) at room temperature was added 4 M HCl in dioxane (3 mL). The reaction mixture was stirred at room temperature overnight and concentrated to give crude amine hydrochloride. The crude amine (90 mg, 0.35 mmol) was then dissolved in MeCN (10 mL) followed by addition of TEA (0.16 mL, 1.1 mmol), (S)-2-phenylpropionic acid (56 mg, 0.37 mmol) and HBTU (170 mg, 0.45 mmol). After stirring for 5 h, the reaction was quenched by H₂O (5 mL), followed by addition of EtOAc (50 mL). The layers were separated. The organic layer was washed with saturated NaHCO₃ (10 mL), brine (10 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to chromatography on silica gel using 0–30% EtOAc in hexanes to furnish 11h (87 mg, 65% yield over two steps) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.20 (m, 5H), 7.08 (d, J = 9.0 Hz, 2H), 6.79 (d, J = 9.0 Hz, 2H), 6.36 (d, J = 6.0 Hz, 1H), 5.44 (d, J = 6.0 Hz, 1H), 3.87 (d, J = 9.0 Hz, 2H), 3.67 (s, 3H), 3.66–3.56 (m, 1H), 2.81–2.65 (m, 1H), 2.18–2.05 (m, 2H), 2.02–1.72 (m, 4H), 1.49 (d, J = 6.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.6, 159.4, 141.1, 128.9, 128.2, 127.6, 127.3, 114.9, 101.6, 72.1, 55.1, 52.6, 46.8, 34.6, 24.8, 18.6, 18.5; MS (ESI) m/z 382.5 [M + H]⁺.

Methyl (2R)-2-[4-(Cyclopropylmethoxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetate (11j)

The procedure for the synthesis of 11h was followed starting with 7j to give 11j (63% yield over two steps) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.33–7.20 (m, 5H), 7.08 (d, J = 9.0 Hz, 2H), 6.78 (d, J = 9.0 Hz, 2H), 6.43 (d, J = 6.0 Hz, 1H), 5.44 (d, J = 6.0 Hz, 1H), 4.74 (d, J = 9.0 Hz, 2H), 3.66 (s, 3H), 3.65–3.56 (m, 1H), 1.49 (d, J = 9.0 Hz, 3H), 1.30–1.14 (m, 1H), 0.66–0.57 (m, 2H), 0.34–0.28 (m, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.6, 159.1, 141.1, 128.9, 128.3, 127.6, 127.3, 114.8, 72.7, 56.0, 52.7, 46.7, 18.5, 10.2, 3.2; MS (ESI) m/z 368.5 [M + H]⁺.

Methyl(2R)-2-{4-[(1-Methylcyclobutyl)methoxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetate (11m).

To a solution of 10 (200 mg, 0.64 mmol) in anhydrous DMF (15 mL) at room temperature was added 1-methylcyclobutyl)methyl 4-methylbenzene-1-sulfonate (189 mg, 0.7 mmol), and K₂CO₃ (264 mg, 1.92 mmol). After stirring for 16 h at 60 °C, the reaction was quenched by H₂O (10 mL), followed by addition of EtOAc (15 mL). The layers were separated, and the aqueous layer was extracted with additional EtOAc (2 x 10 mL). The combined organic layers were washed with brine (4 x 15 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–50% EtOAc in hexanes to furnish 11m (15 mg, 6% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.41–7.23 (m, 5H), 7.17 (d, J = 8.7 Hz, 2H), 6.86 (d, J = 8.7 Hz, 2H), 6.30 (d, J = 6.7 Hz, 1H), 5.43 (d, J = 6.9 Hz, 1H), 3.73 (s, 2H), 3.66 (s, 3H), 3.59 (q, J = 7.2 Hz, 1H), 2.12–1.82 (m, 4H), 1.82–1.66 (m, 2H), 1.51 (d, J = 7.2 Hz, 3H), 1.21 (s, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.5, 159.8, 140.9, 128.9, 128.3, 127.7, 127.3, 114.9, 75.6, 56.0, 52.6, 46.9, 38.7, 30.1, 24.5, 18.4, 15.0; MS (ESI) m/z 396.0 [M + H]⁺.

Methyl (2R)-2-{4-[(3-Methylcyclobutyl)methoxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetate (11n).

The procedure for the synthesis of 7b was followed starting with 10 and (3-methylcyclobutyl)methanol to give 11n (27% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.19 (m, 5H), 7.08 (dd, J = 8.7, 2.2 Hz, 2H), 6.85–6.70 (m, 2H), 6.32 (d, J = 6.6 Hz, 1H), 5.44 (d, J = 6.9 Hz, 1H), 3.91 (d, J = 7.1 Hz, 1H), 3.80 (d, J = 6.3 Hz, 1H), 3.71–3.54 (m, 4H), 2.85–2.12 (m, 3H), 2.04–1.92 (m, 1H), 1.84–1.67 (m, 1H), 1.49 (d, J = 7.2 Hz, 3H), 1.45–1.35 (m, 1H), 1.12 (d, J = 6.9 Hz, 1.4 H), 1.04 (d, J = 6.1 Hz, 1.6 H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.6, 159.3, 141.1, 128.8, 128.2, 127.6, 1272, 114.8, 72.7, 72.2, 56.0, 52.6, 46.8, 31.2, 31.5, 30.6, 30.3, 27.0, 26.8, 22.3, 22.1, 18.4; MS (ESI) m/z 396.0 [M + H]⁺.

Methyl (2R)-2-{4-[(3,3-Dimethylcyclobutyl)methoxy]phenyl}-2-[(2S)-2-phenylpropanami do]acetate (11o).

The procedure for the synthesis of 11m was followed starting with 10 and (3,3-dimethylcyclobutyl)methyl-4-methylbenzene-1-sulfonate to give 11o (21% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.23 (m, 5H), 7.16 (d, J = 8.7 Hz, 2H), 6.82 (d, J = 8.7 Hz, 2H), 6.34 (d, J = 6.8 Hz, 1H), 5.43 (d, J = 6.9 Hz, 1H), 3.86 (d, J = 6.6 Hz, 2H), 3.65 (s, 3H), 3.59 (q, J = 7.2 Hz, 1H), 2.75–2.50 (m, 1H), 1.96–1.81 (m, 2H), 1.68–1.54 (m, 2H), 1.50 (d, J = 7.2 Hz, 3H), 1.17 (s, 3H), 1.08 (s, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.4, 159.4, 140.9, 128.9, 128.3, 127.7, 127.3, 114.9, 73.0, 56.0, 52.6, 46.9, 37.7, 31.9, 30.9, 28.9, 27.2, 18.4; MS (ESI) m/z 410.0 [M + H]⁺.

Methyl (2R)-2-[(2S)-2-Phenylpropanamido]-2-[4-({spiro[2.3]hexan-5-yl}methoxy)phenyl]acetate (11p).

The procedure for the synthesis of 11m (except, MeCN was used instead of DMF) was followed starting with 10 and (3-methylidenecyclobutyl)methyl 4-methylbenzene-1-sulfonate to give the corresponding olefin intermediate: methyl (2R)-2-{4-[(3-methylidenecyclobutyl)methoxy]phenyl}-2-[(2S)-2-phenylpropanamido] acetate (12% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.43–7.21 (m, 5H), 7.16 (d, J = 8.6 Hz, 2H), 6.83 (d, J = 8.6 Hz, 2H), 6.39 (d, J = 6.7 Hz, 1H), 5.42 (d, J = 6.9 Hz, 1H), 4.88–4.68 (m, 2H), 3.93 (d, J = 6.7 Hz, 2H), 3.64 (s, 3H), 3.58 (q, J = 7.2 Hz, 1H), 2.92–2.78 (m, 2H), 2.78–2.62 (m, 1H), 2.56–2.44 (m, 2H), 1.49 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.4, 159.2, 146.3, 140.9, 128.9, 128.6, 128.4, 127.7, 127.3, 114.9, 106.7, 71.6, 56.0, 52.6, 46.8, 34.6, 29.2, 18.5; MS (ESI) m/z 394.0 [M + H]⁺. The olefin function of this material was transformed into the corresponding cyclopropyl function under Shi³⁸ modified Simmons-Smith³⁹ reaction. To a solution of Et₂Zn (171.5 μL, 0.17 mmol) in DCM (1 mL) at 0 °C was added TFA (13.1 μL, 0.17 mmol). After stirring at 0 °C for 1 h, diiodomethane (13.8 μL, 0.17 mmol) was added and stirred for another 40 min at 0 °C. At that time, the olefin intermediate (27 mg, 0.07 mmol) was dissolved in DCM (1 mL) and added slowly to the above reaction via a syringe at 0 °C. The reaction, which resulted, was stirred under N₂ at room temperature for 2 h. After that, the reaction was quenched by cold saturated NH₄Cl (2 mL) and diluted with EtOAc (5 mL). The layers were separated, and the aqueous layer was extracted with additional EtOAc (2 x 10 mL). The combined organic layers were washed with saturated NaHCO₃ (5 mL), brine (3 x 10 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–25% EtOAc in hexanes to furnish 11p (20 mg, 72% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.41–7.22 (m, 5H), 7.17 (d, J = 8.6 Hz, 2H), 6.85 (d, J = 8.7 Hz, 2H), 6.30 (d, J = 6.7 Hz, 1H), 5.43 (d, J = 6.9 Hz, 1H), 4.01 (d, J = 7.1 Hz, 2H), 3.66 (s, 3H), 3.59 (q, J = 7.2 Hz, 1H), 2.92–2.73 (m, 1H), 2.37–2.14 (m, 2H), 1.93 (dd, J = 12.3, 5.8 Hz, 2H), 1.51 (d, J = 7.2 Hz, 3H), 0.41 (s, 4H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 171.4, 159.4, 140.9, 128.9, 128.4, 128.3, 127.7, 127.3, 115.0, 72.5, 56.0, 52.6, 46.9, 33.4, 29.8, 18.4, 16.9, 12.1, 11.7; MS (ESI) m/z 408.0 [M + H]⁺.

Methyl (2R)-2-{4-[(3,3-Difluorocyclobutyl)methoxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetate (11q).

The procedure for the synthesis of 11m was followed starting with 10 and (3,3-difluorocyclobutyl)methyl 4-methylbenzene-1-sulfonate to give 11q (8% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.21 (m, 5H), 7.09 (d, J = 8.6 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 6.34 (d, J = 6.8 Hz, 1H), 5.45 (d, J = 6.9 Hz, 1H), 3.94 (d, J = 5.8 Hz, 2H), 3.72–3.54 (m, 4H), 2.84–2.34 (m, 5H), 1.49 (d, J = 7.1 Hz, 3H); MS (ESI) m/z 418.0 [M + H]⁺.

(2S)-N-[(1R)-1-[4-(Hexan-2-yloxy)phenyl]-2-hydroxyethyl]-2-phenylpropanamide (12).

To a suspension of NaBH₄ (20.9 mg, 0.55 mmol) in EtOH (6 mL) at 0 °C under nitrogen was added LiCl (23.4 mg, 0.55 mmol). After stirring at 0 °C for 10 min, a solution of 11a (88 mg, 0.22 mmol) in THF (6 mL) was added. The reaction mixture was stirred at room temperature for 3 h and quenched with saturated NH₄Cl solution (5 mL), followed by addition of H₂O (10 mL). The mixture was extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (20 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–30% EtOAc in hexanes to furnish 12 (45 mg, 55% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.44–7.20 (m, 5H), 6.93 (d, J = 8.7 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.14 (d, J = 7.1 Hz, 1H), 4.94 (dd, J = 11.2, 6.0 Hz, 1H), 4.42–4.04 (m, 1H), 3.73 (d, J = 5.5 Hz, 2H), 3.62 (q, J = 7.1 Hz, 1H), 2.98 (br s, 1H), 1.79–1.61 (m, 1H), 1.61–1.46 (m, 4H), 1.46–1.28 (m, 4H), 1.25 (d, J = 6.1 Hz, 3H), 0.90 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 157.7, 141.3, 130.6, 128.9, 127.6, 127.3, 116.0, 73.9, 66.5, 55.3, 47.0, 36.2, 27.7, 22.6, 19.7, 18.5, 14.0; HRMS (ESI) m/z calcd for C₂₃H₃₁NO₃ [M + H]⁺ 370.2377, m/z found 370.2371.

(2S)-N-[(1R)-2-Hydroxy-1-{4-[(4-methylpentyl)oxy]phenyl}ethyl]-2-phenylpropanamide (13).

To a solution of 8b (96 mg, 0.35 mmol) in MeCN (10 mL) at room temperature was added TEA (0.16 mL, 1.1 mmol), (S)-2-phenylpropionic acid (56 mg, 0.37 mmol), and HBTU (170 mg, 0.45 mmol). After stirring for 5 h, the reaction was quenched by H₂O (5 mL), followed by addition of EtOAc (50 mL). The layers were separated. The organic layer was washed with saturated NaHCO₃ (10 mL), brine (10 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to chromatography on silica gel using 0–30% EtOAc in hexanes to provide 13 (55 mg, 43% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.20 (m, 5H), 6.95 (d, J = 9.0 Hz, 2H), 6.78 (d, J = 9.0 Hz, 2H), 6.08 (d, J = 9.0 Hz, 1H), 4.99–4.90 (m, 1H), 3.88 (t, J = 7.5 Hz, 2H), 3.75 (br s, 2H), 3.68–3.56 (m, 1H), 2.83 (br s, 1H), 1.81–1.69 (m, 2H), 1.64–1.54 (m, 1H), 1.51 (d, J = 9.0 Hz, 3H), 1.36–1.23 (m, 2H), 0.91 (d, J = 6.0 Hz, 6H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.6, 141.2, 130.6, 128.9, 127.5, 127.3, 114.7, 68.3, 66.6, 55.3, 47.0, 35.1, 27.8, 27.1, 22.5, 18.4; HRMS (ESI) m/z calcd for C₂₃H₃₁NO₃ [M + H]⁺ 370.2377, m/z found 370.2375.

(2S)-N-{(1R)-2-Hydroxy-1-[4-(pentyloxy)phenyl]ethyl}-2-phenylpropanamide (14).

The procedure for the synthesis of 13 was followed starting with 8c to give 14 (43% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.20 (m, 5H), 6.95 (d, J = 9.0 Hz, 2H), 6.77 (d, J = 9.0 Hz, 2H), 6.03 (d, J = 6.0 Hz, 1H), 5.00–4.89 (m, 1H), 3.89 (t, J = 6.0 Hz, 2H), 3.76 (d, J = 6.0 Hz, 2H), 3.68–3.55 (m, 1H), 2.73 (br s, 1H), 1.85–1.67 (m, 2H), 1.51 (d, J = 6.0 Hz, 3H), 1.46–1.25 (m, 4H), 0.92 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.7, 141.2, 130.6, 128.9, 127.6, 127.5, 127.3, 114.7, 68.0, 66.7, 55.4, 47.1, 28.9, 28.2, 22.4, 18.4, 14.0; HRMS (ESI) m/z calcd for C₂₂H₂₉NO₃ [M + H]⁺ 356.2220, m/z found 356.2219.

(2S)-N-[(1R)-2-Hydroxy-1-[4-(2-methylbutoxy)phenyl]ethyl]-2-phenylpropanamide (15).

The procedure for the synthesis of 12 was followed starting with 11d to give 15 (67% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.43–7.16 (m, 5H), 6.95 (d, J = 8.6 Hz, 2H), 6.78 (d, J = 8.6 Hz, 2H), 5.98 (d, J = 6.9 Hz, 1H), 4.95 (dd, J = 11.8, 5.0 Hz, 1H), 3.81–3.72 (m, 3H), 3.71–3.57 (m, 2H), 2.61 (t, J = 5.7 Hz, 1H), 1.94–1.73 (m, 1H), 1.64–1.44 (m, 4H), 1.37–1.12 (m, 1H), 0.99 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.9, 141.2, 130.5, 128.9, 127.6, 127.5, 127.3, 114.8, 72.9, 66.8, 55.4, 47.1, 34.7, 26.1, 18.4, 16.5, 11.2; HRMS (ESI) m/z calcd for C₂₂H₂₉NO₃ [M + H]⁺ 356.2220, m/z found 356.2213.

(2S)-N-[(1R)-2-Hydroxy-1-{4-[(2S)-2-methylbutoxy]phenyl}ethyl]-2-phenylpropanamide (16).

The procedure for the synthesis of 12 was followed starting with 11e to give 16 (69% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.43–7.19 (m, 5H), 6.94 (d, J = 8.6 Hz, 2H), 6.76 (d, J = 8.7 Hz, 2H), 6.11 (d, J = 7.1 Hz, 1H), 4.93 (dd, J = 11.9, 5.3 Hz, 1H), 3.82–3.51 (m, 5H), 2.90 (br s, 1H), 1.94–1.72 (m, 1H), 1.61–1.43 (m, 4H), 1.33–1.14 (m, 1H), 0.99 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.8, 141.3, 130.6, 128.9, 127.6, 127.5, 127.3, 114.7, 72.9, 66.6, 55.3, 47.0, 34.7, 26.1, 18.4, 16.5, 11.3; HRMS (ESI) m/z calcd for C₂₂H₂₉NO₃ [M + H]⁺ 356.2220, m/z found 356.2214.

(2S)-N-[(1R)-1-[4-(Cyclopentylmethoxy)phenyl]-2-hydroxyethyl]-2-phenylpropanamide (17).

The procedure for the synthesis of 12 was followed starting with 11f to give 17 (58% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.44–7.17 (m, 5H), 6.95 (d, J = 8.7 Hz, 2H), 6.77 (d, J = 8.7 Hz, 2H), 6.01 (d, J = 6.9 Hz, 1H), 4.95 (dd, J = 11.9, 5.0 Hz, 1H), 3.76 (dd, J = 6.0, 3.2 Hz, 4H), 3.62 (q, J = 7.1 Hz, 1H), 2.67 (t, J = 6.0 Hz, 1H), 2.45–2.20 (m, 1H), 1.90–1.69 (m, 3H), 1.69–1.55 (m, 3H), 1.52 (t, J = 6.2 Hz, 3H), 1.34 (dt, J = 11.6, 7.2 Hz, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.8, 141.2, 130.6, 128.9, 127.6, 127.5, 127.3, 114.8, 72.3, 66.7, 55.4, 47.1, 39.0, 29.4, 25.4, 18.4; HRMS (ESI) m/z calcd for C₂₃H₂₉NO₃ [M + H]⁺ 368.2220, m/z found 368.2215.

(2S)-N-[(1R)-1-(4-Butoxyphenyl)-2-hydroxyethyl]-2-phenylpropanamide (18).

The procedure for the synthesis of 13 was followed starting with 8g to give 18 (33% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.23 (m, 5H), 6.96 (d, J = 9.0 Hz, 2H), 6.78 (d, J = 9.0 Hz, 2H), 5.99 (d, J = 6.0 Hz, 1H), 5.00–4.89 (m, 1H), 3.91 (t, J = 7.5 Hz, 2H), 3.76 (d, J = 3.0 Hz, 2H), 3.69–3.57 (m, 1H), 2.62 (br s, 1H), 1.82–1.66 (m, 2H), 1.52 (d, J = 6.0 Hz, 3H), 1.51–1.37 (m, 2H), 0.96 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.7, 141.2, 130.6, 128.9, 127.6, 127.5, 127.3, 114.8, 67.7, 66.8, 55.4, 47.1, 31.3, 19.2, 18.4, 13.8; HRMS (ESI) m/z calcd for C₂₁H₂₇NO₃ [M + H]⁺ 342.2064, m/z found 342.2071.

(2S)-N-{(1R)-1-[4-(Cyclobutylmethoxy)phenyl]-2-hydroxyethyl}-2-phenylpropanamide (19).

The procedure for the synthesis of 12 was followed starting with 11h to give 19 (83% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.22 (m, 5H), 6.95 (d, J = 9.0 Hz, 2H), 6.77 (d, J = 9.0 Hz, 2H), 6.09 (d, J = 6.0 Hz, 1H), 4.98–4.88 (m, 1H), 3.86 (d, J = 6.0 Hz, 2H), 3.74 (br s, 2H), 3.68–3.56 (m, 1H), 2.86 (br t, J = 6.0 Hz, 1H), 2.81–2.65 (m, 2H), 2.20–2.04 (m, 2H), 2.20–1.75 (m, 4H), 1.51 (d, J = 9.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.8, 141.2, 130.7, 128.9, 127.5, 127.3, 114.8, 72.1, 66.6, 55.3, 47.0, 34.6, 24.8, 18.5, 18.4; HRMS (ESI) m/z calcd for C₂₂H₂₇NO₃ [M + H]⁺ 354.2064, m/z found 354.2064.

(2S)-N-[(1R)-2-Hydroxy-1-(4-propoxyphenyl)ethyl]-2-phenylpropanamide (20).

The procedure for the synthesis of 13 was followed starting with 8i to give 20 (32% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.22 (m, 5H), 6.95 (d, J = 9.0 Hz, 2H), 6.77 (d, J = 9.0 Hz, 2H), 6.08 (d, J = 6.0 Hz, 1H), 4.99–4.90 (m, 1H), 3.96 (t, J = 7.5 Hz, 2H), 3.75 (br s, 2H), 3.67–3.56 (m, 1H), 2.83 (br t, J = 4.5 Hz, 1H), 1.83–1.68 (m, 2H), 1.51 (d, J = 9.0 Hz, 3H), 1.01 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.6, 141.2, 130.6, 128.9, 127.5, 127.3, 114.7, 69.5, 66.6, 55.3, 47.0, 22.5, 18.4, 10.4; HRMS (ESI) m/z calcd for C₂₀H₂₅NO₃ [M + H]⁺ 328.1907, m/z found 328.1911.

(2S)-N-{(1R)-1-[4-(Cyclopropylmethoxy)phenyl]-2-hydroxyethyl}-2-phenylpropanamide (21).

The procedure for the synthesis of 12 was followed starting with 11j to give 21 (82% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.19 (m, 5H), 6.94 (d, J = 9.0 Hz, 2H), 6.77 (d, J = 9.0 Hz, 2H), 6.11 (d, J = 6.0 Hz, 1H), 4.99–4.88 (m, 1H), 3.73 (d, J = 6.0 Hz, 3H), 3.68–3.54 (m, 1H), 2.89 (br t, J = 6.0 Hz, 1H), 1.50 (d, J = 6.0 Hz, 4H), 1.32–1.14 (m, 1H), 0.69–0.56 (m, 2H), 0.36–0.26 (m, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 174.5, 158.5, 141.2, 130.8, 128.9, 127.5, 127.3, 114.8, 72.7, 66.5, 55.2, 47.0, 18.4, 10.2, 3.1; HRMS (ESI) m/z calcd for C₂₁H₂₅NO₃ [M + H]⁺ 340.1907, m/z found 340.1910.

(2S)-N-[(1R)-2-Hydroxy-1-[4-(4-methoxybutoxy)phenyl]ethyl}-2-phenylpropanamide (22).

The procedure for the synthesis of 13 was followed starting with 8k to give 22 (82% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.20 (m, 5H), 6.94 (d, J = 9.0 Hz, 2H), 6.74 (d, J = 9.0 Hz, 2H), 6.29 (d, J = 6.0 Hz, 1H), 4.97–4.86 (m, 1H), 3.90 (t, J = 6.0 Hz, 2H), 3.70 (br s, 2H), 3.66–3.55 (m, 1H), 3.41 (t, J = 6.0 Hz, 2H), 3.32 (s, 3H), 3.27 (br s, 1H), 1.86–1.63 (m, 4H), 1.48 (d, J = 9.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.5, 158.3, 141.2, 130.8, 128.8, 127.4, 127.1, 114.5, 72.2, 67.5, 66.2, 58.4, 55.1, 46.8, 26.1, 25.9, 18.4; HRMS (ESI) m/z calcd for C₂₂H₂₉NO₄ [M + H]⁺ 372.2169, m/z found 372.2173.

(2S)-N-[(1R)-2-Hydroxy-1-[4-(3-methoxypropoxy)phenyl]ethyl}-2-phenylpropanamide (23).

The procedure for the synthesis of 13 was followed starting with 8l to give 23 (73% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.34–7.20 (m, 5H), 6.93 (d, J = 9.0 Hz, 2H), 6.75 (d, J = 9.0 Hz, 2H), 6.36 (d, J = 6.0 Hz, 1H), 4.96–4.87 (m, 1H), 3.97 (t, J = 6.0 Hz, 2H), 3.76–3.55 (m, 3H), 3.51 (t, J = 6.0 Hz, 2H), 3.45 (br s, 1H), 3.32 (s, 3H), 2.05–1.93 (m, 2H), 1.47 (d, J = 6.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.5, 158.2, 141.2, 130.9, 128.7, 127.4, 127.1, 114.5, 69.1, 66.0, 64.7, 58.5, 55.0, 48.8, 29.4, 18.3; HRMS (ESI) m/z calcd for C₂₁H₂₇NO₄ [M + H]⁺ 358.2013, m/z found 358.2017.

(2S)-N-[(1R)-2-Hydroxy-1-{4-[(1-methylcyclobutyl)methoxy]phenyl}ethyl]-2-phenylpropanamide (24).

The procedure for the synthesis of 12 was followed starting with 11m to give 24 (83% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.46–7.21 (m, 5H), 7.08 (d, J = 8.6 Hz, 2H), 6.86 (d, J = 8.6 Hz, 2H), 5.91 (d, J = 6.5 Hz, 1H), 4.96 (dd, J = 11.1, 5.9 Hz, 1H), 3.86–3.67 (m, 4H), 3.60 (q, J = 7.2 Hz, 1H), 2.58 (t, J = 6.0 Hz, 1H), 2.09–1.81 (m, 4H), 1.81–1.68 (m, 2H), 1.54 (d, J = 7.2 Hz, 3H), 1.22 (s, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.8, 159.4, 141.2, 130.6, 129.0, 127.6, 127.5, 127.4, 114.9, 75.7, 66.8, 55.7, 47.2, 38.7, 30.1, 24.5, 18.5, 15.0; HRMS (ESI) m/z calcd for C₂₃H₂₉NO₃ [M + H]⁺ 368.2220, m/z found 368.2214.

(2S)-N-[(1R)-2-Hydroxy-1-{4-[(3-methylcyclobutyl)methoxy]phenyl}ethyl]-2-phenylpropanamide (25).

The procedure for the synthesis of 12 was followed starting with 11n to give 25 (65% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.09 (m, 5H), 6.94 (dd, J = 8.6, 2.0 Hz, 2H), 6.81–6.72 (m, 2H), 6.08 (d, J = 6.5 Hz, 1H), 4.94 (dd, J = 11.3, 5.5 Hz, 1H), 3.90 (d, J = 7.1 Hz, 1H), 3.80 (d, J = 6.4 Hz, 1H), 3.74 (br s, 2H), 3.61 (q, J = 7.1 Hz, 1H), 2.90–2.80 (m, 1H), 2.78 – 2.60 (m, 0.5 H), 2.60–2.35 (m, 1H), 2.35–2.14 (m, 1.5 H), 2.07–1.91 (m, 1H), 1.82–1.67 (m, 1H), 1.50 (d, J = 7.1 Hz, 3H), 1.46–1.31 (m, 1H), 1.12 (d, J = 6.8 Hz, 1.4H), 1.04 (d, J = 6.1 Hz, 1.6H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 158.8, 158.8, 141.3, 130.7, 130.7, 128.9, 127.6, 127.5, 127.3, 114.8, 72.8, 72.2, 66.6, 55.3, 47.1, 33.2, 31.5, 30.6, 30.3, 27.0, 26.8, 22.3, 22.1, 18.4; HRMS (ESI) m/z calcd for C₂₃H₂₉NO₃ [M + H]⁺ 368.2220, m/z found 368.2213.

(2S)-N-[(1R)-1-{4-[(3,3-Dimethylcyclobutyl)methoxy]phenyl}-2-hydroxyethyl]-2-phenylpropanamide (26).

The procedure for the synthesis of 12 was followed starting with 11o to give 26 (75% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.44–7.20 (m, 5H), 7.06 (d, J = 8.6 Hz, 2H), 6.82 (d, J = 8.6 Hz, 2H), 5.95 (d, J = 6.7 Hz, 1H), 4.94 (dd, J = 11.4, 5.5 Hz, 1H), 3.86 (d, J = 6.6 Hz, 2H), 3.80–3.65 (m, 2H), 3.58 (q, J = 7.2 Hz, 1H), 2.75–2.44 (m, 2H), 1.98–1.76 (m, 2H), 1.67–1.57 (m, 2H), 1.52 (d, J = 7.2 Hz, 3H), 1.17 (s, 3H), 1.08 (s, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.8, 158.9, 141.2, 130.7, 129.0, 127.6, 127.5, 127.4, 114.9, 73.1, 66.7, 55.6, 47.2, 37.7, 31.9, 30.9, 28.9, 27.2, 18.5; HRMS (ESI) m/z calcd for C₂₄H₃₁NO₃ [M + H]⁺ 382.2377, m/z found 382.2368.

(2S)-N-[(1R)-2-Hydroxy-1-[4-({spiro[2.3]hexan-5-yl}methoxy)phenyl]ethyl]-2-phenylpropanamide (27).

The procedure for the synthesis of 12 was followed starting with 11p to give 27 (80% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.44–7.22 (m, 5H), 7.08 (d, J = 8.6 Hz, 2H), 6.86 (d, J = 8.6 Hz, 2H), 5.93 (d, J = 6.7 Hz, 1H), 4.96 (dd, J = 11.4, 5.5 Hz, 1H), 4.01 (d, J = 7.1 Hz, 2H), 3.86–3.67 (m, 2H), 3.60 (q, J = 7.1 Hz, 1H), 2.93–2.71 (m, 1H), 2.56 (t, J = 6.0 Hz, 1H), 2.32–2.11 (m, 2H), 2.04–1.81 (m, 2H), 1.54 (d, J = 7.2 Hz, 3H), 0.58–0.30 (m, 4H); ¹³C NMR (75 MHz, CDCl₃) δ 174.8, 159.0, 141.2, 130.7, 129.0, 127.7, 127.5, 127.4, 115.0, 72.6, 66.7, 55.7, 47.2, 33.4, 29.8, 18.5, 16.9, 12.1, 11.7; HRMS (ESI) m/z calcd for C₂₄H₂₉NO₃ [M + H]⁺ 380.2220, m/z found 380.2214.

(2S)-N-[(1R)-1-{4-[(3,3-Difluorocyclobutyl)methoxy]phenyl}-2-hydroxyethyl]-2-phenylpropanamide (28).

The procedure for the synthesis of 12 was followed starting with 11q to give 28 (50% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.41–7.22 (m, 5H), 6.97 (d, J = 8.6 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 6.00 (d, J = 7.0 Hz, 1H), 4.96 (dd, J = 11.8, 4.9 Hz, 1H), 3.94 (d, J = 5.9 Hz, 2H), 3.77 (d, J = 5.1 Hz, 2H), 3.63 (q, J = 7.1 Hz, 1H), 2.82–2.61 (m, 2H), 2.61–2.33 (m, 4H), 1.52 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.5, 158.2, 141.2, 131.4, 128.9, 127.6, 127.6, 127.3, 114.8, 70.1 (m), 55.2, 47.1, 37.7 (t, J = 23.0 Hz), 22.5 (dd, J = 12.0, 7.4 Hz), 18.4; ¹⁹F NMR (282 MHz, CDCl₃) δ −83.8 (d, J = 193 Hz), −93.2 (d, J = 196 Hz); HRMS (ESI) m/z calcd for C₂₂H₂₅F₂NO₃ [M + H]⁺ 390.1875, m/z found 390.1867.

(2R)-2-{4-[(2-Methylpentyl)oxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetic Acid (29a).²⁸

To a solution of 3 (1.2 g, 3.02 mmol) in THF: H₂O (10 mL, 1:1, v/v) was added 1 N NaOH (6.04 mL, 6.04 mmol) at room temperature. After stirring for 1 h, the reaction was cooled (ice bath) and acidified (pH = ~4) with 1 N HCl and extracted with EtOAc (3 x 20 mL). The combined EtOAc layers were washed with brine (3 x 20 mL), dried (Na₂SO₄), and concentrated under reduced pressure to furnish 29a (1.06 g, 92%) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.17 (m, 5H), 7.11 (d, J = 8.1 Hz, 2H), 6.79 (d, J = 8.2 Hz, 2H), 6.21 (d, J = 6.1 Hz, 1H), 5.43 (d, J = 6.3 Hz, 1H), 3.82–3.72 (m, 1H), 3.71–3.57 (m, 2H), 2.01–1.78 (m, 1H), 1.49 (d, J = 7.1 Hz, 3H), 1.45–1.07 (m, 4H), 0.99 (d, J = 6.6 Hz, 3H), 0.91 (t, J = 6.7 Hz, 3H); MS (ESI) m/z 384.4 [M + H]⁺.

(2R)-2-[4-(Cyclobutylmethoxy)phenyl]-2-[(2S)-2-phenylpropanamido]acetic Acid (29b).

The procedure for the synthesis of 29a was followed starting with 11h to give 29b (100% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.32–7.20 (m, 6H), 7.10 (d, J = 8.7 Hz, 2H), 6.79 (d, J = 8.7 Hz, 2H), 6.25 (d, J = 6.6 Hz, 1H), 5.43 (d, J = 6.7 Hz, 1H), 3.87 (d, J = 6.7 Hz, 2H), 3.63 (q, J = 7.1 Hz, 1H), 2.82–2.62 (m, 1H), 2.22–2.03 (m, 2H), 2.03–1.73 (m, 4H), 1.49 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.3, 174.1, 159.5, 140.7, 128.9, 128.3, 127.6, 127.4, 127.4, 114.9, 72.1, 56.1, 46.7, 34.5, 24.8, 18.5, 18.2; MS (ESI) m/z 368.0 [M + H]⁺.

(2S)-N-[(R)-Carbamoyl({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (30).

To a solution of 29a (230 mg, 0.6 mmol) in dioxane (10 mL) at room temperature were added pyridine (0.1 mL, 1.2 mmol), NH₄HCO₃ (95 mg, 1.2 mmol) and Boc anhydride (262 mg, 1.2 mmol). The reaction mixture was stirred for 4 h. Another portion of NH₄HCO₃ (95 mg, 1.2 mmol) and Boc anhydride (262 mg, 1.2 mmol) was added and the mixture was stirred overnight. The reaction was quenched by 10% citric acid (10 mL) and extracted with EtOAc (3 x 30 mL). The combined extracts were washed with 10% citric acid (10 mL), NaHCO₃ (2 x 10 mL), brine (10 mL), dried (NaSO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–3% MeOH in DCM to provide 30 (100 mg, 44% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.15 (m, 5H), 7.09 (d, J = 9.0 Hz, 2H), 6.96 (d, J = 9.0 Hz, 1H), 6.72 (d, J = 9.0 Hz, 2H), 6.61 (s, 1H), 5.85 (s, 1H), 5.54 (d, J = 6.0 Hz, 1H), 3.84–3.54 (m, 3H), 1.98–1.81 (m, 1H), 1.44 (d, J = 9.0 Hz, 3H), 1.41–1.08 (m, 4H), 0.98 (d, J = 9.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.7, 172.8, 159.2, 141.1, 129.4, 128.7, 128.1, 127.5, 127.1, 114.7, 73.2, 56.0, 46.6, 35.7, 32.8, 20.0, 18.3, 16.9, 14.3; HRMS (ESI) m/z calcd for C₂₃H₃₀N₂O₃ [M + H]⁺ 383.2329, m/z found 383.2332.

(2S)-N-[(R)-(Methylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (31).

To a solution of 29a (115 mg, 0.3 mmol) in DMF (5 mL) under nitrogen were added EDC hydrochloride (63 mg, 0.33 mmol), HOBt (45 mg, 0.33 mmol) and DIPEA (0.16 mL, 0.9 mmol). After cooling to 0 °C, methylamine hydrochloride (22 mg, 0.33 mmol) was added and the reaction mixture was stirred at room temperature overnight. The reaction was quenched with 10% citric acid solution (5 mL) and extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with saturated NaHCO₃ (10 mL) and brine (10 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–100% EtOAc in hexanes to furnish 31 (56 mg, 47% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.18 (m, 5H), 7.17–6.96 (m, 4H), 6.68 (d, J = 9.0 Hz, 2H), 5.62 (d, J = 9.0 Hz, 1H), 3.79–3.54 (m, 3H), 2.70 (d, J = 6.0 Hz, 2H), 1.99–1.81 (m, 1H), 1.55–1.12 (m, 5H), 1.48 (d, J = 9.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.6, 171.0, 159.0, 141.2, 129.9, 128.7, 128.0, 127.5, 127.1, 114.6, 73.1, 56.2, 46.6, 35.7, 32.8, 26.3, 20.0, 18.4, 16.9, 14.2; HRMS (ESI) m/z calcd for C₂₄H₃₂N₂O₃ [M + H]⁺ 397.2486, m/z found 397.2482.

(2S)-N-((1R)-2-(Ethylamino)-1-(4-((2-methylpentyl)oxy)phenyl)-2-oxoethyl)-2-phenylpropanamide (32).

The procedure for the synthesis of 31 was followed starting with 29a and ethylamine to give 32 (53% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.17 (m, 5H), 7.08 (d, J = 8.7 Hz, 2H), 6.77 (d, J = 8.7 Hz, 2H), 6.73 (d, J = 6.5 Hz, 1H), 5.64 (br s, 1H), 5.28 (d, J = 6.6 Hz, 1H), 3.76 (dd, J = 8.8, 5.8 Hz, 1H), 3.70–3.54 (m, 2H), 3.35–3.08 (m, 2H), 2.03–1.77 (m, 1H), 1.53–1.11 (m, 7H), 1.05 (t, J = 7.3 Hz, 3H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃): δ 173.5, 170.2, 159.0, 141.2, 130.0, 128.8, 128.1, 127.5, 127.1, 114.7, 73.2, 56.4, 46.7, 35.7, 34.6, 32.9, 20.0, 18.4, 17.0, 14.5, 14.3; HRMS (ESI) m/z calcd for C₂₅H₃₄N₂O₃ [M + H]⁺ 411.2642, m/z found 411.2639.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}(propylcarbamoyl)methyl]-2-phenylpropanamide (33).

The procedure for the synthesis of 31 was followed starting with 29a and propylamine hydrochloride to give 33 (39% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.17 (m, 5H), 7.06 (d, J = 9.0 Hz, 2H), 6.98 (d, J = 6.0 Hz, 1H), 6.70 (d, J = 9.0 Hz, 2H), 6.68–6.55 (m, 1H), 5.63 (d, J = 9.0 Hz, 1H), 3.78–3.55 (m, 3H), 3.23–3.03 (m, 2H), 1.98–1.81 (m, 1H), 1.55–1.10 (m, 6H), 1.48 (d, J = 9.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H), 0.79 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 170.3, 159.0, 141.2, 130.0, 128.7, 128.0, 127.5, 127.1, 114.6, 73.2, 56.4, 46.7, 41.4, 35.7, 32.8, 22.6, 20.0, 18.4, 16.9, 14.2, 11.2; HRMS (ESI) m/z calcd for C₂₆H₃₆N₂O₃ [M + H]⁺ 425.2799, m/z found 425.2798.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}[(propan-2-yl)carbamoyl]methyl]-2-phenylpropanamide (34).

The procedure for the synthesis of 31 was followed starting with 29a and isopropylamine to give 34 (41% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.16 (m, 5H), 7.03 (d, J = 9.0 Hz, 2H), 6.94 (d, J = 9.0 Hz, 1H), 6.68 (d, J = 9.0 Hz, 2H), 6.39 (d, J = 9.0 Hz, 1H), 5.49 (d, J = 6.0 Hz, 1H), 4.07–3.90 (m, 1H), 3.80–3.57 (m, 3H), 1.98–1.81 (m, 1H), 1.52–1.14 (m, 4H), 1.48 (d, J = 9.0 Hz, 3H), 1.12 (d, J = 6..0 Hz, 3H), 1.00 (d, J = 6.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 169.4, 158.9, 141.2, 130.0, 128.7, 128.0, 127.5, 127.1, 114.6, 73.2, 56.3, 46.7, 41.7, 35.7, 32.9, 22.5, 22.3, 20.0, 18.5, 16.9, 14.2; HRMS (ESI) m/z calcd for C₂₆H₃₆N₂O₃ [M + H]⁺ 425.2799, m/z found 425.2804.

(2S)-N-((1R)-2-(tert-Butylamino)-1-(4-((2-methylpentyl)oxy)phenyl)-2-oxoethyl)-2-phenylpropanamide (35).

To a solution of 29a (50 mg, 0.13 mmol) in THF (2 mL) was added 2-chloro-4,6-dimethoxy-1,3,5-triazine (22.9 mg, 0.13 mmol) and N-methylmorpholine (15 μL, 13.5 mmol). The reaction that resulted was stirred at room temperature. After 1 h tert-butylamine (21 μL, 0.2 mmol) was added to the above cloudy solution and the reaction that resulted was stirred overnight at room temperature. The solvent was evaporated, and the residue was subjected to column chromatography on silica gel using 0–50% EtOAc in hexanes to furnish 35 (28 mg, 49% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.13 (m, 5H), 7.08 (d, J = 8.7 Hz, 2H), 6.77 (d, J = 8.7 Hz, 2H), 6.70 (d, J = 6.6 Hz, 1H), 5.30 (br s, 1H), 5.19 (d, J = 6.7 Hz, 1H), 3.82–3.71 (m, 1H), 3.71–3.50 (m, 2H), 2.05–1.72 (m, 1H), 1.46 (d, J = 7.1 Hz, 3H), 1.44–1.13 (m, 13H), 1.00 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.1 Hz, 3H). ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 169.2, 159.1, 141.4, 130.3, 128.7, 128.2, 127.5, 127.0, 114.8, 73.2, 56.9, 51.7, 46.8, 35.8, 32.9, 28.6, 20.0, 18.5, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₇H₃₈N₂O₃ [M + H]⁺ 439.2955, m/z found 439.2957.

(2S)-N-[(R)-[(Butan-2-yl)carbamoyl]({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (36).

The procedure for the synthesis of 31 was followed starting with 29a and butan-2-amine to give 36 (42% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.16 (m, 5H), 7.07 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 1H), 6.72 (d, J = 9.0 Hz, 2H), 5.94–5.83 (m, 1H), 5.40 (d, J = 6.0 Hz, 1H), 3.90–3.55 (m, 4H), 1.99–1.83 (m, 1H), 1.53–1.12 (m, 6H), 1.48 (d, J = 6.0 Hz, 3H), 1.09 (d, J = 6..0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H), 0.84 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 169.6, 159.0, 141.2, 130.2, 128.8, 128.1, 127.5, 127.1, 114.7, 73.2, 56.6, 47.0, 46.8, 35.7, 32.9, 29.4, 20.4, 20.0, 18.5, 17.0, 14.3, 10.0; HRMS (ESI) m/z calcd for C₂₇H₃₈N₂O₃ [M + H]⁺ 439.2955, m/z found 439.2952.

(2S)-N-((1R)-2-((3-Methylbutan-2-yl)amino)-1-(4-((2-methylpentyl)oxy)phenyl)-2-oxoethyl)-2-phenylpropanamide (37).

The procedure for the synthesis of 31 was followed starting with 29a and 1,2-dimethylpropylamine to give 37 (44% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.16 (m, 5H), 7.08 (d, J = 8.6 Hz, 2H), 6.82 (d, J = 6.5 Hz, 1H), 6.76 (d, J = 8.7 Hz, 2H), 5.53 (d, J = 9.0 Hz, 1H), 5.30 (d, J = 6.5 Hz, 1H), 3.87–3.69 (m, 2H), 3.69–3.41 (m, 2H), 2.04–1.75 (m, 1H), 1.73–1.10 (m, 8H), 1.01 (dd, J = 13.6, 6.7 Hz, 6H), 0.91 (t, J = 7.0 Hz, 3H), 0.69–0.58 (m, 6H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 169.4, 159.1, 141.2, 130.3, 128.8, 128.1, 127.5, 127.1, 114.8, 73.3, 56.8, 50.4, 46.9, 35.7, 32.9, 32.9, 20.0, 18.4, 18.3, 18.0, 17.6, 16.9, 14.3; HRMS (ESI) m/z calcd for C₂₈H₄₀N₂O₃ [M + H]⁺ 453.3112, m/z found 453.3110.

(2S)-N-[(R)-(Cyclopropylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (38).

The procedure for the synthesis of 31 was followed starting with 29a and cyclopropylamine to give 38 (35% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.16 (m, 5H), 7.06–6.93 (m, 3H), 6.86 (d, J = 6.0 Hz, 1H), 6.88 (d, J = 9.0 Hz, 2H), 5.49 (d, J = 6.0 Hz, 1H), 3.79–3.56 (m, 3H), 2.68–2.57 (m, 1H), 1.98–1.85 (m, 1H), 1.51–1.10 (m, 4H), 1.48 (d, J = 6.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H), 0.76–0.57 (m, 2H), 0.52–0.29 (m, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 171.7, 159.0, 141.1, 129.7, 128.8, 128.0, 127.5, 127.1, 114.6, 73.2, 56.1, 46.7, 35.7, 32.9, 22.7, 20.0, 18.4, 17.0, 14.2, 6.4, 6.2; HRMS (ESI) m/z calcd for C₂₆H₃₄N₂O₃ [M + H]⁺ 423.2642, m/z found 423.2647.

(2S)-N-((1R)-2-((Cyclopropylmethyl)amino)-1-(4-((2-methylpentyl)oxy)phenyl)-2-oxoethyl)-2-phenylpropanamide (39).

The procedure for the synthesis of 31 was followed starting with 29a and cyclopropylmethylamine hydrochloride to give 39 (49% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.18 (m, 5H), 7.07 (d, J = 8.7 Hz, 2H), 6.93 (d, J = 7.2 Hz, 1H), 6.70 (d, J = 8.7 Hz, 2H), 6.57 (t, J = 5.4 Hz, 1H), 5.53 (d, J = 7.2 Hz, 1H), 3.82–3.52 (m, 3H), 3.15–2.92 (m, 2H), 2.04–1.78 (m, 1H), 1.48 (d, J = 7.1 Hz, 3H), 1.45–1.11 (m, 4H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H), 0.88–0.77 (m, 1H), 0.47–0.35 (m, 2H), 0.15–0.05 (m, 2H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 170.3, 159.1, 141.3, 130.0, 128.8, 128.1, 127.5, 127.1, 114.7, 73.2, 56.4, 46.8, 44.3, 35.7, 32.9, 20.0, 18.5, 17.0, 14.3, 10.5, 3.3, 3.2; HRMS (ESI) m/z calcd for C₂₇H₃₆N₂O₃ [M + H]⁺ 437.2799, m/z found 437.2793.

(2S)-N-[(R)-(Cyclobutylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (40).

The procedure for the synthesis of 31 was followed starting with 29a and cyclobutylamine to give 40 (35% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.17 (m, 5H), 7.04 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 6.0 Hz, 1H), 6.70 (d, J = 9.0 Hz, 2H), 6.59 (d, J = 9.0 Hz, 1H), 5.45 (d, J = 6.0 Hz, 1H), 4.36–4.20 (m, 1H), 3.80–3.56 (m, 3H), 2.35–2.13 (m, 2H), 1.97–1.57 (m, 5H), 1.54–1.10 (m, 4H), 1.49 (d, J = 6.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 169.3, 159.0, 141.2, 129.9, 128.8, 128.1, 127.5, 127.1, 114.7, 73.2, 58.3, 46.8, 45.0, 35.7, 32.9, 30.8, 30.7, 20.0, 18.4, 17.0, 15.0, 14.3; HRMS (ESI) m/z calcd for C₂₇H₃₆N₂O₃ [M + H]⁺ 437.2799, m/z found 437.2806.

(2S)-N-((1R)-2-((1-Methylcyclobutyl)amino)-1-(4-((2-methylpentyl)oxy)phenyl)-2-oxoethyl)-2-phenylpropanamide (41).

The procedure for the synthesis of 31 was followed starting with 29a and 1-methylcyclobutylamine to give 41 (48% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.13 (m, 5H), 7.07 (d, J = 8.7 Hz, 2H), 6.75 (d, J = 8.7 Hz, 2H), 5.89 (s, 1H), 5.29 (d, J = 6.8 Hz, 1H), 3.76 (dd, J = 8.9, 5.8 Hz, 1H), 3.71–3.54 (m, 2H), 2.26–2.03 (m, 2H), 2.03–1.85 (m, 3H), 1.83–1.67 (m, 2H), 1.54–1.10 (m, 10H), 1.00 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃): δ 173.2, 168.9, 159.1, 141.3, 130.2, 128.8, 128.2, 127.5, 127.1, 114.8, 73.2, 56.6, 54.5, 46.8, 35.7, 34.4, 34.3, 32.9, 25.0, 20.0, 18.5, 17.0, 14.5, 14.3; HRMS (ESI) m/z calcd for C₂₈H₃₈N₂O₃ [M + H]⁺ 451.2955, m/z found 451.2950.

(2S)-N-[(R)-(Cyclopentylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (42).

The procedure for the synthesis of 31 was followed starting with 29a and cyclopentylamine to give 42 (33% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.15 (m, 5H), 7.06 (d, J = 8.6 Hz, 2H), 6.87–6.64 (m, 3H), 5.90 (d, J = 7.3 Hz, 1H), 5.35 (d, J = 6.9 Hz, 1H), 4.28–4.02 (m, 1H), 3.75 (dd, J = 8.9, 5.8 Hz, 1H), 3.70–3.55 (m, 2H), 2.04–1.76 (m, 3H), 1.66–1.28 (m, 11H), 1.27–1.10 (m, 2H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 169.7, 159.1, 141.2, 130.0, 128.8, 128.1, 127.5, 127.1, 114.8, 73.2, 56.5, 51.5, 46.8, 35.7, 32.9, 32.8, 32.8, 23.7, 23.6, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₈H₃₈N₂O₃ [M + H]⁺ 451.2955, m/z found 451.2952.

(2S)-N-[(R)-(Cyclohexylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (43).

The procedure for the synthesis of 31 was followed starting with 29a and cyclohexylamine to give 43 (54% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.15 (m, 5H), 7.06 (d, J = 8.7 Hz, 2H), 6.85–6.64 (m, 3H), 5.75 (d, J = 7.6 Hz, 1H), 5.33 (d, J = 6.9 Hz, 1H), 3.83–3.53 (m, 4H), 2.01–1.77 (m, 2H), 1.76–1.51 (m, 5H), 1.48 (d, J = 7.1 Hz, 3H), 1.46–1.03 (m, 8H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 169.2, 159.1, 141.2, 130.1, 128.8, 128.2, 127.5, 127.1, 114.8, 73.2, 56.6, 48.6, 46.9, 35.7, 32.9, 32.8, 32.6, 25.4, 24.7, 24.6, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₉H₄₀N₂O₃ [M + H]⁺ 465.3122, m/z found 465.3111.

Methyl 2-[(2R)-2-{4-[(2-Methylpentyl)oxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetamido]acetate (44).

The procedure for the synthesis of 31 was followed starting with 29a and methyl 2-aminoacetate hydrochloride to give 44 (46% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.16 (m, 5H), 7.10 (d, J = 9.0 Hz, 2H), 6.88 (t, J = 4.5 Hz, 1H), 6.82–6.67 (m, 3H), 5.54 (d, J = 6.0 Hz, 1H), 3.93 (t, J = 6.0 Hz, 2H), 3.79–3.58 (m, 6H), 1.97–1.85 (m, 1H), 1.54–1.13 (m, 4H), 1.47 (d, J = 9.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 170.6, 169.8, 159.2, 141.1, 129.3, 128.8, 128.2, 127.5, 127.1, 114.7, 73.2, 56.4, 52.3, 46.7, 41.3, 35.7, 32.8, 20.0, 18.4, 16.9, 14.3; HRMS (ESI) m/z calcd for C₂₆H₃₄N₂O₅ [M + H]⁺ 455.2540, m/z found 455.2541.

(2S)-N-[(R)-[(2-Hydroxyethyl)carbamoyl]({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (45).

To a suspension of NaBH₄ (11 mg, 0.29 mmol) in EtOH (2 mL) at 0 °C under nitrogen was added LiCl (12 mg, 0.29 mmol). After stirring at 0 °C for 10 min, a solution of 44 (50 mg, 0.11 mmol) in THF (2 mL) was added. The reaction mixture was stirred at room temperature for 3 h and quenched with saturated NH₄Cl solution (5 mL), followed by addition of H₂O (10 mL). The mixture was extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (20 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 20–70% EtOAc in hexanes afforded 45 (38 mg, 81% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.28–7.08 (m, 5H), 7.07–6.88 (m, 3H), 6.76 (d, J = 6.0 Hz, 1H), 6.65 (d, J = 9.0 Hz, 2H), 5.39 (d, J = 6.0 Hz, 1H), 3.71–3.44 (m, 4H), 3.44–3.26 (m, 1H), 3.20–3.04 (m, 1H), 1.92–1.74 (m, 1H), 1.46–1.01 (m, 6H), 1.40 (d, J = 9.0 Hz, 3H), 0.92 (d, J = 9.0 Hz, 3H), 0.84 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.1, 171.2, 159.2, 141.0, 129.2, 128.8, 128.1, 127.5, 127.2, 114.8, 73.2, 61.6, 56.7, 46.7, 42.5, 35.7, 32.9, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₅H₃₄N₂O₄ [M + H]⁺ 427.2591, m/z found 427.2589.

(2S)-N-[(R)-[(2-Methoxyethyl)carbamoyl]({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (46)

The procedure for the synthesis of 31 was followed starting with 29a and 2-methoxyethan-1-amine to give 46 (42% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.29–7.09 (m, 5H), 7.00 (d, J = 9.0 Hz, 2H), 6.74 (d, J = 6.0 Hz, 1H), 6.66 (d, J = 9.0 Hz, 2H), 6.37 (br s, 1H), 5.35 (d, J = 6.0 Hz, 1H), 3.73–3.62 (m, 1H), 3.61–3.50 (m, 2H), 3.42–3.18 (m, 4H), 3.17 (s, 3H), 1.91–1.74 (m, 1H), 1.49–1.05 (m, 4H), 1.40 (d, J = 6.0 Hz, 3H), 0.91 (d, J = 6.0 Hz, 3H), 0.84 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 170.3, 159.1, 141.2, 129.8, 128.7, 128.1, 127.5, 127.1, 114.7, 73.2, 70.8, 58.7, 56.5, 46.7, 39.4, 35.7, 32.8, 20.0, 18.4, 16.9, 14.2; HRMS (ESI) m/z calcd for C₂₆H₃₆N₂O₄ [M + H]⁺ 441.2748, m/z found 441.2743.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}({[(oxolan-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (47).

The procedure for the synthesis of 31 was followed starting with 29a and tetrahydrofurfurylamine to give 47 (88% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.18 (m, 5H), 7.08 (d, J = 8.6 Hz, 2H), 6.82–6.66 (m, 3H), 6.19–6.03 (m, 1H), 5.35 (t, J = 7.2 Hz, 1H), 3.95–3.49 (m, 6H), 3.50–3.32 (m, 1H), 3.32–3.06 (m, 1H), 2.01–1.64 (m, 3H), 1.64–1.08 (m, 8H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 173.2, 170.4, 170.3, 159.2, 159.2, 141.3, 141.2, 130.2, 129.8, 128.8, 128.2, 128.1, 127.6, 127.1, 114.8, 77.3, 73.3, 73.2, 68.1, 68.1, 56.8, 56.7, 46.8, 43.2, 42.9, 35.7, 32.9, 28.5, 28.2, 25.8, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₈H₃₈N₂O₄ [M + H]⁺ 467.2904, m/z found 467.2905.

tert-Butyl N-{2-[(2R)-2-{4-[(2-Methylpentyl)oxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetamido] ethyl}carbamate (48).

The procedure for the synthesis of 31 was followed starting with 29a and N-Boc-ethylenediamine hydrochloride to give 48 (55% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.17 (m, 5H), 7.06 (d, J = 8.7 Hz, 2H), 6.84–6.65 (m, 4H), 5.35 (d, J = 6.7 Hz, 1H), 4.92 (d, J = 5.5 Hz, 1H), 3.74 (dd, J = 8.9, 5.8 Hz, 1H), 3.69–3.55 (m, 2H), 3.40–3.04 (m, 4H), 2.05–1.73 (m, 1H), 1.47 (d, J = 7.1 Hz, 3H), 1.45–1.11 (m, 13H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 170.8, 159.2, 141.2, 129.6, 128.8, 128.1, 127.5, 127.2, 114.8, 79.7, 73.2, 56.8, 46.8, 40.8, 40.0, 35.7, 32.9, 28.3, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₃₀H₄₃N₃O₅ [M + H]⁺ 526.3275, m/z found 526.3267.

(2S)-N-[(R)-[(2-Aminoethyl)carbamoyl]({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide Hydrochloride (49).

To a solution of 48 (14.1 mg, 0.03 mmol) in DCM, HCl (4 M in dioxane, 67 μ, 0.27 mmol) was added at 0 °C. The reaction that resulted was warmed to room temperature and stirred until completion. After the completion of the reaction, the solvent was evaporated, and the residue was re-dissolved in DCM and evaporated under reduced pressure (3 times). The residue was dried in vacuo to furnish 49 as hydrochloride salt (9.7 mg, 78%) as a thick colorless oil. ¹H NMR (300 MHz, CD₃OD) δ 7.36–7.15 (m, 7H), 6.86 (d, J = 8.7 Hz, 2H), 5.11 (s, 1H), 3.84–3.53 (m, 4H), 3.37–3.22 (m, 1H), 3.19–2.94 (m, 2H), 2.00–1.76 (m, 1H), 1.57–1.14 (m, 7H), 1.00 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 177.3, 174.4, 161.0, 142.8, 130.2, 129.5, 129.1, 128.5, 128.0, 115.8, 74.3, 59.4, 48.4, 41.0, 38.3, 36.9, 34.1, 21.1, 18.9, 17.3, 14.6; HRMS (ESI) free base m/z calcd for C₂₅H₃₅N₃O₃ [M + H]⁺ 426.2751, m/z found 426.2750.

tert-Butyl N-Methyl-N-{2-[(2R)-2-{4-[(2-methylpentyl)oxy]phenyl}-2-[(2S)-2-phenylpropanamido]acetamido]ethyl}carbamate (50).

The procedure for the synthesis of 31 was followed starting with 29a and N-Boc-N-methylethylenediamine to give 50 (32% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.15 (m, 5H), 7.06 (d, J = 8.6 Hz, 2H), 6.97 (br s, 1H), 6.84 (d, J = 6.7 Hz, 1H), 6.72 (d, J = 8.6 Hz, 2H), 5.62–4.88 (m, 1H), 3.80–3.57 (m, 3H), 3.27 (br s, 4H), 2.74 (br s, 3H), 2.01–1.78 (m, 1H), 1.46 (d, J = 7.2 Hz, 3H), 1.44–1.10 (m, 13H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 170.6, 159.1, 156.8, 141.3, 129.9, 128.8, 128.1, 127.5, 127.1, 114.7, 79.9, 73.2, 56.6, 47.6, 46.7, 39.1, 35.7, 35.0, 32.9, 28.4, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₃₁H₄₅N₃O₅ [M + H]⁺ 540.3432, m/z found 540.3424.

(2S)-N-[(R)-{[2-(Methylamino)ethyl]carbamoyl}({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide Hydrochloride (51).

The procedure for the synthesis of 49 was followed starting with 50 to give 51 as hydrochloride salt (94%) as a thick colorless oil. ¹H NMR (300 MHz, CD₃OD) δ 8.62–8.38 (m, 1H), 7.37–7.12 (m, 7H), 6.85 (d, J = 8.7 Hz, 2H), 5.17–5.06 (m, 1H), 3.89–3.61 (m, 4H), 3.40–3.32 (m, 1H), 3.26–3.03 (m, 2H), 2.73 (s, 3H), 2.05–1.75 (m, 1H), 1.57–1.13 (m, 7H), 0.99 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 179.9, 176.9, 163.6, 145.3, 132.7, 132.1, 131.6, 131.0, 130.5, 118.3, 76.9, 62.0, 53.0, 49.3, 39.6, 39.4, 36.6, 36.4, 23.6, 21.4, 19.8, 17.2; HRMS (ESI) free base m/z calcd for C₂₆H₃₇N₃O₃ [M + H]⁺ 440.2908, m/z found 440.2923.

(2S)-N-[(R)-{[2-(Dimethylamino)ethyl]carbamoyl}({4- [(2-methylpentyl)oxy]phenyl})methyl]-2 -phenylpropanamide (52).

A solution of 29a (77 mg, 0.20 mmol) in DCM (5 mL) was cooled to 0 °C. To that above solution oxalyl chloride (0.4 mmol, 34 μL) and DMF (catalytic) was added sequentially. The reaction that resulted was stirred at room temperature for 2 h. At that time, the solvent was removed, and the residue was dried in vacuo for 1 h. The yellow color residue was re-dissolved in DCM (5 mL) and DIPEA (350 μL, 2.0 mmol) and N, N-dimethylethylenediamine (88.5 mg, 1.0 mmol) was added. The reaction that resulted was stirred at room temperature for 14 h. The solvent was evaporated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–50% CMA80 in DCM to furnish the amide 52 (37 mg, 41%) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.08 (m, 6H), 6.95 (d, J = 6.7 Hz, 1H), 6.79 (dd, J = 18.1, 8.7 Hz, 2H), 6.69 (s, 1H), 5.33 (d, J = 6.7 Hz, 1H), 3.82–3.52 (m, 3H), 3.47–3.28 (m, 1H), 3.28–2.97 (m, 1H), 2.56–2.32 (m, 2H), 2.32–2.16 (m, 6H), 2.03–1.78 (m, 1H), 1.62–1.09 (m, 8H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.1, 170.0, 158.8, 140.7, 129.5, 128.3, 128.0, 127.3, 126.7, 114.4, 72.8, 57.0, 56.4, 46.5, 44.2, 36.0, 35.3, 32.4, 19.6, 18.1, 16.5, 13.8; HRMS (ESI) m/z calcd for C₂₇H₃₉N₃O₃ [M + H]⁺ 454.3064, m/z found 454.3059.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}({[(pyrrolidin-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (53).

The procedure for the synthesis of 31 was followed starting with 29a and 1-Boc-2-(aminomethyl)pyrrolidine to give N-Boc-53 (68% yield). The N-Boc compound (100.0 mg, 0.18 mmol) was dissolved in DCM (2 mL) and cooled to 0 °C. To that above solution HCl (4 M in dioxane, 440 μL, 1.77 mmol) was added and the reaction that resulted was stirred at room temperature until complete conversion. After that the reaction was diluted with DCM (10 mL) and basified with a saturated NaHCO₃ (10 mL). The organic layer was separated, and the aqueous layer was extracted with additional DCM (2 x 10 mL). The combined organic layers were washed with brine, dried (K₂CO₃), and evaporated to furnish the free base 53 (72 mg, 87%) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.18 (m, 5H), 7.08 (d, J = 8.6 Hz, 2H), 6.96 (dd, J = 6.8, 2.3 Hz, 1H), 6.90–6.81 (m, 1H), 6.72 (d, J = 8.6 Hz, 2H), 5.42 (dd, J = 6.9, 2.0 Hz, 1H), 3.82–3.53 (m, 3H), 3.37–2.90 (m, 3H), 2.90–2.57 (m, 2H), 2.32 (br s, 1H), 2.07–1.82 (m, 1H), 1.79–1.06 (m, 11H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 173.4, 170.5, 170.4, 159.1, 141.3, 130.1, 130.1, 128.7, 128.1, 127.5, 127.1, 114.7, 73.2, 57.4, 57.4, 56.6, 46.7, 46.4, 46.3, 44.0, 43.8, 35.7, 32.9, 29.0, 28.8, 25.6, 25.6, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₈H₃₉N₃O₃ [M + H]⁺ 466.3064, m/z found 464.3077.

(2S)-N-[(R)-{[(Azetidin-3-yl)methyl]carbamoyl}({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (54).

The procedure for the synthesis of 53 was followed starting with 29a and 1-Boc-3-(aminomethyl)azetidine to furnish N-Boc-54 (88% yield) as a colorless oil. The amide, N-Boc-54 (75 mg, 0.13 mmol) was dissolved in DCM (6 mL) and cooled to 0 °C. To that above solution, TFA (0.4 mL) was added dropwise with a syringe and the reaction, which resulted, was stirred at 0 °C for 30 min. At that time, the reaction was diluted with DCM (10 mL) and basified with saturated aqueous NaHCO₃ (15 mL). The organic layer was separated, and the aqueous layer was extracted with additional DCM (2 x 5 mL). The combined organic layers were dried with K₂CO₃ and the residue was subjected to column chromatography on silica gel using 0–40% CMA80/DCM to furnish 54 (38 mg, 62%) as a free base. ¹H NMR (300 MHz, CDCl₃) δ 9.68–8.74 (br s, 2H), 8.53 (s, 1H), 7.40–7.08 (m, 6H), 6.95 (d, J = 6.6 Hz, 1H), 6.76 (dd, J = 26.9, 8.7 Hz, 2H), 5.59 (d, J = 6.6 Hz, 1H), 4.22–3.82 (m, 3H), 3.77–3.47 (m, 4H), 3.27–3.04 (m, 1H), 3.03–2.84 (m, 1H), 2.72–2.24 (m, 1H), 1.96–1.78 (m, 1H), 1.59–1.05 (m, 7H), 0.96 (d, J = 6.7 Hz, 3H), 0.90 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 174.6, 171.8, 159.3, 140.9, 128.8, 128.1, 127.5, 127.3, 114.8, 73.2, 57.0, 48.7, 46.5, 40.1, 35.7, 32.8, 32.0, 20.0, 18.2, 16.9, 14.3; HRMS (ESI) m/z calcd for C₂₇H₃₇N₃O₃ [M + H]⁺ 452.2908, m/z found 452.2908.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}(phenylcarbamoyl)methyl]-2-phenylpropanamide (55).

The procedure for the synthesis of 35 was followed starting with 29a and aniline to give 55 (65% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.22 (br s, 1H), 7.41 (d, J = 7.9 Hz, 2H), 7.37–7.19 (m, 7H), 7.15 (d, J = 8.7 Hz, 2H), 7.06 (t, J = 7.4 Hz, 1H), 6.81 (d, J = 7.2 Hz, 1H), 6.75 (d, J = 8.7 Hz, 2H), 5.67 (d, J = 7.0 Hz, 1H), 3.83–3.52 (m, 3H), 2.01–1.78 (m, 1H), 1.51 (d, J = 7.1 Hz, 3H), 1.47–1.07 (m, 4H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.9, 168.5, 159.4, 141.0, 137.4, 129.0, 128.9, 128.3, 127.6, 127.3, 124.5, 120.0, 115.0, 73.3, 57.3, 46.9, 35.7, 32.8, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₉H₃₄N₂O₃ [M + H]⁺ 459.2642, m/z found 459.2642.

(2S)-N-[(R)-(Benzylcarbamoyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (56).

The procedure for the synthesis of 31 was followed starting with 29a and benzylamine to give 56 (45% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.37–7.15 (m, 8H), 7.09 (d, J = 8.8 Hz, 4H), 6.80–6.68 (m, 3H), 6.12 (t, J = 5.6 Hz, 1H), 5.39 (d, J = 6.7 Hz, 1H), 4.51–4.13 (m, 2H), 3.88–3.44 (m, 3H), 2.06–1.63 (m, 1H), 1.60–1.08 (m, 7H), 0.99 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 170.2, 159.3, 141.2, 137.6, 129.7, 128.8, 128.6, 128.3, 127.5, 127.4, 127.1, 114.9, 73.3, 56.8, 46.8, 43.7, 35.7, 32.9, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₃₀H₃₆N₂O₃ [M + H]⁺ 473.2799, m/z found 473.2797.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}({[(pyridin-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (57).

The procedure for the synthesis of 31 was followed starting with 29a and 2-picolylamine to give 57 (75% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.47 (d, J = 4.3 Hz, 1H), 7.61 (td, J = 7.7, 1.8 Hz, 1H), 7.36–7.19 (m, 5H), 7.19–7.06 (m, 4H), 6.93 (d, J = 4.4 Hz, 1H), 6.78 (d, J = 8.7 Hz, 2H), 6.71 (d, J = 6.6 Hz, 1H), 5.43 (d, J = 6.6 Hz, 1H), 4.63–4.36 (m, 2H), 3.79–3.71 (m, 1H), 3.70–3.48 (m, 2H), 1.94–1.82 (m, 1H), 1.47 (d, J = 7.1 Hz, 3H), 1.45–1.09 (m, 4H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 170.2, 159.2, 155.7, 149.0, 141.2, 136.7, 129.8, 128.8, 128.2, 127.6, 127.1, 122.4, 121.7, 114.9, 73.2, 56.8, 46.9, 44.5, 35.7, 32.9, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₉H₃₅N₃O₃ [M + H]⁺ 474.2751, m/z found 474.2749.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}({[(pyridin-3-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (58).

The procedure for the synthesis of 31 was followed starting with 29a and 3-picolylamine to give 58 (71 % yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.46 (dd, J = 4.8, 1.5 Hz, 1H), 8.39 (d, J = 1.8 Hz, 1H), 7.45–7.38 (m, 1H), 7.34–7.22 (m, 3H), 7.22–7.10 (m, 3H), 7.05 (d, J = 8.7 Hz, 2H), 7.02–6.94 (m, 1H), 6.79–6.68 (m, 3H), 5.54 (d, J = 7.0 Hz, 1H), 4.48–4.19 (m, 2H), 3.74 (dd, J = 8.9, 5.8 Hz, 1H), 3.69–3.53 (m, 2H), 1.98–1.84 (m, 1H), 1.58–1.07 (m, 7H), 0.99 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.6, 170.6, 159.3, 148.9, 148.8, 141.0, 135.1, 133.5, 129.3, 128.8, 128.1, 127.5, 127.2, 123.4, 114.8, 73.3, 56.6, 46.8, 41.1, 35.7, 32.9, 20.0, 18.3, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₉H₃₅N₃O₃ [M + H]⁺ 474.2751, m/z found 474.2744.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}({[(pyridin-4-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (59).

The procedure for the synthesis of 31 was followed starting with 29a and 4-picolylamine to give 59 (74% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.43 (d, J = 5.8 Hz, 2H), 7.39–7.20 (m, 4H), 7.19–7.13 (m, 2H), 7.08 (d, J = 8.6 Hz, 2H), 6.95 (d, J = 5.7 Hz, 2H), 6.85 (d, J = 7.1 Hz, 1H), 6.73 (d, J = 8.5 Hz, 2H), 5.66 (d, J = 7.2 Hz, 1H), 4.33 (ddd, J = 48.7, 16.1, 6.0 Hz, 2H), 3.81–3.70 (m, 1H), 3.70–3.50 (m, 2H), 2.09–1.72 (m, 1H), 1.57–1.08 (m, 7H), 1.00 (d, J = 6.7 Hz, 3H), 0.90 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.8, 170.8, 159.4, 149.9, 147.0, 140.9, 129.3, 128.9, 128.1, 127.5, 127.3, 121.9, 114.8, 73.3, 56.5, 46.7, 42.3, 35.7, 32.9, 20.0, 18.3, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₉H₃₅N₃O₃ [M + H]⁺ 474.2751, m/z found 474.2750.

(2S)-N-[(R)-{[(Furan-2-yl)methyl]carbamoyl}({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (60).

The procedure for the synthesis of 31 was followed starting with 29a and furfurylamine to give 60 (75% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.41–7.12 (m, 6H), 7.06 (d, J = 8.6 Hz, 2H), 6.74 (d, J = 8.6 Hz, 3H), 6.46–6.15 (m, 2H), 6.09 (d, J = 3.1 Hz, 1H), 5.43 (d, J = 6.9 Hz, 1H), 4.56–4.20 (m, 2H), 3.86–3.49 (m, 3H), 1.99–1.81 (m, 1H), 1.54–1.09 (m, 7H), 0.99 (d, J = 6.7 Hz, 3H), 0.90 (t, J = 7.0, 3H); ¹³C NMR (75 MHz, CDCl₃) δ173.4, 170.1, 159.2, 150.7, 142.2, 141.1, 129.5, 128.8, 128.3, 127.5, 127.2, 114.8, 110.4, 107.4, 73.2, 56.6, 46.8, 36.8, 35.7, 32.9, 20.0, 18.4, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₈H₃₄N₂O₄ [M + H]⁺ 463.2591, m/z found 463.2585.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(furan-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (61).

The procedure for the synthesis of 31 was followed starting with 29b and furfurylamine to give 61 (73% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.44–7.12 (m, 6H), 7.04 (d, J = 8.7 Hz, 2H), 6.82 (d, J = 7.1 Hz, 1H), 6.76 (t, J = 5.6 Hz, 1H), 6.70 (d, J = 8.7 Hz, 2H), 6.25 (dd, J = 3.1, 1.9 Hz, 1H), 6.08 (d, J = 2.7 Hz, 1H), 5.54 (d, J = 7.1 Hz, 1H), 4.35 (qd, J = 15.6, 5.6 Hz, 2H), 3.85 (d, J = 6.7 Hz, 2H), 3.61 (q, J = 7.1 Hz, 1H), 2.82–2.64 (m, 1H), 2.20–2.04 (m, 2H), 2.02–1.66 (m, 4H), 1.43 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 170.2, 159.1, 150.8, 142.1, 141.1, 129.7, 128.8, 128.2, 127.5, 127.2, 114.8, 110.4, 107.3, 72.1, 56.4, 46.8, 36.7, 34.6, 24.8, 18.5, 18.3; HRMS (ESI) m/z calcd for C₂₇H₃₀N₂O₄ [M + H]⁺ 447.2278, m/z found 447.2277.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(thiophen-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (62).

The procedure for the synthesis of 31 was followed starting with 29b and 2-thiophenemethylamine to give 62 (80% yield) as a white waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.35–7.23 (m, 3H), 7.23–7.14 (m, 3H), 7.07 (d, J = 8.7 Hz, 2H), 6.89 (dd, J = 5.1, 3.5 Hz, 1H), 6.86–6.80 (m, 1H), 6.76 (d, J = 8.7 Hz, 2H), 6.70 (d, J = 6.8 Hz, 1H), 6.26–6.17 (m, 1H), 5.38 (d, J = 6.7 Hz, 1H), 4.67–4.36 (m, 2H), 3.87 (d, J = 6.7 Hz, 2H), 3.61 (q, J = 7.1 Hz, 1H), 2.88–2.52 (m, 1H), 2.20–2.04 (m, 2H), 2.03–1.73 (m, 4H), 1.45 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 169.9, 159.2, 141.1, 140.2, 129.5, 128.8, 128.3, 127.5, 127.2, 126.8, 125.9, 125.2, 114.9, 72.1, 56.7, 46.8, 38.6, 34.6, 24.8, 18.5, 18.4; HRMS (ESI) m/z calcd for C₂₇H₃₀N₂O₃S [M + H]⁺ 463.2050, m/z found 463.2046.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(1H-pyrrol-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (63).

The procedure for the synthesis of 31 was followed starting with 29b and (1H-pyrrol-2-yl)methanamine to give 63 (58% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 8.87 (br s, 1H), 7.45–7.15 (m, 5H), 7.11 (t, J = 5.7 Hz, 1H), 7.01 (d, J = 8.7 Hz, 2H), 6.79–6.55 (m, 4H), 6.04 (dd, J = 5.8, 2.8 Hz, 1H), 5.97–5.92 (m, 1H), 5.45 (d, J = 7.1 Hz, 1H), 4.33 (dd, J = 15.1, 6.0 Hz, 1H), 4.21–4.08 (m, 1H), 3.84 (d, J = 6.7 Hz, 2H), 3.60 (q, J = 7.1 Hz, 1H), 2.81–2.61 (m, 1H), 2.20–2.04 (m, 2H), 2.00–1.73 (m, 4H), 1.44 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.8, 171.3, 159.2, 141.0, 129.3, 128.9, 128.5, 128.1, 127.5, 127.3, 118.2, 114.9, 107.7, 106.8, 72.1, 56.6, 46.7, 37.1, 34.6, 24.8, 18.5, 18.4, HRMS (ESI) m/z calcd for C₂₇H₃₁N₃O₃ [M + H]⁺ 446.2438, m/z found 446.2437.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(5-methyl-1,3-oxazol-2-yl)methyl]carbamoyl}) methyl]-2-phenylpropanamide (64).

The procedure for the synthesis of 31 was followed starting with 29b and (5-methyloxazol-2-yl)methanamine to give 64 (46% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.36–7.15 (m, 5H), 7.09 (d, J = 8.7 Hz, 2H), 6.87 (t, J = 5.4 Hz, 1H), 6.81–6.67 (m, 3H), 6.59 (d, J = 1.2 Hz, 1H), 5.50 (d, J = 7.0 Hz, 1H), 4.51 (dd, J = 16.4, 5.8 Hz, 1H), 4.32 (dd, J = 16.4, 5.1 Hz, 1H), 3.86 (d, J = 6.7 Hz, 2H), 3.62 (q, J = 7.1 Hz, 1H), 2.84–2.52 (m, 1H), 2.24 (d, J = 1.1 Hz, 3H), 2.19–2.03 (m, 2H), 2.03–1.70 (m, 4H), 1.45 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 170.4, 159.2, 158.8, 149.4, 141.2, 129.5, 128.8, 128.3, 127.6, 127.2, 122.7, 114.8, 72.1, 56.5, 46.8, 37.0, 34.6, 24.8, 18.5, 18.4, 10.7; HRMS (ESI) m/z calcd for C₂₇H₃₁N₃O₄ [M + H]⁺ 462.2387, m/z found 462.2385.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(1,2-oxazol-3-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (65).

The procedure for the synthesis of 31 was followed starting with 29b and isoxazol-3-ylmethanamine hydrochloride to give 65 (60% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.25 (d, J = 1.6 Hz, 1H), 7.50–7.36 (m, 1H), 7.35–7.12 (m, 5H), 7.04 (d, J = 8.7 Hz, 2H), 6.84 (d, J = 7.2 Hz, 1H), 6.69 (d, J = 8.7 Hz, 2H), 6.13 (d, J = 1.6 Hz, 1H), 5.60 (d, J = 7.2 Hz, 1H), 4.51–4.31 (m, 2H), 3.84 (d, J = 6.7 Hz, 2H), 3.64 (q, J = 7.1 Hz, 1H), 2.84–2.60 (m, 1H), 2.23–2.02 (m, 2H), 2.02–1.69 (m, 4H), 1.43 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.8, 170.8, 160.0, 159.2, 158.7, 141.0, 129.3, 128.8, 128.1, 127.5, 127.2, 114.8, 103.6, 72.1, 56.4, 46.7, 35.2, 34.6, 24.8, 18.5, 18.3; HRMS (ESI) m/z calcd for C₂₆H₂₉N₃O₄ [M + H]⁺ 448.2231, m/z found 448.2229.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(1,3-thiazol-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (66).

The procedure for the synthesis of 31 was followed starting with 29b and 1,3-thiazol-2-ylmethylamine hydrochloride to give 66 (65% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 7.63 (d, J = 3.3 Hz, 1H), 7.49–7.14 (m, 6H), 7.07 (d, J = 8.7 Hz, 2H), 6.81 (d, J = 7.1 Hz, 1H), 6.72 (d, J = 8.7 Hz, 2H), 5.58 (d, J = 7.1 Hz, 1H), 4.66 (d, J = 5.9 Hz, 2H), 3.85 (d, J = 6.7 Hz, 2H), 3.62 (q, J = 7.1 Hz, 1H), 2.95–2.50 (m, 1H), 2.23–2.03 (m, 2H), 2.03–1.64 (m, 4H), 1.43 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 170.5, 167.0, 159.2, 142.4, 141.1, 129.3, 128.8, 128.3, 127.5, 127.2, 126.2, 119.5, 114.9, 72.1, 56.5, 46.7, 41.2, 34.6, 24.8, 18.5, 18.4; HRMS (ESI) m/z calcd for C₂₆H₂₉N₃O₃S [M + H]⁺464.2002, m/z found 448.2000.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(1H-imidazol-2-yl)methyl]carbamoyl}) methyl]-2-phenylpropanamide (67).

The procedure for the synthesis of 31 was followed starting with 29b and (1H-imidazol-2-yl)methanamine hydrochloride to give 67 (53% yield) as a waxy solid. ¹H NMR (300 MHz, CDCl₃) δ 8.35 (t, J = 5.7 Hz, 1H), 7.38–7.15 (m, 6H), 7.02 (d, J = 8.7 Hz, 2H), 6.89–6.82 (m, 3H), 6.70 (d, J = 8.7 Hz, 2H), 5.41 (d, J = 6.8 Hz, 1H), 4.42 (dd, J = 15.3, 6.3 Hz, 1H), 4.22 (dd, J = 15.3, 5.5 Hz, 1H), 3.83 (d, J = 6.7 Hz, 2H), 3.66 (q, J = 7.1 Hz, 1H), 2.81–2.52 (m, 1H), 2.18–1.66 (m, 6H), 1.46 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 229.0, 174.1, 171.9, 159.3, 145.2, 141.1, 128.8, 128.2, 127.5, 127.2, 122.0, 114.9, 72.1, 57.0, 46.6, 37.4, 34.6, 24.8, 18.5, 18.4; HRMS (ESI) m/z calcd for C₂₆H₃₀N₄O₃ [M + H]⁺ 447.2391, m/z found 447.2387.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(5-methyl-1,3,4-oxadiazol-2-yl)methyl]carbamoyl})methyl]-2-phenylpropanamide (68).

The procedure for the synthesis of 31 was followed starting with 29b and (5-methyl-1,3,4-oxadiazol-2-yl)methanamine oxalate to give 68 (55% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.16 (m, 5H), 7.10 (d, J = 8.7 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 6.59 (d, J = 6.4 Hz, 2H), 5.44 (d, J = 6.7 Hz, 1H), 4.57 (ddd, J = 38.1, 16.6, 5.8 Hz, 2H), 3.87 (d, J = 6.6 Hz, 2H), 3.63 (q, J = 7.1 Hz, 1H), 2.81–2.65 (m, 1H), 2.48 (s, 3H), 2.21–2.02 (m, 2H), 2.02–1.73 (m, 4H), 1.47 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 229.0, 173.6, 170.6, 164.4, 163.0, 159.4, 141.0, 128.9, 128.4, 127.5, 127.2, 115.0, 77.2, 72.1, 56.7, 46.8, 34.6, 34.5, 24.8, 18.5, 18.4, 10.9; HRMS (ESI) m/z calcd for C₂₆H₃₀N₄O₄ [M + H]⁺ 463.2340, m/z found 463.2337.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl]({[(1H-1,2,4-triazol-3-yl)methyl]carbamoyl}) methyl]-2-phenylpropanamide (69).

The procedure for the synthesis of 31 was followed starting with 29b and (4H-1,2,4-triazol-3-yl)methanamine hydrochloride to give 69 (47% yield) as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 8.20 (s, 1H), 7.39–7.06 (m, 7H), 6.80 (d, J = 8.7 Hz, 2H), 5.37 (s, 1H), 4.52 (dd, J = 52.1, 15.8 Hz, 2H), 3.89 (d, J = 6.6 Hz, 2H), 3.80 (q, J = 7.0 Hz, 1H), 2.85–2.58 (m, 1H), 2.19–2.03 (m, 2H), 2.03–1.75 (m, 4H), 1.46 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 176.6, 173.1, 160.7, 158.4, 148.1, 142.9, 130.3, 129.9, 129.6, 128.5, 128.0, 115.7, 73.2, 58.4, 47.0, 37.2, 36.1, 25.7, 19.4, 18.8; HRMS (ESI) m/z calcd for C₂₅H₂₉N₅O₃ [M + H]⁺ 448.2343, m/z found 448.2342.

(2S)-N-[(R)-(Hydrazinecarbonyl)({4-[(2-methylpentyl)oxy]phenyl})methyl]-2-phenylpropanamide (70a).

A mixture of 3 (596 mg, 1.5 mmol) and hydrazine hydrate (2 mL) in EtOH (10 mL) was refluxed for 3 h. After cooling to room temperature, the mixture was concentrated under reduced pressure. The residue was crystallized from MeOH/hexanes to give 70a (572 mg, 96% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.55 (s, 1H), 7.42–7.16 (m, 5H), 7.07 (d, J = 9.0 Hz, 2H), 6.76 (d, J = 9.0 Hz, 2H), 6.61 (d, J = 6.0 Hz, 1H), 5.43 (d, J = 6.0 Hz, 1H), 3.81 (s, 2H), 3.80–3.69 (m, 1H), 3.69–3.55 (m, 2H), 2.01–1.81 (m, 1H), 1.56–1.12 (m, 4H), 1.50 (d, J = 9.0 Hz, 3H), 0.99 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); MS (ESI) m/z 398.4 [M + H]⁺.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl](hydrazinecarbonyl)methyl]-2-phenylpropanamide (70b).

The procedure for the synthesis of 70a was followed starting with 11h to give 70b (90% yield) as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 7.33–7.19 (m, 5H), 7.16 (d, J = 8.6 Hz, 2H), 6.79 (d, J = 8.7 Hz, 2H), 5.28 (s, 1H), 3.87 (d, J = 6.6 Hz, 2H), 3.78 (q, J = 7.1 Hz, 1H), 2.85–2.63 (m, 1H), 2.19–2.02 (m, 2H), 2.02–1.77 (m, 4H), 1.44 (d, J = 7.1 Hz, 3H); ¹H NMR (300 MHz, CD₃OD) δ 176.5, 172.2, 160.7, 142.9, 130.4, 129.6, 129.6, 128.5, 128.0, 115.7, 73.2, 56.8, 47.0, 36.1, 25.7, 19.3, 18.8; MS (ESI) m/z 382.0 [M + H]⁺.

(2S)-N-[(R)-{4-[(2-Methylpentyl)oxy]phenyl}(1,3,4-oxadiazol-2-yl)methyl]-2-phenylpropanamide (71).

A mixture of 70a (70 mg, 0.18 mmol), trimethyl orthoformate (5 mL), and PTSA (45 mg, 0.23 mmol) was heated at 80 °C for 2 h. After cooling to room temperature, the mixture was diluted with EtOAc (50 mL), washed with NaHCO₃ (3 x 10 mL), brine (10 mL), dried (Na₂SO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–20% CMA80 in DCM to furnish 71 (40 mg, 56% yield) as an off-white solid. ¹H NMR (300 MHz; CDCl₃) δ 8.31 (s, 1H), 7.43–7.19 (m, 5H), 7.05 (d, J = 9.0 Hz, 2H), 6.79 (d, J = 9.0 Hz, 2H), 6.54 (d, J = 9.0 Hz, 1H), 6.33 (d, J = 6.0 Hz, 1H), 3.80–3.60 (m, 3H), 1.98–1.81 (m, 1H), 1.56–1.09 (m, 4H), 1.51 (d, J = 6.0 Hz, 3H), 0.98 (d, J = 6.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 165.9, 159.9, 163.2, 140.8, 128.9, 128.2, 127.7, 127.6, 127.4, 115.0, 73.3, 49.5, 46.7, 35.7, 32.8, 20.0, 18.4, 16.9, 14.2; HRMS (ESI) m/z calcd for C₂₄H₂₉N₃O₃ [M + H]⁺ 408.2282, m/z found 408.2278.

(2S)-N-[(R)-(5-Methyl-1,3,4-oxadiazol-2-yl)({4-[(2-methylpentyl)oxy]phenyl})-2-phenylpropanamide (72).

A mixture of 70a (100 mg, 0.25 mmol), trimethyl orthoacetate (0.065 mL, 0.5 mmol), and acetic acid (0.1 mL) in m-xylene (5 mL) was refluxed for 6 h. After cooling to room temperature, the mixture was diluted with EtOAc (50 mL), washed with NaHCO₃ (3 x 10 mL), brine (10 mL), dried (NaSO₄), and concentrated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–20% CMA80 in DCM to provide 72 (60 mg, 57% yield) as an off-white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.21 (m, 5H), 7.06 (d, J = 9.0 Hz, 2H), 6.78 (d, J = 9.0 Hz, 2H), 6.60 (d, J = 9.0 Hz, 1H), 6.25 (d, J = 6.0 Hz, 1H), 3.79–3.60 (m, 3H), 2.44 (s, 3H), 1.95–1.83 (m, 1H), 1.53–1.12 (m, 4H), 1.50 (d, J = 6.0 Hz, 3H), 0.98 (d, J = 9.0 Hz, 3H), 0.91 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 165.8, 164.2, 159.5, 140.8, 128.8, 128.2, 127.5, 127.2, 114.8, 73.1, 49.4, 46.8, 35.6, 32.7, 20.0, 18.4, 16.8, 14.2, 10.8; HRMS (ESI) m/z calcd for C₂₅H₃₁N₃O₃ [M + H]⁺ 422.2438, m/z found 422.2440.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl](1,3,4-oxadiazol-2-yl)methyl]-2-phenylpropanamide (73).

The procedure for the synthesis of 71 was followed starting with 70b and trimethyl orthoformate to give 73 (65% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.31 (s, 1H), 7.43–7.18 (m, 5H), 7.05 (d, J = 8.7 Hz, 2H), 6.79 (d, J = 8.7 Hz, 2H), 6.52 (d, J = 7.6 Hz, 1H), 6.33 (d, J = 7.7 Hz, 1H), 3.86 (d, J = 6.6 Hz, 2H), 3.67 (q, J = 7.1 Hz, 1H), 2.90–2.53 (m, 1H), 2.22–2.02 (m, 2H), 2.02–1.66 (m, 4H), 1.51 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.4, 165.9, 159.7, 153.2, 140.8, 128.9, 128.2, 127.8, 127.6, 127.4, 115.1, 72.1, 49.5, 46.8, 34.5, 24.8, 18.5, 18.4; HRMS (ESI) m/z calcd for C₂₃H₂₅N₃O₃ [M + H]⁺ 392.1969, m/z found 392.1967.

(2S)-N-[(R)-[4-(Cyclobutylmethoxy)phenyl](5-methyl-1,3,4-oxadiazol-2-yl)methyl]-2-phenylpro panamide (74).

The procedure for the synthesis of 72 was followed starting with 70b and trimethyl orthoacetate to give 74 (72% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.43–7.18 (m, 5H), 7.06 (d, J = 8.7 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 6.55 (d, J = 7.7 Hz, 1H), 6.24 (d, J = 7.7 Hz, 1H), 3.87 (d, J = 6.6 Hz, 2H), 3.66 (q, J = 7.1 Hz, 1H), 2.86–2.61 (m, 1H), 2.45 (s, 3H), 2.20–2.04 (m, 2H), 2.04–1.68 (m, 4H), 1.50 (d, J = 7.1 Hz, 3H); ¹³C NMR (75MHz, CDCl₃) δ 173.3, 165.8, 164.3, 159.5, 140.9, 128.9, 128.3, 128.2, 127.6, 127.3, 115.0, 72.1, 49.5, 46.8, 34.5, 24.8, 18.5, 18.5, 10.9; HRMS (ESI) m/z calcd for C₂₄H₂₇N₃O₃ [M + H]⁺ 406.2125, m/z found 406.2123.

Methyl (3S)-3-Amino-3-(4-hydroxyphenyl)propanoate (76).

(S)-3-Amino-3-(4-hydroxyphenyl)propanoic acid (75) (6.0 g, 33.15 mmol) was dissolved in dry MeOH (60 mL). Acetyl chloride (5 mL) was added to that above solution slowly via a syringe. The reaction that resulted was stirred at 65 °C for 12 h. After that, the solvent was evaporated under reduced pressure and the residue was dried in vacuo to furnish the methyl ester 76 as hydrochloride salt (8.63 g, 115% crude yield) as a foamy solid which was used for the next transformation without purification. ¹H NMR (300 MHz, CD₃OD) δ 7.35–7.24 (m, 2H), 6.84 (d, J = 8.6 Hz, 2H), 4.63 (t, J = 7.2 Hz, 1H), 3.68 (s, 3H), 3.12 (ddd, J = 16.8, 7.9, 1.6 Hz, 1H), 2.98 (dd, J = 16.8, 6.6 Hz, 1H); ¹³C NMR (75 MHz, CD₃OD) δ 171.8, 159.8, 129.8, 127.8, 117.0, 52.8, 52.7, 39.2; MS (ESI) m/z 196.0 [M + H]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-(4-hydroxyphenyl)propanoate (77).

To a solution of 76 (8.6 g, 37.1 mmol) in DCM (200 mL), DIPEA (38.8 mL, 0.22 mol) was added slowly via a syringe at 0 °C. To that above solution, Boc₂O (8.09 g, 37.1 mmol) was added and the reaction which resulted was stirred at room temperature under nitrogen for 12 h. At that time, water (100 mL) was to the reaction and the organic layer was separated. The aqueous layer was extracted with additional DCM (2 x 50 ML) and the combined organic layers were washed with brine and dried (Na₂SO₄). The solvent was evaporated under reduced pressure and the residue was crystallized with DCM/hexanes to furnish 77 (11.5 g, 78%) as an off-white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.04 (t, J = 9.2 Hz, 3H), 6.67 (d, J = 8.1 Hz, 2H), 5.54 (br s, 1H), 5.00 (br s, 1H), 3.61 (s, 3H), 2.96–2.60 (m, 2H), 1.43 (s, 9H); ¹³C NMR (75 MHz, CDCl₃) δ 171.7, 155.7, 155.4, 132.4, 127.3, 115.6, 80.0, 51.8, 50.9, 40.9, 28.3; MS (ESI) m/z 318.0 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-{4-[(2-methylpentyl)oxy]phenyl}propanoate (78a).

The procedure for the synthesis of 7b was followed starting with 77 and 2-methylpentanol to give 78a (87% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.6 Hz, 2H), 6.84 (d, J = 8.7 Hz, 2H), 5.32 (br s, 1H), 5.04 (br s, 1H), 3.79 (dd, J = 9.0, 5.8 Hz, 1H), 3.69 (dd, J = 8.9, 6.7 Hz, 1H), 3.61 (s, 3H), 2.82 (qd, J = 15.3, 6.2 Hz, 2H), 2.07–1.78 (m, 1H), 1.55–1.11 (m, 13H), 1.00 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.1 Hz, 3H); MS (ESI) m/z 402.0 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-[4-(cyclobutylmethoxy)phenyl]propanoate (78b).

The procedure for the synthesis of 7b was followed starting with 77 and cyclobutylmethanol to give 78b (87% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.6 Hz, 2H), 6.90–6.77 (m, 2H), 5.36 (s, 1H), 5.04 (d, J = 5.0 Hz, 1H), 3.90 (d, J = 6.7 Hz, 2H), 3.61 (s, 3H), 2.94–2.59 (m, 3H), 2.23–2.04 (m, 2H), 2.01–1.78 (m, 4H), 1.42 (s, 9H); ¹³C NMR (75 MHz, CDCl₃) δ 171.4, 158.7, 155.0, 133.1, 127.2, 114.7, 79.6, 72.1, 51.7, 50.9, 40.9, 34.6, 28.3, 24.8, 18.5; MS (ESI) m/z 386.0 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-(4-{[(2S)-2-methylpentyl]oxy}phenyl)propanoate (78c).

The procedure for the synthesis of 7b was followed starting with 77 and (S)-2-methylpentanol to give 78c (30% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.6 Hz, 2H), 6.84 (d, J = 8.7 Hz, 2H), 5.35 (br s, 1H), 5.04 (br s, 1H), 3.79 (dd, J = 9.0, 5.8 Hz, 1H), 3.69 (dd, J = 8.9, 6.7 Hz, 1H), 3.61 (s, 3H), 2.82 (qd, J = 15.3, 6.2 Hz, 2H), 2.01–1.77 (m, 1H), 1.55–1.09 (m, 13H), 1.00 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.1 Hz, 3H); MS (ESI) m/z 402.0 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-(4-{[(2R)-2-methylpentyl]oxy}phenyl)propanoate (78d).

The procedure for the synthesis of 11m was followed starting with 77 and (2R)-2-methylpentyl 4-methylbenzene-1-sulfonate to give 78d (35% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.6 Hz, 2H), 6.84 (d, J = 8.7 Hz, 2H), 5.38 (br s, 1H), 5.03 (br s, J = 5.4 Hz, 1H), 3.78 (dd, J = 9.0, 5.8 Hz, 1H), 3.68 (dd, J = 9.0, 6.7 Hz, 1H), 3.61 (s, 3H), 2.82 (qd, J = 15.3, 6.3 Hz, 2H), 2.00–1.76 (m, 1H), 1.56–1.11 (m, 13H), 1.00 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.4, 158.7, 155.0, 133.1, 127.2, 114.6, 79.6, 73.2, 51.6, 50.9, 40.9, 35.7, 32.9, 28.3, 20.0, 17.0, 14.3; MS (ESI) m/z 402.0 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-{4-[(2S)-2-methylbutoxy]phenyl}propanoate (78e).

The procedure for the synthesis of 7b was followed starting with 77 and (S)-2-methylbutanol to give 78e (95% yield) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.20 (d, J = 8.7 Hz, 2H), 6.84 (d, J = 8.7 Hz, 2H), 5.42 (br s, 1H), 5.03 (br s, 1H), 3.79 (dd, J = 9.0, 6.0 Hz, 1H), 3.70 (dd, J = 9.0, 6.6 Hz, 1H), 3.61 (s, 3H), 2.82 (qd, J = 15.3, 6.3 Hz, 2H), 1.90–1.75 (m, 1H), 1.564–1.48(m, 1H), 1.42 (s, 9H), 1.33–1.15 (m, 4H), 1.00 (d, J = 6.7 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.3, 158.7, 155.0, 133.1, 127.2, 114.6, 79.5, 72.8, 51.6, 50.9, 40.9, 34.7, 28.3, 26.1, 16.5, 11.2; MS (ESI) m/z 388 [M + Na]⁺.

Methyl (3S)-3-{[(tert-Butoxy)carbonyl]amino}-3-{4-[(2R)-2-methylbutoxy]phenyl}propanoate (78f).

The procedure for the synthesis of 11m was followed starting with 77 and (2R)-2-methylbutyl 4-methylbenzene-1-sulfonate to give 78f as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.19 (d, J = 8.7 Hz, 2H), 6.85 (d, J = 8.7 Hz, 2H), 5.34 (br s, 1H), 5.04 (br s, 1H), 3.83–3.66 (m, 2H), 3.61 (s, 3H), 2.82 (qd, J = 15.3, 6.2 Hz, 2H), 1.96–1.71 (m, 1H), 1.65–1.48 (m, 1H), 1.42 (br s, 9H), 1.33–1.16 (m, 1H), 1.00 (d, J = 6.7 Hz, 3H), 0.94 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 171.4, 158.7, 155.0, 133.0, 127.3, 114.6, 79.6, 72.9, 51.7, 50.9, 40.9, 34.7, 28.3, 26.1, 16.5. 11.3; MS (ESI) m/z 388 [M + Na]⁺.

Methyl (3S)-3-Amino-3-{4-[(2-methylpentyl)oxy]phenyl}propanoate Hydrochloride (79a).

A solution of 78a (1.63 g, 4.29 mmol) in DCM (20 mL) was cooled to 0 °C. To that above solution, HCl (4 M in dioxane, 10.74 mL, 42.95 mmol) was added via a syringe. The reaction which resulted was stirred at room temperature for overnight. At that time, the solvent was removed under reduced pressure and the residue was re-dissolved in DCM and evaporated (repeated 3 times). The residue was dried in vacuo to furnish 79a (1.35 g, 99%) as a foamy solid. ¹H NMR (300 MHz, CDCl₃) δ 8.72 (br s, 3H), 7.42 (d, J = 8.3 Hz, 2H), 6.86 (d, J = 8.1 Hz, 2H), 4.64 (br s, 1H), 3.87–3.64 (m, 2H), 3.60 (s, 3H), 3.35–3.15 (m, 1H), 3.00 (dd, J = 16.4, 6.5 Hz, 1H), 2.00–1.82 (m, 1H), 1.55–1.09 (m, 4H), 1.00 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); MS (ESI) free base m/z 263.0 [M + H⁺ - NH₃]⁺.

Methyl (3S)-3-Amino-3-[4-(cyclobutylmethoxy)phenyl]propanoate Hydrochloride (79b).

The procedure for the synthesis of 79a was followed starting with 78b to give 79b as a waxy solid. MS (ESI) free base m/z 247.0 [M + H⁺ - NH₃]⁺. This material was used for the next transformation without further characterization.

Methyl (3S)-3-Amino-3-(4-{[(2S)-2-methylpentyl]oxy}phenyl)propanoate Hydrochloride (79c).

The procedure for the synthesis of 79a was followed starting with 78c to give 79c as a waxy solid. MS (ESI) free base m/z 263.0 [M + H⁺ - NH₃]⁺. This material was used for the next transformation without further characterization.

Methyl (3S)-3-Amino-3-(4-{[(2R)-2-methylpentyl]oxy}phenyl)propanoate Hydrochloride (79d).

The procedure for the synthesis of 79a was followed starting with 78d to give 79d as a waxy solid. MS (ESI) free base m/z 263.0 [M + H⁺ - NH₃]⁺. This material was used for the next transformation without further characterization.

Methyl (3S)-3-Amino-3-{4-[(2S)-2-methylbutoxy]phenyl}propanoate Hydrochloride (79e).

The procedure for the synthesis of 79a was followed starting with 78e to give 79e as a waxy solid. MS (ESI) free base m/z 249.0 [M + H⁺ - NH₃]⁺. This material was used for the next transformation without further characterization.

Methyl (3S)-3-Amino-3-{4-[(2R)-2-methylbutoxy]phenyl}propanoate Hydrochloride (79f).

The procedure for the synthesis of 79a was followed starting with 78f to give 79f as a waxy solid. MS (ESI) free base m/z 249.0 [M + H⁺- NH₃]⁺.

Methyl (3S)-3-{4-[(2-Methylpentyl)oxy]phenyl}-3-[(2S)-2-phenylpropanamido]propanoate (80a).

To a solution of 79a (1.35 g, 4.27 mmol) in MeCN (50 mL), HBTU (2.43 g, 6.41 mmol), Et₃N (1.80 mL, 12.82 mmol) and (S)-(+)-2-phenylpropionic acid (642 mg, 4.27 mmol) was added and the reaction that resulted was stirred at room temperature 5 h. After the completion of the reaction, EtOAC (20 mL) and saturated NaHCO₃ (20 mL) was added and the organic layer was separated. The aqueous layer was extracted with additional EtOAc and the combined organic layers were washed with brine and dried (Na₂SO₄). The solvent was evaporated under reduced pressure. The residue was subjected to column chromatography on silica gel using 0–50% EtOAc in hexanes to furnish 80a (1.60 g, 91%) as a colorless oil. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.19 (m, 5H), 6.95 (d, J = 8.7 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.38 (d, J = 8.4 Hz, 1H), 5.31 (dt, J = 8.3, 5.9 Hz, 1H), 3.75 (dd, J = 8.9, 5.8 Hz, 1H), 3.69–3.57 (m, 2H), 3.55 (s, 3H), 2.77 (qd, J = 15.5, 5.9 Hz, 2H), 2.00–1.77 (m, 1H), 1.51 (d, J = 57.2 Hz, 3H), 1.48–1.09 (m, 4H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 171.5, 158.6, 141.3, 132.3, 128.9, 127.6, 127.2, 127.0, 114.5, 73.2, 51.7, 49.0, 47.1, 40.0, 35.7, 32.9, 20.0, 18.3, 17.0, 14.3; MS (ESI) m/z 412.0 [M + H]⁺.

Methyl (3S)-3-[4-(Cyclobutylmethoxy)phenyl]-3-[(2S)-2-phenylpropanamido]propanoate (80b).

The procedure for the synthesis of 80a was followed starting with 79b to give 80b (65% yield over 2 steps) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.29 (dd, J = 13.5, 6.6 Hz, 5H), 6.95 (d, J = 8.7 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.50 (d, J = 8.4 Hz, 1H), 5.31 (dt, J = 8.3, 6.0 Hz, 1H), 3.85 (d, J = 6.7 Hz, 2H), 3.59 (q, J = 7.1 Hz, 1H), 3.54 (s, 3H), 2.91–2.64 (m, 3H), 2.18–2.00 (m, 2H), 1.96–1.63 (m, 4H), 1.50 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 13.2, 171.5, 158.5, 141.3, 132.4, 128.9, 127.6, 127.2, 127.0, 114.6, 72.1, 51.7, 49.0, 47.1, 40.0, 34.6, 24.8, 18.5, 18.3; MS (ESI) m/z 396.0 [M + H]⁺.

Methyl (3S)-3-(4-{[(2S)-2-Methylpentyl]oxy}phenyl)-3-[(2S)-2-phenylpropanamido]propanoate (80c).

The procedure for the synthesis of 80a was followed starting with 79c to give 80c (61% yield) as a sticky solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.21 (m, 5H), 6.95 (d, J = 8.7 Hz, 2H), 6.73 (d, J = 8.7 Hz, 2H), 6.46 (d, J = 8.4 Hz, 1H), 5.32 (dt, J = 8.2, 5.9 Hz, 1H), 3.74 (dd, J = 8.9, 5.8 Hz, 1H), 3.69–3.58 (m 2H), 3.55 (s, 3H), 2.78 (qd, J = 15.5, 5.9 Hz, 2H), 2.01–1.80 (m, 1H), 1.52 (d, J = 7.2 Hz, 3H), 1.49–1.02 (m, 4H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.1 Hz, 3H); MS (ESI) m/z 412.0 ([M + H]⁺.

Methyl (3S)-3-(4-{[(2R)-2-Methylpentyl]oxy}phenyl)-3-[(2S)-2-phenylpropanamido]propanoate (80d).

The procedure for the synthesis of 80a was followed starting with 79d to give 80d (63% yield over two steps) as a sticky solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.15 (m, 5H), 6.95 (d, J = 8.6 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.43 (d, J = 8.4 Hz, 1H), 5.36–5.25 (m, 1H), 3.79–3.70 (m, 1H), 3.69–3.56 (m, 2H), 3.54 (s, 3H), 2.77 (qd, J = 15.5, 5.9 Hz, 2H), 1.98–1.81(m, 1H), 1.51 (d, J = 7.2 Hz, 3H), 1.49–1.11 (m, 4H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.3, 171.5, 158.6, 141.3, 132.3, 128.9, 127.6, 127.2, 127.0, 114.5, 73.2, 51.7, 49.0, 47.1, 40.0, 35.7, 32.9, 20.0, 18.3, 17.0, 14.3; MS (ESI) m/z 412.0 ([M + H]⁺.

Methyl (3S)-3-{4-[(2S)-2-Methylbutoxy]phenyl}-3-[(2S)-2-phenylpropanamido]propanoate (80e).

The procedure for the synthesis of 80a was followed starting with 79e to give 80e (64% yield over two steps) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.48–7.03 (m, 5H), 6.95 (d, J = 8.7 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.43 (d, J = 8.4 Hz, 1H), 5.30 (dd, J = 14.2, 6.0 Hz, 1H), 3.79–3.66 (m, 3H), 3.54 (s, 3H), 2.77 (qd, J = 15.5, 6.0 Hz, 2H), 1.90–1.75 (m, 1H), 1.64–1.37 (m, 4H), 1.36–1.08 (m, 1H), 0.98 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 171.5, 158.6, 141.3, 132.3, 128.8, 127.6, 127.2, 127.1, 114.5, 72.8, 51.7, 49.0, 47.1, 40.0, 34.7, 26.1, 18.3,16.5, 11.3; MS (ESI) m/z 398.0 [M + H]⁺.

Methyl (3S)-3-{4-[(2R)-2-Methylbutoxy]phenyl}-3-[(2S)-2-phenylpropanamido]propanoate (80f).

The procedure for the synthesis of 80a was followed starting with 79f to give 80f (54% yield over two steps) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.20 (m, 5H), 6.99–6.88 (m, 2H), 6.80–6.69 (m, 2H), 6.38 (d, J = 8.4 Hz, 1H), 5.35–5.25 (m, 1H), 3.79–3.71 (m, 1H), 3.70–3.56 (m, 2H), 3.55 (s, 6H), 2.77 (qd, J = 15.5, 5.9 Hz, 2H), 1.89–1.72 (m, 1H), 1.62–1.45 (m, 4H), 1.32–1.13 (m, 1H), 0.98 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.2, 171.5, 158.6, 141.3, 132.3, 128.9, 127.6, 127.2, 127.0, 114.5, 72.9, 51.7, 49.0, 47.1, 40.0, 34.7, 26.1, 18.3, 16.5, 11.3; MS (ESI) m/z 398.0 [M + H]⁺.

(2S)-N-[(1S)-2-(Hydrazinecarbonyl)-1-{4-[(2-methylpentyl)oxy]phenyl}ethyl]-2-phenylpropanamide (81a).

The procedure for the synthesis of 70a was followed starting with 80a to give 81a (92% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.21 (m, 5H), 6.93 (d, J = 8.7 Hz, 2H), 6.73 (d, J = 8.7 Hz, 2H), 6.68 (br s, 1H), 5.27–5.14 (m, 1H), 3.79–3.57 (m, 3H), 2.59 (qd, J = 14.6, 5.5 Hz, 2H), 1.99–1.82 (m, 1H), 1.65–1.09 (m, 9H), 0.99 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); MS (ESI) m/z 412.0 [M + H]⁺.

(2S)-N-[(1S)-1-[4-(Cyclobutylmethoxy)phenyl]-2-(hydrazinecarbonyl)ethyl]-2-phenylpropanamide (81b).

The procedure for the synthesis of 70a was followed starting with 80b to give 81b (90% yield) as a white solid. MS (ESI) m/z 396.0 [M + H]⁺. This material was used for the next transformation without further characterization.

(2S)-N-[(1S)-2-(Hydrazinecarbonyl)-1-(4-{[(2S)-2-methylpentyl]oxy}phenyl)ethyl]-2-phenylpropanamide (81c).

The procedure for the synthesis of 70a was followed starting with 80c to give 81c (67% yield) as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 7.30–7.06 (m, 5H), 6.94 (d, J = 8.6 Hz, 2H), 6.63 (d, J = 8.7 Hz, 2H), 5.15 (t, J = 7.1 Hz, 1H), 3.72 – 3.49 (m, 3H), 2.58–2.39 (m, 2H), 1.87–1.62 (m, 1H), 1.44–0.93 (m, 7H), 0.88 (d, J = 6.7 Hz, 3H), 0.82 (t, J = 7.1 Hz, 3H); MS (ESI) m/z 412.0 [M + H]⁺.

(2S)-N-[(1S)-2-Hydrazinecarbonyl)-1-(4-{[(2R)-2-methylpentyl]oxy}phenyl)ethyl]-2-phenylpropanamide (81d).

The procedure for the synthesis of 70a was followed starting with 80d to give 81d (75% yield) as a white solid. MS (ESI) m/z 412.0 [M + H]⁺. This material was used for the next transformation without further characterization.

(2S)-N-[(1S)-2-(Hydrazinecarbonyl)-1-{4-[(2S)-2-methylbutoxy]phenyl}ethyl]-2-phenylpropanamide (81e).

The procedure for the synthesis of 70a was followed starting with 80e to give 81e (97% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.43–7.17 (m, 5H), 6.95 (m, 4H), 6.73 (d, J = 8.7 Hz, 2H), 5.26–5.15 (m, 1H), 3.80–3.45 (m, 4H), 2.58 (qd, J = 14.6, 5.6 Hz, 2H), 1.91–1.41 (m, 6H), 1.34–1.12 (m, 1H), 0.98 (d, J = 6.7 Hz, 3H), 0.93 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 171.2, 158.6, 141.3, 132.3, 128.8, 127.6, 127.2, 126.9, 114.6, 72.9, 49.9, 47.2, 40.1, 34.7, 26.1, 18.2, 16.5, 11.3; (ESI) m/z 398.0 [M + H]⁺.

(2S)-N-[(1S)-2-(Hydrazinecarbonyl)-1-{4-[(2R)-2-methylbutoxy]phenyl}ethyl]-2-phenylpropanamide (81f).

The procedure for the synthesis of 70a was followed starting with 80f to give 81f (90% yield) as a white solid. MS (ESI) m/z 398.0 [M + H]⁺. This material was used for the next transformation without further characterization.

(2S)-N-[(1S)-1-{4-[(2-Methylpentyl)oxy]phenyl}-2-(1,3,4-oxadiazol-2-yl)ethyl]-2-phenylpropanamide (82).

The procedure for the synthesis of 71 was followed starting with 81a and trimethyl orthoformate to give 82 (35% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.27 (s, 1H), 7.28 (m, 5H), 6.96 (d, J = 8.6 Hz, 2H), 6.74 (d, J = 8.6 Hz, 2H), 6.29 (d, J = 8.3 Hz, 1H), 5.42 (dd, J = 14.1, 7.3 Hz, 1H), 3.78–3.69 (m, 1H), 3.68–3.51 (m, 2H), 3.43–3.15 (m, 2H), 1.99–1.80 (m, 1H), 1.46 (d, J = 7.2 Hz, 3H), 1.43–1.07 (m, 4H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 164.0, 158.9, 153.0, 141.0, 131.2, 128.9, 127.6, 127.2, 127.1, 114.7, 73.2, 50.0, 47.0, 35.7, 32.8, 31.7, 20.0, 18.3, 17.0, 14.3; HRMS (ESI) m/z calcd for C₂₅H₃₁N₃O₃ [M + H]⁺ 422.2438, m/z found 422.2432.

(2S)-N-[(1S)-2-(5-Methyl-1,3,4-oxadiazol-2-yl)-1-{4-[(2-methylpentyl)oxy]phenyl}ethyl]-2-phenylpropanamide (83).

The procedure for the synthesis of 72 was followed starting with 81a and trimethyl orthoacetate to give 83 (93% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.38–7.16 (m, 5H), 6.97 (d, J = 8.6 Hz, 2H), 6.73 (d, J = 8.7 Hz, 2H), 6.41 (d, J = 8.4 Hz, 1H), 5.46–5.29 (m, 1H), 3.78–3.69 (m, 1H), 3.68–3.48 (m, 2H), 3.35–3.07 (m, 2H), 2.42 (s, 3H), 1.98–1.79 (m, 1H), 1.52–1.10 (m, 7H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 164.0, 163.9, 158.8, 141.1, 131.6, 128.8, 127.6, 127.2, 127.2, 114.7, 73.2, 49.9, 47.0, 35.7, 32.8, 31.8, 20.0, 18.3, 17.0, 14.3, 10.8; HRMS (ESI) m/z calcd for C₂₆H₃₃N₃O₃ [M + H]⁺ 436.2595, m/z found 436.2609.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-{4-[(2-methylpentyl)oxy]phenyl}ethyl]-2-phenylpropanamide Hydrochloride (84).

To a solution of 81a (28 mg, 0.07 mmol) in dry MeOH (2.5 mL), cyanogen bromide (8 mg, 0.075 mmol) was added and the reaction that resulted was heated to reflux under nitrogen for 4 h. The reaction was cooled to room temperature and quenched with saturated NaHCO₃ (5 mL) and extracted with EtOAc (3 x 10 mL). The combined organic layers were washed with brine (2 x 10 mL) and dried (Na₂SO₄). The solvent was evaporated, and the residue was subjected to column chromatography on silica gel using 0–50% CMA80 in DCM to furnish 84 (free base, 22 mg, 75% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.34–7.17 (m, 5H), 6.99 (d, J = 8.6 Hz, 2H), 6.74 (d, J = 8.7 Hz, 2H), 6.51 (d, J = 8.5 Hz, 1H), 5.45 (br s, 2H), 5.37 (dd, J = 14.4, 7.6 Hz, 1H), 3.73 (dd, J = 8.9, 5.8 Hz, 1H), 3.68–3.48 (m, 2H), 3.26–3.01 (m, 2H), 1.96–1.80 (m, 1H), 1.52–1.05 (m, 7H), 0.98 (d, J = 6.7 Hz, 3H), 0.90 (t, J = 7.0 Hz, 3H). The above solid was suspended in DCM (2 mL) and cooled to 0 °C. To that above suspension, HCl (126 μL, 2.0 M in diethyl ether) was added dropwise. The solution that resulted was stirred for 30 min at 0 °C. The solvent was evaporated and the solid was triturated with MeOH and hexanes to furnish the hydrochloride salt 84 as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 8.52 (d, J = 8.3 Hz, 1H), 7.34–7.19 (m, 5H), 7.16 (d, J = 8.6 Hz, 2H), 6.83 (d, J = 8.7 Hz, 2H), 5.37–5.18 (m, 1H), 3.86–3.59 (m, 3H), 3.24 (d, J = 7.3 Hz, 2H), 2.05–1.78 (m, 1H), 1.61–1.30 (m, 6H), 1.30–1.11 (m, 1H), 1.01 (d, J = 6.7 Hz, 3H), 0.97–0.87 (m, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 176.5, 163.3, 160.4, 159.4, 142.7, 133.0, 129.5, 128.7, 128.4, 127.9, 115.7, 74.3, 51.4, 47.3, 36.9, 34.1, 32.8, 21.1, 18.8, 17.3, 14.6; HRMS (ESI) free base m/z calcd for C₂₅H₃₂N₄O₃ [M + H]⁺ 437.2547, m/z found 437.2546.

(2S)-N-[(1S)-1-[4-(Cyclobutylmethoxy)phenyl]-2-(1,3,4-oxadiazol-2-yl)ethyl]-2-phenylpropanamide (85).

The procedure for the synthesis of 71 was followed starting with 81b and trimethyl orthoformate to give 85 (57% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 8.27 (s, 1H), 7.39–7.17 (m, 5H), 6.96 (d, J = 8.7 Hz, 2H), 6.75 (d, J = 8.7 Hz, 2H), 6.17 (d, J = 8.3 Hz, 1H), 5.42 (dd, J = 14.0, 7.3 Hz, 1H), 3.85 (d, J = 6.6 Hz, 2H), 3.56 (q, J = 7.1 Hz, 1H), 3.45–3.22 (m, 2H), 2.79–2.63 (m, 1H), 2.24–1.98 (m, 2H), 1.97–1.70 (m, 4H), 1.47 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.6, 164.0, 158.9, 153.0, 141.0, 131.3, 128.9, 127.6, 127.3, 127.1, 114.8, 72.1, 50.0, 47.0, 34.6, 31.7, 24.8, 24.5, 18.6, 18.3; HRMS (ESI) m/z calcd for C₂₄H₂₇N₃O₃ [M + H]⁺ 406.2125, m/z found 406.2122.

(2S)-N-[(1S)-1-[4-(Cyclobutylmethoxy)phenyl]-2-(5-methyl-1,3,4-oxadiazol-2-yl)ethyl]-2-phenylpropanamide (86).

The procedure for the synthesis of 72 was followed starting with 81b and trimethyl orthoacetate to give 86 (88% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.39–7.17 (m, 5H), 6.96 (d, J = 8.6 Hz, 2H), 6.83–6.65 (m, 2H), 6.25 (d, J = 8.3 Hz, 1H), 5.39 (dd, J = 14.7, 6.6 Hz, 1H), 3.85 (d, J = 6.6 Hz, 2H), 3.57 (q, J = 7.1 Hz, 1H), 3.39–3.09 (m, 2H), 2.80–2.63 (m, 1H), 2.44 (s, 3H), 2.20–2.02 (m, 2H), 2.02–1.65 (m, 4H), 1.47 (d, J = 7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.5, 163.9, 158.8, 141.1, 131.6, 128.9, 127.6, 127.2, 127.1, 114.7, 72.1, 49.9, 47.0, 34.6, 31.9, 24.8, 18.5, 18.3, 10.8; HRMS (ESI) m/z calcd for C₂₅H₂₉N₃O₃ [M + H]⁺ 420.2282, m/z found 420.2281.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-[4-(cyclobutylmethoxy)phenyl]ethyl]-2-phenylpropanamide Hydrochloride (87).

The procedure for the synthesis of 84 was followed starting with 81b and cyanogen bromide to give 87 (80% yield) as a white solid. ¹H NMR [free base] (300 MHz, CD₃OD) δ 7.36–7.14 (m, 5H), 7.10 (d, J = 8.7 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 5.28 (t, J = 7.6 Hz, 1H), 3.87 (d, J = 6.6 Hz, 2H), 3.65 (q, J = 7.0 Hz, 1H), 3.16 (d, J = 7.6 Hz, 2H), 2.84–2.46 (m, 1H), 2.19–2.03 (m, 2H), 2.03–1.64 (m, 4H), 1.39 (d, J = 7.1 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 176.3, 165.8, 160.2, 159.3, 142.8, 133.6, 129.5, 128.6, 128.4, 127.9, 115.6, 73.2, 51.6, 47.4, 36.1, 32.9, 25.7, 19.3, 18.9; HRMS (ESI) free base m/z calcd for C₂₄H₂₈N₄O₃ [M + H]⁺ 421.2234, m/z found 421.2231.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-(4-{[(2S)-2-methylpentyl]oxy}phenyl)ethyl]-2-phenylpropanamide Hydrochloride (88).

The procedure for the synthesis of 84 was followed starting with 81c and cyanogen bromide to give 88 (60% yield) as a white solid. ¹H NMR (300 MHz, CDCl₃) δ 7.40–7.16 (m, 5H), 6.98 (d, J = 8.7 Hz, 2H), 6.75 (d, J = 8.6 Hz, 2H), 6.30 (d, J = 8.4 Hz, 1H), 5.35 (dd, J = 14.8, 6.8 Hz, 1H), 5.06 (br s, 2H), 3.81–3.45 (m, 3H), 3.13 (d, J = 6.7 Hz, 2H), 1.95–1.76 (m, 1H), 1.46 (d, J = 7.2 Hz, 3H), 1.43–1.10 (m, 4H), 0.98 (d, J = 6.7 Hz, 3H), 0.91 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.6, 162.8, 158.8, 158.2, 141.1, 131.7, 128.8, 127.6, 127.2, 127.1, 114.7, 73.2, 49.9, 47.0, 35.7, 32.8, 32.0, 20.0, 18.3, 17.0, 14.3; HRMS (ESI) free base m/z calcd for C₂₅H₃₂N₄O₃ [M + H]⁺ 437.2547, m/z found 437.2537.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-(4-{[(2R)-2-methylpentyl]oxy}phenyl)ethyl]-2-phenylpropanamide Hydrochloride (89).

The procedure for the synthesis of 84 was followed starting with 81d and cyanogen bromide to give 89 (75% yield) as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 7.33–7.16 (m, 5H), 7.12 (d, J = 8.7 Hz, 2H), 6.80 (d, J = 8.6 Hz, 2H), 5.30 (t, J = 7.6 Hz, 1H), 3.84–3.60 (m, 3H), 3.18 (d, J = 7.5 Hz, 2H), 1.97–1.88 (m, 1H), 1.57–1.12 (m, 7H), 1.01 (d, J = 6.7 Hz, 3H), 0.94 (t, J = 7.0 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 176.3, 165.8, 160.2, 159.3, 142.8, 133.5, 129.5, 128.6, 128.4, 127.9, 115.6, 74.3, 51.6, 47.0, 36.9, 34.1, 32.9, 21.1, 18.9, 17.3, 14.6; HRMS (ESI) free base m/z calcd for C₂₅H₃₂N₄O₃ [M + H]⁺ 437.2547, m/z found 437.2539.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-{4-[(2S)-2-methylbutoxy]phenyl}ethyl]-2-phenylpropanamide Hydrochloride (90).

The procedure for the synthesis of 84 was followed starting with 81e and cyanogen bromide to give 90 (94% yield) as a white solid. ¹H NMR [free base] (300 MHz, CDCl₃) δ 7.36–7.11 (m, 5H), 7.10–6.97 (m, 3H), 6.71 (d, J = 8.6 Hz, 2H), 6.20 (br s, 2H), 5.41 (dt, J = 14.2, 7.2 Hz, 1H), 3.71 (dd, J = 9.0, 6.0 Hz, 1H), 3.58 (ddd, J = 21.3, 11.5, 6.8 Hz, 2H), 3.16 (qd, J = 15.2, 7.2 Hz, 2H), 1.89–1.71 (m, 1H), 1.63–1.44 (m, 1H), 1.39 (d, J = 7.1 Hz, 3H), 1.30–1.11 (m, 1H), 0.97 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 7.4 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ 173.9, 163.7, 158.8, 157.9, 141.2, 132.0, 128.7, 127.6, 127.2, 127.0, 114.7, 72.9, 49.9, 46.8, 34.7, 32.1, 26.1, 18.4, 16.5, 11.3; HRMS (ESI) free base m/z calcd for C₂₄H₃₀N₄O₃ [M + H]⁺ 423.2391, m/z found 423.2388.

(2S)-N-[(1S)-2-(5-Amino-1,3,4-oxadiazol-2-yl)-1-{4-[(2R)-2-methylbutoxy]phenyl}ethyl]-2-phenylpropanamide Hydrochloride (91).

The procedure for the synthesis of 84 was followed starting with 81f and cyanogen bromide to give 91 (59% yield) as a white solid. ¹H NMR (300 MHz, CD₃OD) δ 7.32–7.02 (m, 5H), 6.93 (d, J = 8.6 Hz, 2H), 6.63 (d, J = 8.7 Hz, 2H), 5.14 (t, J = 7.1 Hz, 1H), 3.77–3.48 (m, 3H), 2.50 (d, J = 7.1 Hz, 2H), 1.76–1.60 (m, 1H), 1.53–1.37 (m, 1H), 1.32 (d, J = 7.1 Hz, 3H), 1.23–1.06 (m, 1H), 0.88 (d, J = 6.7 Hz, 3H), 0.83 (t, J = 7.5 Hz, 3H); ¹³C NMR (75 MHz, CD₃OD) δ 175.9, 172.0, 160.0, 142.9, 134.4, 129.5, 128.5, 128.5, 127.9, 115.4, 73.9, 51.4, 47.5, 41.3, 36.0, 27.2, 18.8, 16.8, 11.6; HRMS (ESI) free base m/z calcd for C₂₄H₃₀N₄O₃ [M + H]⁺ 423.2391, m/z found 423.2383.

Pharmacology.

Materials.

Cell culture materials were purchased from Fisher SSI. Forskolin was purchased from Sigma-Aldrich. The Lance Ultra kit (TRF0262) was purchased from PerkinElmer.

Lance™ Ultra cAMP assay using stable PPLS-HA-GPR88 CHO cells.

All cAMP assays were performed using our previously published methods.²⁶ Stimulation buffer containing 1X Hank’s Balanced Salt Solution (HBSS), 5 mM HEPES, 0.1% BSA stabilizer, and 0.5 mM final IBMX was prepared and titrated to pH 7.4 at room temperature. Serial dilutions of the test compounds (5 μL) and 300 nM forskolin (5 μL), both prepared at 4x the desired final concentration in 2% DMSO/stimulation buffer, were added to a 96-well white ½ area microplate (PerkinElmer). A cAMP standard curve was prepared at 4x the desired final concentration in stimulation buffer and 5 μL was added to the assay plate. Stable PPLS-HA-GPR88 CHO cells were lifted with versene and spun at 270g for 10 minutes. The cell pellet was resuspended in stimulation buffer and 4,000 cells (10 μL) were added to each well except wells containing the cAMP standard curve. After incubating for 30 min at room temperature, Eu-cAMP tracer and uLIGHT-anti-cAMP working solutions were added per the manufacturer’s instructions. After incubation at room temperature for 1 hour, the TR-FRET signal (ex 337 nm) was read on a CLARIOstar multimode plate reader (BMG Biotech, Cary, NC).

Data Analysis.

The TR-FRET signal (665 nm) was converted to fmol cAMP by interpolating from the standard cAMP curve. Fmol cAMP was plotted against the log of compound concentration and data were fit to a three-parameter logistic curve to generate EC₅₀ values (Prism, version 6.0, GraphPad Software, Inc., San Diego, CA). The E_max value for each test compound relative to the control compounds RTI-13951-33 or 2-PCCA was calculated with the equation % control E_max = (maximal test compound signal / maximal control signal) x 100.

[³⁵S]GTPγS Binding Assay.

[³⁵S]GTPγS binding assays were performed on membrane preparations from wild-type (WT) mice or GPR88 KO mice, following our previously published methods.^{14, 15} To assess [³⁵S]GTPγS binding in the whole striatal region, brains were quickly removed after cervical dislocation and the whole striatal region was dissected out, frozen, and stored at −80 °C until use. Membranes were prepared by homogenizing brain samples in ice-cold 0.25 M sucrose solution 10 vol (mL/g wet weight of tissue). The obtained suspensions were then centrifuged at 2500 g for 10 min. Supernatants were collected and diluted ten times in buffer containing 50 mM TrisHCl (pH 7.4), 3 mM MgCl₂, 100 mM NaCl, 0.2 mM EGTA, and then centrifuged at 23,000 g for 30 min. The pellets were homogenized in 800 μL ice-cold sucrose solution (0.32 M), aliquoted and kept at −80 °C. For [³⁵S]GTPγS binding assays, 2 μg of protein was used per well. Samples were incubated with and without the test compound for 1 h at 25 °C in an assay buffer containing 30 mM GDP and 0.1 nM [³⁵S]GTPγS. Bound radioactivity was quantified using a liquid scintillation counter. Non-specific binding was defined as binding in the presence of 10 μM GTPγS; basal binding refers to binding in the absence of the agonist. Data were expressed as a mean percentage of activation above the basal binding. GTPγS binding by agonist was plotted with X axis representing concentration and Y axis representing the percentage of activation against background. EC₅₀ values were calculated using GraphPad Prism software.

Solubility Determination.

For kinetic solubility experiments, 10 mM DMSO stocks of compounds were directly diluted into 10 mM phosphate buffer at pH 7.4 and shaken for 90 min at room temperature. The final concentration of DMSO was 1%. After the incubation, samples were filtered through a 0.4 μm filter plate (Millipore). Filtrates were carefully collected. On each experimental occasion, tamoxifen and caffeine were assessed as reference compounds for low and high solubility, respectively. All samples were assessed in triplicate and analyzed by LC-MS/MS using electrospray ionization against standards prepared in the same matrix.

Pharmacokinetic Analysis.

PK study of 90 was performed using male Long-Evans rats (Paraza Pharma Inc., Montreal, Canada). Doses were formulated in 5% dimethylacetamide in sesame oil. On the morning of the PK study, animals were weighed, and dosing formulation volumes were calculated accordingly. The compound was injected intraperitoneally to all animals. At selected time points (0.5, 1, 2, 4, and 8 h postdose), animals were anesthetized to perform a cardiac puncture to collect blood for pooled plasma analysis, followed by whole body perfusion with phosphate saline buffer (pH 7.4) to wash out any remaining blood from the organs. Brains were harvested and homogenized by polytron 1:4 (w/v) in 25% isopropanol in water. Brain homogenates were further pooled per corresponding time point and extracted for drug quantification of LC-MS/MS. Samples were prepared and analyzed as follows: Plasma (10 μL) was mixed with 10 μL of 0.5% formic acid in water, 100 μL internal standard working solution (0.1 μM Glyburide/Labetalol in 0.5% ammonium formate in methanol/acetonitrile), vortexed, and centrifuged at 10000 g for 10 min at 4 °C. Supernatant (100 μL) was transferred to a 2 mL deepwell plate and diluted with 200 μL 30% acetonitrile/water. Brain homogenate (25 μL) was mixed with 25 μL of 0.5% formic acid in water, 150 μL internal standard working solution (0.1 μM Glyburide/Labetalol in methanol/acetonitrile), vortexed, and centrifuged at 10000 g for 10 min at 4 °C. Supernatant (100 μL) was transferred to a 2 mL deepwell plate and diluted with 100 μL water. LC-MS/MS was conducted using an Applied Biosystems API 4000 HPLC system. Chromatography was performed with an Xbridge BEH C18 (2.1 × 30 mm, 2.5 μm) column. Mobile phases were 0.1% formic acid in water (A), and 0.1% formic acid in 25% isopropanol/acetonitrile (B). Initial conditions were 5% B and held for 0.5 min, followed by a linear gradient to 95% B over 1.8 min. 95% B was held for 2.6 min before returning to initial conditions.

ABBREVIATIONS USED

ADME

absorption, distribution, metabolism, and excretion

2-AMPP

(2S)-N-((1R)-2-amino-1-(4-(2-methyl-pentyloxy)phenyl)ethyl)-2-phenylpropanamide

cAMP

cyclic adenosine monophosphate

BBB

blood–brain barrier

BRET

bioluminescence resonance energy transfer

CHO cells

Chinese hamster ovary cells

DCM

dichloromethane

DEAD

diethyl azodicarboxylate

DIPEA

N,N-diisopropylethylamine

DMF

N,N-dimethylformamide

EDC

1-ethyl-3-(3-dimethylaminopropyl)carbodiimide

GPCR

G protein-coupled receptor

HA

human influenza hemagglutinin

HBTU

2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate

HEK293 cells

human embryonic kidney cells

HOBt

hydroxybenzotriazole

KO

knockout

MS

mass spectroscopy

2-PCCA

(1R,2R)-2-(pyridin-2-yl)cyclopropane carboxylic acid ((2S,3S)-2-amino-3-methylpentyl)-(4′-propylbiphenyl-4-yl)amide

Pgp

P-glycoprotein

PPLS

pre-prolactin leader sequence

PTSA

p-toluenesulfonic acid

TBAF

tetrabutylammonium fluoride

TEA

triethylamine

TFA

trifluoroacetic acid

THF

tetrahydrofuran

TIPS

triisopropylsilyl

TIPSCl

triisopropylsilyl chloride

TLC

thin layer chromatography

TPSA

topological polar surface area

SAR

structure-activity relationship