Discovery of Cytochrome P450 4F11 Activated Inhibitors of Stearoyl

Article Cite This: J. Med. Chem. 2018, 61, 5199−5221

pubs.acs.org/jmc

Discovery of Cytochrome P450 4F11 Activated Inhibitors of Stearoyl Coenzyme A Desaturase Sarah E. Winterton,† Emanuela Capota,‡ Xiaoyu Wang,† Hong Chen,† Prema L. Mallipeddi,† Noelle S. Williams,† Bruce A. Posner,† Deepak Nijhawan,*,†,‡ and Joseph M. Ready*,† †

Department of Biochemistry, ‡Department of Internal Medicine, UT Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, Texas 75390-9038, United States

Downloaded via UNIV OF TOLEDO on June 30, 2018 at 06:21:16 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

S Supporting Information *

ABSTRACT: Stearoyl-CoA desaturase (SCD) catalyzes the first step in the conversion of saturated fatty acids to unsaturated fatty acids. Unsaturated fatty acids are required for membrane integrity and for cell proliferation. For these reasons, inhibitors of SCD represent potential treatments for cancer. However, systemically active SCD inhibitors result in skin toxicity, which presents an obstacle to their development. We recently described a series of oxalic acid diamides that are converted into active SCD inhibitors within a subset of cancers by CYP4F11-mediated metabolism. Herein, we describe the optimization of the oxalic acid diamides and related N-acyl ureas and an analysis of the structure−activity relationships related to metabolic activation and SCD inhibition.

■

INTRODUCTION Cells require unsaturated fatty acids (UFAs) for properly functioning membranes. Dividing cells utilize UFAs to form new membranes, and UFAs can be obtained either through uptake from the environment or from de novo synthesis. Tumor cells in particular rely on UFA biosynthesis because they frequently exist in a nutrient-poor environments. They synthesize UFAs to maintain membrane fluidity and to buffer the cell from the toxic effects of saturated fatty acids and free cholesterol.1 For these reasons, inhibition of de novo UFA biosynthesis represents an attractive approach to treating solid tumors. The enzyme stearoyl-CoA desaturase (SCD) catalyzes the introduction of unsaturation at the C9−C10 position of saturated fatty acids (Figure 1).2 Humans express two isoforms, SCD1 and SCD5.3 Human SCD1 is highly homologous to the mouse and rat isoforms and is ubiquitously expressed. In humans, SCD1 appears to play the predominant biological role.2 For simplicity, we will refer to SCD1 as SCD hereafter. SCD is a nonheme diiron oxidase that functions in the endoplasmic reticulum. It primarily converts stearoyl-CoA and palmitoyl-CoA to oleate and palmitoleate, respectively. Because of its role in the generation of UFAs, SCD has emerged as an attractive target for treating cancer. Indeed, a recent functional genomic screen of 66 genes involved in lipid biosynthesis identified SCD as essential in breast- and prostatecancer cells.4 Similarly, patient samples from clear-cell-renalcell-carcinoma tumors showed elevated expression of SCD and increased amounts of unsaturated fatty acids, and the levels of both enzyme and lipids were correlated with tumor growth rates.5 © 2018 American Chemical Society

Inhibitors of SCD have been developed for the treatment of metabolic diseases and cancer. Several pharmaceutical companies have developed potent and bioavailable inhibitors of SCD, and the area has been reviewed recently.6 For example, Abbott disclosed urea 3 as a low-nanomolar inhibitor of SCD (Figure 2).7 Scientists from Bristol-Myers Squibb reported that this SCD inhibitor slowed the growth of gastric cancers in tumor-xenograft experiments.8 In similar studies, 3 slowed the growth of a tumor xenograft derived from a renalcell-carcinoma line when dosed in combination with an mTOR inhibitor.9 Additional studies from Abbott led to pyridazine 4, a potent and orally bioavailable SCD inhibitor,10 which was found to slow the growth of a tumor xenograft derived from the human colorectal-cancer cell line HCT116.11 CV Therapeutics described a series of quinazolin-4-ones, exemplified by compound 5, with an EC50 of 119 nM against human SCD and excellent oral bioavailability.12 These inhibitors blocked proliferation of lung-cancer H460 cells without affecting fibroblasts under the same conditions.13 Finally, several potent SCD inhibitors have been shown to affect triglyceride levels in vivo, including thiazolylpyridinone 6 from Novartis and Xenon Pharmaceuticals,14 isoxazole 7 (MK8245) from Merck, 15 and pyridazine 8 from Xenon Pharmaceuticals.16 Inhibition of SCD in the skin results in toxicity that presents an obstacle to the clinical development of conventional SCD inhibitors. Specifically, mice treated with known inhibitors suffer atrophy of sebocytes.17 These skin cells require SCD to Received: January 12, 2018 Published: June 5, 2018 5199

DOI: 10.1021/acs.jmedchem.8b00052 J. Med. Chem. 2018, 61, 5199−5221

Journal of Medicinal Chemistry

Article

Figure 1. SCD enzymatic reaction and inhibition by tumor-activated oxalamides.

Figure 2. Inhibitors of SCD.

determined that, in sensitive cells, active oxalamides covalently bound to and inhibited SCD. Consistent with previous reports, inhibition of SCD enzymatic function starved cells for UFAs, leading to cell death. The selective toxicity displayed by the oxalamides arises from the overexpression of cytochrome P450 isoform CYP4F11 within sensitive cells.23 This enzyme metabolizes compounds such as 1 (Figure 1), which are in fact prodrugs, into an active SCD inhibitors (e.g., 2) through O-demethylation. The CYP-mediated activation of SCD inhibitors provides a potential means to increase the therapeutic index of SCD inhibitors by minimizing systemic exposure. Herein, we describe the discovery of optimized SCD inhibitors derived from the oxalamide scaffold.

synthesize sebum, which contains esters of fatty acids and fatty alcohols. Sebum is excreted onto the skin by hair follicles to reduce heat loss and onto the eyes and eyelids for lubrication. As a result of sebocyte atrophy, mice treated with SCD inhibitors suffer from dry eye, dry skin, and hypothermia. These effects are severe: the global-SCD-knockout mouse shows alopecia and closed eyes and will die from hypothermia after only a few hours at 4 °C.18 By contrast, inhibition of SCD in the liver appears well tolerated, as mice with liver-specific SCD knockouts were indistinguishable from wild-type mice.19 Similarly, Merck reported no toxicity in a Phase II trial of an SCD inhibitor that utilizes organic anion transporters to accumulate in the liver.20 We recently described the discovery of tumor-targeted inhibitors of SCD.21 These compounds emerged from a highthroughput screen against 12 non-small-cell-lung-cancer (NSCLC) lines in conjunction with the Cancer Target Discovery and Development Network.22 The NSCLC lines were selected to maximize diversity in terms of expression profiles and copy-number variation. In this experiment, each of approximately 200 000 compounds was tested at 2.5 μM against 12 lung-cancer lines, and viability was assessed by measuring ATP levels with CellTiter-Glo. The experiment was designed to reveal small molecules with selective toxicity toward a subset of the NSCLC lines. We therefore developed an algorithm to cluster cells into a “Sensitive” and “Insensitive” groups for each compound. We then identified the compounds with the largest difference between the two groups, reflecting a high degree of selective toxicity. Among the most selective compounds were a series of compounds containing an oxalic acid diamide moiety, referred to herein as oxalamides and exemplified by compound 9 (see Table 1). Ultimately, we

■

RESULTS AND DISCUSSION The HTS experiment described above returned compound 9 as a selective and modestly potent cytotoxin (Table 1). It showed EC50 values 95% toxicity to each cell line. By contrast, eight other NSCLC cell lines were substantially less sensitive to 9 (EC50 > 10 μM; H1155, H1395, H1819, H1993, H2009, H2073, HCC366, and HCC4017). We set out to obtain potent and selective toxins and to generate probe reagents to facilitate mechanism-ofaction studies. At the outset of our project, we did not know the biological target of the oxalamides, so we used cytotoxicity to H2122 cells to drive our optimization efforts. The anilide ring of 9 features a chlorine substituent, and other small substituents in this position (H, F, CN) provided similar activity (Table 1).24 However, we found that introduction of an aryl ketone at the 4-position of the anilide improved potency against H2122 cells. 5200



Article

Table 1. Anilide Substitutions

Table 2. Heteroaryl Anilide Replacements

the nitrogen linker were acceptable (40−42, 44, and 45), albeit without conferring obvious improvement in activity. In an effort to avoid the aniline substructure, we attempted to cyclize one of the amide groups into a heterocyclic ring. Unfortunately, the heterocyclic replacements shown in Table 3 Table 3. Cyclic Anilide Replacements

For example, the simple benzophenone 13 and the methoxysubstituted benzophenone 14 displayed EC50’s around 80 nM. We were hopeful to use the benzophenone as a photo-crosslinking agent, so we incorporated a propargyl ether (15) and were gratified to see a further increase in potency to 30 nM, with essentially complete cell killing at >1 μM. We briefly explored substitution at the other positions of the anilide ring. Even small substituents ortho to the nitrogen (18) totally abrogated activity. By contrast, substitution at the meta position was tolerated (20−22), but offered no improvement, whereas 3,5-disubstitution (23−25) was not tolerated. A series of 3,4-disubstituted analogues revealed the SAR within fairly narrow limits. For example, small electron-withdrawing groups at the 3-position (26−30) supported submicromolar EC50’s, but larger groups like nitro (31) or electron-donating methoxy (34) groups decreased activity substantially. We attempted to replace the N-aryl ring of 9 with an Nheteroaryl ring (Table 2). Although a 4-pyridine (36) and a pyrazine (37) were not tolerated, 2- and 3-pyridyl rings were viable replacements. In general, the SAR within the 3aminopyridines (38−42) and the 2-aminopyridines (43−47) was consistent with what we observed in the anilide series. For example, substitution ortho to the exocyclic nitrogen (38 and 39) was not tolerated. However, small groups meta and para to

were all inactive against H2122 cells. We tested both 5,6- and 6,6-bicyclic structures. Additionally, no activity was observed with heteroaromatic analogues that included (49 and 51−53) or lacked (48, 50, and 54) acidic hydrogens to mimic the amide NH. Similarly, the anilide group could not be replaced by a simple carboxylic acid (55) or a variety of amides derived from basic amines (Table 4). Thus, benzylic amides (56 and 57) and a series of amides derived from acyclic amines (58−62) showed EC50 values >50 μM. Tertiary amides derived from cyclic (63 and 64) or bicyclic (65−67) amines were also inactive. Parallel efforts focused on modifying the tyramine side chain (Table 5). These studies revealed a remarkably sharp SAR. The methoxy group on the phenyl ring could not be removed (68) or moved to the 3- or 2-position (69 and 70, respectively). Neither could it be replaced with a halogen (71−73), an ethoxy group (74), or a methyl group (75). A 3,4-dimethoxy phenyl ring was also not active (76). By 5201



Article

Table 4. Attempted Replacement of the Anilide Group

Table 6. Variation of the Linker

a

EC50 values calculated from 12-point dose−response study, in triplicate, following 4 days in cell culture.

Table 5. Variation of the Tyramine Side Chain

compd

R1

R2

R3

EC50 vs H2122 (μM)a

9 68 69 70 71 72 73 74 75 76 77 78 79

OCH3 H H H Br Cl F OEt CH3 OCH3 OH H H

H H OCH3 H H H H H H OCH3 H OH H

H H H OCH3 H H H H H H H H OH

0.22 >50 >50 >50 >50 >50 48 >50 >50 >50 0.26 2.40 1.72

a


generated urea 86, which was inactive. We reasoned that extending the methylene linker might maintain the spacing between the two aromatic rings, and indeed the extended compound, 87, showed modest activity. Low-micromolar EC50’s were observed with a simple amide linkage (88−90 and 93), but these were significantly less active than the initial hit (9). We were excited to discover that the oxalic acid diamide linker could be replaced with an N-acyl urea. Coupling tyramine to a benzoic acid derivative provided an analogue, 94, that was >10-fold more potent than its isomer, 95. The high activity of N-acyl urea 94 provided the starting point for additional optimization efforts. The SAR of the N-acyl ureas mirrored what was observed in the oxalamide series (Table 7). Thus, a variety of nonpolar substituents on the 4-position of benzamide ring, including halogens (96) and a phenyl ring (97), had minimal effects on activity. A 4-methoxy group was modestly deactivating (98), whereas substantial loss of activity was observed with larger polar groups, such as an ester (99), a carboxylic acid (100), or tertiary amides (101 and 102). By contrast, we observed improved activity when a ketone (103) or ether (104−106)

a


contrast, the para-substituted phenol (77) group retained full activity, whereas the 3- and 2-hydroxyl variants (78, 79) were approximately 10-fold less active against H2122 cells. The final section of the oxalamide cytotoxins to be evaluated was the linking group between the anilide and the methoxyphenyl ring (Table 6). Shortening the linker as in oxalamide 80 provided an inactive compound. Likewise, introducing a methyl group at either position on the ethylene linker (81 and 82) or on the tyramine nitrogen (83) resulted in at least a 10-fold loss in activity. Similarly, removing a carbonyl oxygen to provide α-aryloxy amide 84 or α-amino amide 85 was not successful. Removing the carbonyl CO 5202



Article

Table 7. N-Acyl Ureas

Following elution from the beads and purification with SDSPAGE, mass spectrometry revealed that both the 30 and 37 kDa bands were SCD.25 Functional studies using microsomes derived from H2122 cells showed that active oxalamides inhibited SCD’s enzymatic function. Moreover, we confirmed that toxicity was a result of unsaturated fatty acid deficiency in the presence of oxalamides. Thus, inhibition of SCD starved cells of the unsaturated fatty acids needed to maintain membrane fluidity and function. All dividing cells require SCD activity, so the selective toxicity does not result from differential expression or requirement of SCD. Rather the selectivity observed with the oxalamides and N-acyl ureas was found to result from the expression of CYP4F11 enzymes in sensitive cells lines. This metabolic enzyme can demethylate the 4-methoxy group found on all selective compounds to generate an active inhibitor. To summarize, the 4-methoxy forms of the oxalamides and N-acyl ureas act as prodrugs. CYP-mediated demethylation reveals the free phenol, which functions as a covalent and irreversible inhibitor of SCD. The selectivity observed with this series emerges from differential expression of CYP4F11; the hydroxy forms of the drugs are universally toxic. The significance of the CYP-mediated activation emerges from the on-target toxicity associated with known SCD inhibitors. As described above, inhibition of SCD in sebocytes results in hair loss, thermal dysregulation, and dry eyes in mice. CYP4F11 is not expressed in sebocytes, offering the possibility of inhibiting SCD within tumors while sparing sebocytes. Initial studies from our group provide some support for this hypothesis.18,21 Although the requirement for tumor-mediated activation of the oxalamide SCD inhibitors provides a potential means to enhance their therapeutic index, it complicates the interpretation of our structure−activity relationship. In particular, it was unclear if nontoxic analogues were unable to be metabolized by CYP4F11 or if the resulting metabolite was unable to inhibit SCD. To address this question, we compared the SCD inhibitory activity and toxicity toward H2122 cells for a series of drug (−OH form) and prodrug (−OCH3 form) analogues (Table 8). Three pairs of compounds were tested in which the −OCH3 form was inactive against H2122 cells (114 vs 58, 115 vs 80, and 119 vs 48), and in all three cases, the −OH form was also inactive. For these analogues, the −OH forms showed little to no SCD inhibitory activity. Additionally, we tested a series of moderately active cytotoxins (EC50 = 0.5−5 μM), including the pairs 116 and 88, 117 and 81, 118 and 82, and 120 and 121, using an in vitro SCD assay. In all of these cases, the activity against H2122 cells of the −OH and −OCH3 forms was similar in each case. Moreover, the −OH forms generally showed only modest inhibition of SCD activity at 100 nM in an in vitro assay. The lack of SCD inhibition by the simple amide 116 may suggest that the micromolar toxicity observed with 116 and 88 does not arise from SCD inhibition (see below). Finally, a pair of N-acyl ureas (122 and 106) and oxalamides (123 and 15) were potent as both the −OCH3 forms and the respective −OH forms showed nearly complete SCD inhibition under the assay conditions. We used an isogenic-cell-culture system to precisely determine the relationship between compound metabolism, toxicity, and SCD inhibition. We engineered H1155 cells (which do not normally express CYP4F11) to stably express either a vector control (pLVX) or CYP4F11. To determine the impact of CYP4F11, we compared the stability and toxic

a


was incorporated in the 4-position. The analogous benzylic alcohol (107) and benzylic amine (108) retained activity, but they were less potent than the ethers, 104−106. As described previously (see Table 1), the oxalamide series tolerated substitution in the 3-position of the aniline ring. However, in the N-acyl urea series, substitution in the analogous 3-position returned inactive analogues (109 and 110). The two series overlapped, however, in that N-methylation on the linker resulted in substantial loss of activity (111−113). During the course of our optimization efforts, we used propargyl ether 15 to identify SCD as the functional target of the oxalamides and N-acyl ureas (see Figure 1, above). Specifically, as previously described,21 compound 15 was first incubated with H2122 cells, which were then lysed. The terminal alkyne of 15 was used as a click tag to introduce a fluorescent dye onto compound-bound proteins. The labeled lysates were resolved on a denaturing gel and visualized with a fluorescence reader, which revealed that oxalamide 15 formed covalent adducts with two major proteins. Subsequent competition experiments indicated the biological relevance of those binding partners. Next, we used a biotin-azide to immobilize the binding partners on streptavidin-coated beads. 5203



Article

Table 8. Comparison of Phenols and Methyl Ethers

compd

drug or prodruga

EC50 H2122 (μM)b

77 9 114 58 115 80 116 88 117 81 118 82 119 48 120 121 122 106 123 15

drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug drug prodrug

0.26 0.22 >50 >50 >50 >50 2.7 0.76 4.0 3.7 5.1 3.5 >50 >50 1.5 0.32 0.011 0.010 0.058 0.028

% SCD inhibition at 100 nM

0 0 0 28 24 31

EC50 H1155 (μM)b (+oleate)c

EC50 H1155+Cyp4F11 (μM)b,d (+oleate)c

0.39 (37) 36 (>50) >50 (>50) >50 (>50) 39 (17) >50 (>50) 6.3 (>50) 3.4 (>50) >50 (>50) >50 (>50) 11.7 (10) 9.5 (12) 18 (46) >50 (>50)

0.37 (21) 0.67 (>50) >50 (>50) >50 (>50) 12 (13) >50 (>50) 4.9 (>50) 2.9 (>50) >50 (>50) >50 (>50) 11.2 (13) 16.4 (7.6) 17 (35) >50 (>50)

0.007 (12) >50 (>50) 0.008 (>50) 8.5 (16)

0.051 (6.4) 0.034 (3.6) 0.21 (>50) 0.12 (>50)

stability in H1155 (t1/2, h)

stability in H1155+Cyp4F11 (t1/2, h)d

murine S9 stability (t1/2, min) 71 7

>48

4.7

>48

>48

>48

7.5

>48

28

57 96 >98e

>240 158 13 126

Drug indicates the −OH-containing analogue; prodrug indicates the −OCH3-containing analogue. bEC50’s calculated from 12-point dose− response study, in triplicate, following 4 days in cell culture. 95% confidence intervals are provided in the Supporting Information. cValues in parentheses refer to EC50’s obtained when the cell-culture media was supplemented with 10 μM sodium oleate. dH1155 cell line stably expressing Cyp4F11. e3 μM. a

effects of different analogues in these two cell lines. In addition, we tested both conditions in the presence and absence of oleate supplementation. Compounds rescued by oleate supplementation were considered inhibitors of SCD. Finally, as a control, we confirmed that there were no major differences in cell accumulation that could explain the differences in toxicity that we observed.26 Results with analogue 106 are representative (Figure 3). This compound shows complete toxicity toward H2122 cells with an EC50 of 10 nM. By contrast, it shows little toxicity toward H1155 cells expressing an empty vector (H1155 pLVX). However, 106 is toxic to H1155 cells ectopically expressing CYP4F11 with an EC50 of 34 nM. In the presence of 10 μM oleate, the EC50 is shifted 100-fold to 3.6 μM. The incomplete killing could be due to variable expression of CYP4F11, but the clear rescue by oleate confirms that the activity is on target. To confirm that the

Figure 3. Dose−response curves for H2122 and H1155 cells. H1155 Cyp4F11 cells stably express CYP4F11, and H1155 pLVX cells contain the empty vector.

5204



■

Article

CHEMISTRY The oxalamide scaffold was synthesized through two amine additions to ethyl chlorooxoacetate (Scheme 1A). In the first

ectopically expressed CYP4F11 is active, we monitored the stability of compound 106 in the presence of H1155 cells that did and did not express CYP4F11. As expected, 106 was stable for over 48 h in the presence of H1155 control cells, but it was metabolized relatively rapidly in the presence of H1155 cells stably expressing CYP4F11. As further confirmation of our model, we observed the formation of the des-methyl form of N-acyl urea 106 (i.e., 122) only in the presence of H1155 cells expressing CYP4F11 (Figure S1). Consistently, the drug form, phenol 122, was potently toxic to H1155 irrespective of CYP4F11 status, and the dose−response curve was rightshifted at least 100-fold in the presence of oleate. All of the compounds with submicromolar potencies fit the model described for 106 and 122. For example, the initial hit, 9, is only toxic in the presence of CYP4F11, and that toxicity is rescued by oleate. Similar results are obtained with the optimized analogue, 15. By contrast, the weak potency (10−40 μΜ) associated with 115, 118, and 119 may not be related to SCD inhibition; their activity is not affected by CYP4F11 expression in H1155 cells, and the weak toxicity is not rescued by oleate. The moderate activity of 116 (∼5 μM) is currently unresolved. It did not show inhibition of SCD, and its activity is not dependent on CYP4F11 expression. Nonetheless, its modest toxicity was rescued by oleate. It is possible that 116 and 88 target a different step in the UFA synthesis pathway. Taken together, the results shown in Table 8 indicate that in sensitive cell lines (e.g., H2122), CYP-mediated demethylation is not dominating the structure−activity relationship with regard to cytotoxicity. In other words, inactive compounds with the p-methoxyphenyl ethylamine groups are likely inactive because the −OH form is not an efficient inhibitor of SCD not because demethylation by CYP4F11 fails to occur. This observation holds true for changes to the linker length and substitution pattern (115−118) and the right-hand aryl ring as drawn (114 and 119). In the case of the benzyl amide, 115, CYP-mediated demethylation appears retarded, but this appears irrelevant because the drug form (80) is nontoxic. Finally, the data in Table 8 confirm that potent SCD inhibition is required for potent cytotoxicity, as demonstrated by the correlation between SCD inhibition and EC50 against H2122 cells. All active SCD inhibitors from the oxalamide and N-acyl urea series contained a 4-substituted phenol subunit. Furthermore, prior work from our group revealed that these inhibitors formed covalent adducts with SCD. Although we do not understand the nature of the chemistry involved in forming these adducts, we have made some informative observations. First, these SCD inhibitors are not highly reactive. For example, N-acyl urea 122 shows excellent stability in the presence of murine liver S9 fractions (t1/2 > 4 h). Second, NADPH is required for both SCD enzymatic activity and for generating the adduct. These observations suggest that the catalytic activity of SCD may be required for covalent-adduct formation. Third, the oxalamides form covalent adducts with two other proteins besides SCD: heme-oxygenase 2 and CYP4F11. Cross-links to the CYP enzyme are only observed when it is overexpressed in H293T cells. Of note, all three of these enzymes are iron-containing oxygenases. We speculate that the phenol moiety of active SCD inhibitors could be oxidized to a phenolic radical, which could then react with amino acid side chains within the active site.27

Scheme 1. Synthesis of Oxalamides

step, typically anilines were condensed with ethyl chlorooxoacetate. Nucleophilic addition to the resulting aminooxoacetate, 124, at elevated temperatures then yielded the oxalamides. Usually, the desired oxalamide began to precipitate within five minutes. With this procedure, aminoheterocycles and anilines, as well as primary and secondary amines were introduced on the right-hand side as drawn. To provide comparisons between the methoxyphenyl and hydroxyphenyl derivatives (see Table 8), the methyl groups of several derivatives were removed with BBr3 (Scheme 1B). Alternatively, to modify the arylethylamines, a nitroaldol condensation reaction between aldehydes and nitroalkanes was used to generate nitroalkene 126, which could be diverted down several paths (Scheme 1C). First, hydrogenation with palladium on carbon under acid conditions formed amine salt 127. Second, nitroalkene 126 underwent conjugate addition with methylmagnesium bromide to yield nitroalkane 128.28 Finally, the α-methyl amine, 131, was synthesized through a similar procedure with nitroethane, followed by reduction with LiAlH4 (Scheme 1D). To synthesize oxalamides for biological-target identification, we synthesized oxalamide 15, which contains a benzophenone and propargyl ether to be used as a photo-cross-linking moiety and to undergo click-chemistry conjugation with an azide, respectively (Scheme 2). 132, a known29 methoxybenzophenone, was hydrolyzed by refluxing concentrated hydrobromic acid in acetic acid and then subjected to etherification with 5205



Article

amide 88, was synthesized through an EDC−amide coupling with 4-chloroaniline and 3-(4-hydroxyphenyl)propionic acid (Scheme 3E). Lastly, substitution of 4-chlorobenzoyl chloride with different lengths of 4-methoxyphenyl alkyl amines yielded benzamides 91−93. The oxalic acid diamide linker was replaced with an N-acyl urea as shown in Scheme 4. Thus, addition of oxalyl chloride to an amide yielded isocyanate 138, which underwent nucleophilic addition with phenethylamines to form a variety of Nacyl ureas (Scheme 4A).30 N-acyl urea regioisomer 95 was prepared utilizing the same isocyanation and addition steps with 4-chloroaniline and 3-(4-methoxyphenyl)propionic acid (Scheme 4B). Next, several N-acyl ureas were further modified to provide additional inhibitors. For example, urea 99 formed the basis for installing different amides through sequential hydrolysis (100) and amide couplings (101 and 102). Additionally, N-acyl urea 103 underwent reduction to form alcohol 107. Then, chlorination, azidation, and reduction yielded amine 108 (Scheme 4D). Lastly, N-methylation of Nacyl urea 103 provided monomethylated and dimethylated ureas 112 and 113, depending on the base utilized.

Scheme 2. Synthesis of 15

■

propargyl bromide to install the alkyne. Subsequent nitro reduction followed by addition to ethyl chlorooxoacetate and the addition of 4-methoxyphenethylamine yielded oxalamide 15. To evaluate replacements of the linking group between the anilide and the methoxyphenyl ring, several routes were devised (Scheme 3). First, to remove a carbonyl oxygen from the oxalic acid linker, 4-methoxyphenethylamine was acylated with bromoacetyl bromide to form bromoacetamide 136, which underwent nucleophilic substitution with either 4chlorophenol or 4-chloroaniline (see 84 and 85). Second, analogues lacking one carbonyl CO were assembled from carbamate 137 and 4-methoxyphenyl-alkyl amines of varying lengths (Scheme 3B). A third set of analogues consisting of simple amides with extended methylene linkers originated from the corresponding carboxylic acids (Scheme 3C). Similarly, EDC couplings provided amides 48−54, derived from different heteroaryl carboxylic acids (Scheme 3D). Alternatively, compound 116, which is a phenol version of

CONCLUSIONS Oxalamide 9 was originally identified from an unbiased phenotypic screen for compounds that showed toxicity toward a subset of NSCLC cell lines. Subsequent optimization efforts showed that the tyramine chain is required for activity and that the p-OCH3 is cleaved by CYP4F11 to reveal the active phenol. In turn, the phenol serves as an irreversible inhibitor of SCD, perhaps through the formation of reactive phenolic radicals. The ethylene linker of 9 could not be substituted, shortened, or lengthened, and the two amide groups appeared to be required. However, the oxalic acid diamide linker could be replaced with a more stable N-acyl urea. This change, in turn, allowed us to replace the aniline ring with a benzoic acid derivative. A variety of small substituents were accommodated on the right-hand (as drawn) aromatic ring. The most successful substitutions included modestly polar groups at the 4-position (ketone or ether) connected to aromatic rings. These inhibitors were cytotoxic to H2122 cells with EC50

Scheme 3. Varying the Linker between the Anilide and the 4-Methoxyphenyl Ring

5206



Article

Scheme 4. Synthesis of N-Acyl Ureas

General Procedure 1, for Synthesis of Oxalamides, As Exemplified by the Synthesis of N1-(4-Methoxyphenethyl)-N2phenyloxalamide (10). Step 1. A solution of ethyl chlorooxoacetate (0.15 mmol) in anhydrous CH2Cl2 (0.86 M) was added dropwise slowly to a solution of 4-chloroaniline (0.15 mmol) and Et3N (0.3 mmol) in anhydrous CH2Cl2 (0.1 M). The reaction was stirred at least 3 h at room temperature, typically overnight. Once complete, the reaction was diluted with CH2Cl2, washed with saturated NaHCO3, dried over Na2SO4, filtered, and concentrated under reduced pressure to yield ethyl 2-((4-chlorophenyl)amino)-2-oxoacetate as a white solid, which was used immediately without purification. Step 2. 4-Methoxyphenethylamine (0.75 mmol) was added to a solution of the ethyl oxalate from Step 1 (0.15 mmol) in anhydrous EtOH (0.1 M). The solution was heated at 80 °C for 3−5 h. A precipitate formed immediately. After cooling to rt, the precipitate was filtered and washed with EtOH before drying under vacuum to give the final product as a white solid, 18.0 mg, 31% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.61 (br s, 1H), 9.01 (t, J = 5.7 Hz, 1H), 7.81 (d, J = 8.0 Hz, 2H), 7.38−7.32 (m, 2H), 7.16−7.11 (m, 3H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.42−3.37 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 158.6, 157.7, 137.6, 130.9, 129.6, 128.7, 124.5, 120.3, 113.8, 55.0, 40.8, 33.7. ESIMS (m/z): 299.0 [M + H]+. N1-(4-Fluorophenyl)-N2-(4-methoxyphenethyl)oxalamide (11). The title compound was prepared according to General Procedure 1 to provide 63.7 mg of a white solid, 75% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.72 (br s, 1H), 9.00 (t, J = 6.0 Hz, 1H), 7.87−7.81 (m, 2H), 7.22−7.16 (m, 2H), 7.16−7.11 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.7, 158.7 (d, 1JCF = 241.3 Hz), 158.5, 157.7, 134.1 (d, JCF = 2.7 Hz), 130.9, 129.6, 122.2 (d, JCF = 8.1 Hz), 115.3 (d, JCF = 22.3 Hz), 113.8, 55.0, 40.8, 33.7. ESI-MS (m/z): 315.2 [M − H]−. N1-(4-Cyanophenyl)-N2-(4-methoxyphenethyl)oxalamide (12). The title compound synthesized according to General Procedure 1 to provide a white solid, 14.6 mg, 30% isolated yield. 1H NMR (400 MHz, DMSO-d6) δ 11.06 (br s, 1H), 9.09 (t, J = 5.7 Hz, 1H), 8.03 (d, J = 8.6 Hz, 2H), 7.83 (d, J = 8.6 Hz, 2H), 7.13 (d, J = 8.4 Hz, 2H), 6.85 (d, J = 8.4 Hz, 2H), 3.71 (s, 3H), 3.44−3.36 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.3, 159.2, 157.7, 141.9, 133.2, 130.9, 129.6, 120.5, 118.9, 113.8, 106.3, 55.0, 40.9, 33.6. ESI-MS (m/z): 323.9 [M + H]+. N1-(4-Benzoylphenyl)-N2-(4-methoxyphenethyl)oxalamide (13). The title compound was synthesized according to General Procedure 1 to provide a white solid, 42.1 mg, 69% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.99 (br s, 1H), 9.09 (t, J = 5.6 Hz, 1H), 8.02 (d, J = 8.0 Hz, 2H), 7.74 (dd, J = 17.9, 8.0 Hz, 4H), 7.70−7.64 (m, 1H), 7.56 (t, J = 7.5 Hz, 2H), 7.14 (d, J = 8.0 Hz, 2H), 6.86 (d, J = 8.0 Hz, 2H), 3.71 (s, 3H), 3.46−3.38 (m, 2H), 2.77 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 194.7, 159.5, 159.1, 157.7, 141.7, 137.3, 132.6, 132.5, 130.9, 130.8, 129.6, 129.5, 128.5, 119.9, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 402.9 [M + H]+. N1-(4-(4-Methoxybenzoyl)phenyl)-N2-(4-methoxyphenethyl)oxalamide (14). Step 1. A solution of tin(II) chloride dihydrate (35 mmol) in concentrated HCl (1.1 M) was added dropwise over 20 min to a suspension of 4-methoxyphenyl(4-nitrophenyl)methanone29 (132, 11.7 mmol) in 1,2-dimethoxyethane (1.1 M) and ethanol (1 M). The solution was stirred at room temperature for 1 h. Then, the mixture was poured into ice water (200 mL) and CH2Cl2 (100 mL) with stirring. NaOH (15 g) in 50 mL of water was added slowly with stirring. The mixture was filtered to break the emulsion. The aqueous layer was back-extracted two times with CH2Cl2, and the organic extracts were dried over MgSO4, filtered, and concentrated under reduced pressure. The residue was recrystallized in benzene/hexanes to yield (4-aminophenyl)(4-methoxyphenyl)methanone31 as a lightorange solid, 1.43 g, 54% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.78−7.74 (m, 2H), 7.71−7.66 (m, 2H), 6.97−6.93 (m, 2H), 6.70− 6.66 (m, 2H), 4.10 (br s, 2H), 3.88 (s, 3H). ESI-MS (m/z): 228.0 [M + H]+.

values 95% as judged by HPLC analysis (UV detection at 210 nM) except as noted. Specifically, several oxalamides contained tyramine dimer 57, which is formed from 4-methoxyphenethylamine and ethyl chlorooxoacetate and is difficult to remove via purification methods. However, it is inactive against H2122 cells. The presence of this impurity is indicated below. Chemical shifts, δ, are in parts per million, and spectra are referenced using the residual solvent peak. The following abbreviations are used: singlet (s), doublet (d), triplet (t), quartet (q), double doublet (dd), quintet (quint), multiplet (m), and broad signal (br s). For signals with multiple coupling patterns, the coupling constants are listed in the same order as the pattern (e.g., for “dt, J = 2.0, 4.0”, 2.0 is the coupling constant for the doublet, and 4.0 is for the coupling constant for the triplet). Mass spectra (m/z) were recorded on an Agilent LC-MS 1100 or 1290 Infinity using ESI ionization or on an Agilent GC-MS 7820A/5975 using EI ionization. All chemicals were used as received unless otherwise noted. 5207



Article

Step 2. Oxalamide 14 was synthesized using the amine from Step 1 following General Procedure 1 to yield the product as a white solid, 7 mg, 5% isolated yield. 1H NMR (400 MHz, DMSO-d6) δ 10.95 (br s, 1H), 9.06 (t, J = 6.0 Hz, 1H), 8.02−7.98 (m, 2H), 7.77−7.69 (m, 4H), 7.17−7.12 (m, 2H), 7.12−7.07 (m, 2H), 6.88−6.82 (m, 2H), 3.86 (s, 3H), 3.72 (s, 3H), 3.45−3.38 (m, 2H), 2.78 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 193.4, 162.8, 159.5, 159.0, 157.7, 141.2, 133.4, 132.1, 130.9, 130.5, 129.62, 129.59, 119.7, 113.9, 113.8, 55.6, 55.0, 40.9, 33.7. ESI-MS (m/z): 432.9 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(4-(4-(prop-2-yn-1-yloxy)benzoyl)phenyl)oxalamide (15). Step 1. (4-Hydroxyphenyl)(4-nitrophenyl)methanone29 (133, 1.1 mmol) and K2CO3 (1.5 mmol) were heated at reflux in acetone (0.2 M) for 30 min before propargyl bromide (80 wt % in toluene, 2.2 mmol) was added. After 2 h, the reaction was complete, as judged by TLC analysis, and the reaction mixture was cooled to rt and concentrated under reduced pressure. The residue was partitioned between water and EtOAc (5 mL). The layers were separated, and the aqueous fraction was extracted with EtOAc (2 × 5 mL). The combined organic extracts were washed with brine, dried with Na2SO4, filtered, and concentrated under reduced pressure to give (4-nitrophenyl)(4-(prop-2-yn-1-yloxy)phenyl)methanone (134) as a yellow solid, 287.5 mg, 93% yield, which was used without further purification. 1H NMR (400 MHz, DMSO-d6) δ 8.39−8.35 (m, 2H), 7.95−7.90 (m, 2H), 7.82−7.77 (m, 2H), 7.19−7.15 (m, 2H), 4.95 (d, J = 2.4 Hz, 2H), 3.66 (t, J = 2.4 Hz, 1H). ESI-MS (m/z): 282.1 [M + H]+. Step 2. A solution of tin(II) chloride dihydrate (22 mmol) in concentrated HCl (0.39 M) was added dropwise over 20 min to a solution of the nitroarene from Step 1 (7.3 mmol) in 1,2dimethoxyethane (0.39 M) and EtOH (0.33 M). After being stirred overnight, the reaction mixture was poured into ice water, and CH2Cl2 was added with stirring until the ice melted. The aqueous layer was neutralized with 7.5 M NaOH, and the solution was extracted with CH2Cl2 (3×). The organic layer was dried with Na2SO4 and concentrated under reduced pressure to yield (4aminophenyl)(4-(prop-2-yn-1-yloxy)phenyl)methanone (135) as an orange oil, 654.4 mg, 35% yield. 1H NMR (500 MHz, DMSO-d6) δ 7.65−7.61 (m, 2H), 7.52−7.49 (m, 2H), 7.11−7.07 (m, 2H), 6.62− 6.59 (m, 2H), 4.90 (d, J = 2.4 Hz, 2H), 3.65 (t, J = 2.4 Hz, 1H). ESIMS (m/z): 252.0 [M + H]+. Step 3. The oxalamide 15 was synthesized using the amine from Step 2 following General Procedure 1 to yield the product as a creamcolored solid, 385.7 mg, 32% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.06 (t, J = 5.8 Hz, 1H), 8.03−7.98 (m, 2H), 7.78−7.70 (m, 4H), 7.17−7.12 (m, 4H), 6.88−6.83 (m, 2H), 4.93 (d, J = 2.2 Hz, 2H), 3.72 (s, 3H), 3.65 (t, J = 2.2 Hz, 1H), 3.45−3.38 (m, 2H), 2.78 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 193.4, 160.7, 159.5, 159.0, 157.8, 141.2, 133.3, 131.9, 130.9, 130.5, 130.3, 129.6, 119.7, 114.7, 113.8, 94.6, 78.8, 55.8, 55.0, 40.8, 33.7. ESI-MS (m/z): 456.9 [M + H]+. HRMS (ESI) m/z: calcd for C27H24N2O5 (M + H)+ 457.175 80, found 457.1750. N 1-(4-(4-Hydroxybenzoyl)phenyl)-N 2-(4-methoxyphenethyl)oxalamide (16). Step 1. A solution of tin(II) chloride dihydrate (1.2 mmol) in concentrated HCl (0.39 M) was added dropwise over 20 min to a solution of (4-hydroxyphenyl)(4-nitrophenyl)methanone29 (133, 0.41 mmol) in 1,2-dimethoxyethane (0.39 M) and EtOH (0.33 M). After being stirred overnight, the reaction mixture was poured into ice water, and CH2Cl2 was added with stirring until the ice melted. The aqueous layer was neutralized with 7.5 M NaOH, and the solution was extracted with CH2Cl2 (3×). The organic extracts were dried over Na2SO4, filtered, and concentrated under reduced pressure to yield (4-aminophenyl)(4-hydroxyphenyl)methanone (S1) as a solid, 50.1 mg, 57% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.14 (br s, 1H), 7.57−7.51 (m, 2H), 7.51−7.45 (m, 2H), 6.87−6.82 (m, 2H), 6.62−6.57 (m, 2H). ESI-MS (m/z): 214.0 [M + H]+. Step 2. The oxalamide 16 was synthesized using the amine from Step 1 following General Procedure 1 with purification by flash chromatography over silica gel (0 to 100% EtOAc/hexanes) to give a cream-colored solid, 19.8 mg, 20% yield, contaminated with 23% dimer 57. 1H NMR (500 MHz, DMSO-d6) δ 10.96 (br s, 1H), 10.42

(br s, 1H), 9.09 (t, J = 6.0 Hz, 1H), 8.00−7.97 (m, 2H), 7.71−7.67 (m, 2H), 7.67−7.63 (m, 2H), 7.16−7.13 (m, 2H), 6.91−6.88 (m, 2H), 6.87−6.85 (m, 2H), 3.71 (s, 3H), 3.43−3.38 (m, 2H), 2.77 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 193.2, 161.8, 159.6, 159.0, 157.7, 140.9, 133.7, 132.4, 130.9, 130.3, 129.6, 128.1, 119.7, 115.2, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 418.9 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(2-methoxyphenyl)oxalamide (17). The title compound was prepared according to General Procedure 1 to provide 31.2 mg of a white solid, 35% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.77 (br s, 1H), 9.15 (t, J = 6.0 Hz, 1H), 8.17 (dd, J = 8.1, 1.3 Hz, 1H), 7.19−7.08 (m, 4H), 7.02−6.96 (m, 1H), 6.88−6.83 (m, 2H), 3.89 (s, 3H), 3.71 (s, 3H), 3.42−3.35 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.5, 157.7, 157.3, 148.7, 130.8, 129.6, 125.7, 125.2, 120.6, 119.3, 113.8, 111.2, 56.0, 55.0, 41.0, 33.6. ESI-MS (m/z): 328.9 [M + H]+. N1-(2-Fluorophenyl)-N2-(4-methoxyphenethyl)oxalamide (18). The title compound was prepared according to General Procedure 1 to provide 44.3 mg of a white solid, 52% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.18 (br s, 1H), 9.06 (t, J = 5.9 Hz, 1H), 7.70 (td, J = 7.9, 1.6 Hz, 1H), 7.34−7.19 (m, 3H), 7.17−7.11 (m, 2H), 6.88−6.83 (m, 2H), 3.72 (s, 3H), 3.43−3.35 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13 C NMR (100 Hz, DMSO-d6) δ 159.3, 158.5, 157.7, 154.8 (d, 1JCF = 247.1 Hz), 130.9, 129.6, 127.1 (d, 3JCF = 7.7 Hz), 125.2, 124.6 (d, JCF = 3.7 Hz), 124.5 (d, JCF = 4.4 Hz), 115.8 (d, 2JCF = 19.5 Hz), 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 317.1 [M + H]+. N1-(2-Bromophenyl)-N2-(4-methoxyphenethyl)oxalamide (19). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel (0 to 30% EtOAc/hexanes) to give a white solid, 34.4 mg, 30% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.12 (br s, 1H), 9.18 (t, J = 6.0 Hz, 1H), 7.98 (dd, J = 8.1, 1.6 Hz, 1H), 7.72 (dd, J = 8.1, 1.4 Hz, 1H), 7.44 (td, J = 7.7, 1.4 Hz, 1H), 7.18 (td, J = 7.7, 1.6 Hz, 1H), 7.16−7.12 (m, 2H), 6.88−6.84 (m, 2H), 3.71 (s, 3H), 3.42−3.36 (m, 2H), 2.77 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.3, 158.1, 157.7, 134.8, 132.7, 130.8, 129.6, 128.5, 127.2, 123.6, 116.0, 113.8, 55.0, 41.1, 33.6. ESI-MS (m/z): 376.9 [M + H]+. N1-(3-Chlorophenyl)-N2-(4-methoxyphenethyl)oxalamide (20). The title compound was prepared according to General Procedure 1 to provide 29.1 mg of a white solid, 33% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.82 (br s, 1H), 9.03 (t, J = 5.9 Hz, 1H), 7.96 (t, J = 2.0 Hz, 1 Hz), 7.78 (dd, J = 8.2, 0.8 Hz, 1H), 7.38 (t, J = 8.1 Hz, 1H), 7.19 (dd, J = 8.1, 0.8 Hz, 1H), 7.16−7.11 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.5, 158.9, 157.7, 139.2, 133.0, 130.9, 130.4, 129.6, 124.2, 119.8, 118.8, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/ z): 333.1 [M + H]+. N1-(3-Ethoxyphenyl)-N2-(4-methoxyphenethyl)oxalamide (21). The title compound was synthesized according to General Procedure 1 to give a white solid, 40.5 mg, 83% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.53 (br s, 1H), 8.98 (t, J = 6.0 Hz, 1H), 7.46 (dd, J = 2.2, 2.2 Hz, 1H), 7.40 (ddd, J = 8.2, 2.2, 0.9 Hz, 1H), 7.22 (dd, J = 8.2, 8.2 Hz, 1H), 7.16−7.12 (m, 2H), 6.87−6.83 (m, 2H), 6.69 (ddd, J = 8.2, 2.2, 0.9 Hz, 1H), 3.99 (q, J = 7.0 Hz, 2H), 3.71 (s, 3H), 3.43− 3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H), 1.32 (t, J = 7.0 Hz, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 158.6, 158.5, 157.7, 138.7, 130.9, 129.6, 129.5, 113.8, 112.4, 110.4, 106.6, 62.9, 55.0, 40.8, 33.7, 14.6. ESI-MS (m/z): 343.0 [M + H]+. N1-(3-Cyanophenyl)-N2-(4-methoxyphenethyl)oxalamide (22). The title compound was synthesized according to General Procedure 1 to give a white solid, 20.2 mg, 23% yield. 1H NMR (400 MHz, DMSO-d6) δ 11.00 (br s, 1H), 9.07 (t, J = 6.0 Hz, 1H), 8.25−8.23 (m, 1H), 8.13 (dt, J = 7.7, 1.9 Hz, 1H), 7.62−7.55 (m, 2H), 7.16− 7.11 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.44−3.37 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.3, 159.1, 157.7, 138.6, 130.9, 130.2, 129.6, 128.0, 125.0, 123.2, 118.6, 113.8, 111.5, 55.0, 40.9, 33.7. ESI-MS (m/z): 324.1 [M + H]+. N 1 -(3-Chloro-5-fluorophenyl)-N 2 -(4-methoxyphenethyl)oxalamide (23). The title compound was prepared according to General Procedure 1 to provide 30.8 mg of a white solid, 33% yield. 1 H NMR (400 MHz, DMSO-d6) δ 9.07 (t, J = 5.9 Hz, 1 Hz), 7.85− 5208



Article

N1-(3,4-Dichlorophenyl)-N2-(4-methoxyphenethyl)oxalamide (30). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 93.3 mg, 69% yield contaminated with 5% dimer 57. 1H NMR (500 MHz, DMSOd6) δ 10.96 (br s, 1H), 9.05 (t, J = 6.0 Hz, 1H), 8.16 (d, J = 2.5 Hz, 1H), 7.84 (dd, J = 8.9, 2.5 Hz, 1H), 7.62 (d, J = 8.9 Hz, 1H), 7.15− 7.12 (m, 2H), 6.87−6.84 (m, 2H), 3.71 (s, 3H), 3.42−3.36 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.3, 159.0, 157.7, 137.9, 130.93, 130.87, 130.7, 129.6, 126.1, 121.6, 120.4, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 368.8 [M + H]+. N 1 -(4-Chloro-3-nitrophenyl)-N 2 -(4-methoxyphenethyl)oxalamide (31). The title compound was prepared according to General Procedure 1 to provide 83.0 mg of an orange solid, 82% yield. 1 H NMR (400 MHz, DMSO-d6) δ 11.12 (br s, 1H), 9.09 (t, J = 5.9 Hz, 1H), 8.62 (d, J = 2.2 Hz, 1H), 8.11 (dd, J = 8.9, 2.2 Hz, 1H), 7.77 (d, J = 8.9 Hz, 1H), 7.16−7.10 (m, 2H), 6.88−6.82 (m, 2H), 3.71 (s, 3H), 3.44−3.36 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.2, 159.1, 157.7, 147.2, 137.7, 132.0, 130.9, 129.6, 125.4, 119.9, 116.8, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 376.1 [M − H]−. N 1 -(4-Chloro-3-cyanophenyl)-N 2 -(4-methoxyphenethyl)oxalamide (32). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 34.1 mg, 66% yield. 1H NMR (400 MHz, DMSO-d6) δ 11.11 (br s, 1H), 9.08 (t, J = 6.0 Hz, 1H), 8.35 (d, J = 2.6 Hz, 1H), 8.16 (dd, J = 9.0, 2.6 Hz, 1H), 7.75 (d, J = 9.0 Hz, 1H), 7.16−7.10 (m, 2H), 6.87−6.84 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 159.15, 159.12, 157.7, 137.4, 130.9, 130.6, 129.6, 130.1, 126.5, 125.2, 115.9, 113.8, 112.0, 55.0, 40.9, 33.6. ESI-MS (m/z): 358.1 [M + H]+. N1-(4-Chloro-3-(trifluoromethyl)phenyl)-N2-(4methoxyphenethyl)oxalamide (33). The title compound was prepared according to General Procedure 1 to provide 60.6 mg of a white solid in 56% yield. 1H NMR (400 MHz, DMSO-d6) δ 11.06 (br s, 1H), 9.07 (t, J = 6.0 Hz, 1H), 8.43 (d, J = 2.5 Hz, 1H), 8.14 (dd, J = 8.8, 2.5 Hz, 1H), 7.72 (d, J = 8.8 Hz, 1H), 7.16−7.11 (m, 2H), 6.87− 6.82 (m, 2H), 3.71 (s, 3H), 3.44−3.37 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.2, 159.1, 157.7, 137.3, 132.1, 130.9, 129.6, 126.7 (q, 2JCF = 30.6 Hz), 125.3 (q, 4JCF = 1.9 Hz), 125.2, 122.7 (q, 1JCF = 273.4 Hz), 119.3 (q, 3JCF= 5.6 Hz), 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 399.1 [M − H]−. N 1 -(3-Methoxy-4-methylphenyl)-N 2 -(4-methoxyphenethyl)oxalamide (34). The title compound was prepared according to General Procedure 1 to provide 45.7 mg of a white solid, 50% yield. 1 H NMR (400 MHz, DMSO-d6) δ 10.49 (br s, 1H), 8.97 (t, J = 5.9 Hz, 1H), 7.50 (d, J = 1.7 Hz, 1H), 7.36 (dd, J = 8.0, 1.8 Hz, 1H), 7.16−7.11 (m, 2H), 7.07 (d, J = 8.2 Hz, 1H), 6.88−6.83 (m, 2H), 3.74 (s, 3H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H), 2.10 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 159.9, 158.3, 157.7, 157.1, 136.7, 130.9, 130.1, 129.6, 121.6, 113.8, 111.8, 103.1, 55.1, 55.0, 40.8, 33.7, 15.6. ESI-MS (m/z): 341.2 [M − H]−. N 1 -(4-Methoxy-3-methylphenyl)-N 2 -(4-methoxyphenethyl)oxalamide (35). The title compound was prepared according to General Procedure 1 to provide 60.4 mg of a white solid, 66% yield. 1 H NMR (400 MHz, DMSO-d6) δ 10.39 (br s, 1H), 8.91 (t, J = 6.0 Hz, 1H), 7.62−7.55 (m, 2H), 7.16−7.11 (m, 2H), 6.89 (d, J = 8.8 Hz, 1H), 6.87−6.83 (m, 2H), 3.76 (s, 3H), 3.71 (s, 3H), 3.42−3.35 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H), 2.12 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 160.0, 158.0, 157.7, 154.2, 130.9, 130.2, 129.6, 125.5, 122.9, 119.1, 113.8, 110.2, 55.4, 55.0, 40.8, 33.7, 16.2. ESI-MS (m/z): 343.1 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(pyridin-4-yl)oxalamide (36). The title compound was synthesized according to General Procedure 1 to give a white solid, 44.6 mg, 49% yield, contaminated with 21% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 9.09 (t, J = 5.9 Hz, 1H), 8.49 (dd, J = 4.8 1.5 Hz, 2H), 7.82 (dd, J = 4.8, 1.5 Hz, 2H), 7.16−7.12 (m, 2H), 6.87−6.84 (m, 2H), 3.71 (s, 3H), 3.44−3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.7, 159.2, 157.7, 150.4, 144.5, 130.9, 129.6, 114.2, 113.8, 55.0, 40.9, 33.6. ESIMS (m/z): 300.0 [M + H]+.

7.82 (m, 1H), 7.73 (dt, J = 11.1, 2.1 Hz, 1H), 7.19 (dt, J = 8.6, 2.1 Hz, 1H), 7.16−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.43− 3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSOd6) δ 162.0 (d, 1JCF = 244.7 Hz), 159.2, 159.1, 157.7, 140.3 (d, JCF = 12.8 Hz), 133.9 (d, JCF = 12.8 Hz), 130.9, 129.6, 116.1 (d, JCF = 3.1 Hz), 113.8, 111.7 (d, JCF = 25.7 Hz), 106.0 (d, JCF = 26.4 Hz), 55.0, 40.9, 33.7. ESI-MS (m/z): 351.1 [M + H]+. N1-(3-Chloro-5-(trifluoromethyl)phenyl)-N2-(4methoxyphenethyl)oxalamide (24). The title compound was prepared according to General Procedure 1 to provide 53.9 mg of a white solid, 50% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.99 (br s, 1H), 9.08 (t, J = 6.0 Hz, 1H), 8.28−8.22 (m, 2H), 7.63−7.60 (m, 1H), 7.16−7.12 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.44− 3.37 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSOd6) δ 159.3, 159.1, 157.7, 140.0, 134.2, 131.2, 130.9, 129.6, 124.5, 123.4, 120.7 (q, JCF = 4.0 Hz), 115.3 (q, JCF = 4.1 Hz), 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 399.1 [M − H]−. N1-(3,5-Dichlorophenyl)-N 2-(4-methoxyphenethyl)oxalamide (25). The title compound was prepared according to General Procedure 1 to provide 71.6 mg of a light-brown solid, 73% yield. 1 H NMR (400 MHz, DMSO-d6) δ 10.98 (br s, 1H), 9.05 (t, J = 5.9 Hz, 1H), 7.95 (app d, J = 1.9 Hz, 2H), 7.36 (app t, J = 1.9 Hz, 1H), 7.16−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.14, 159.13, 157.7, 140.1, 134.0, 130.9, 129.6, 123.7, 118.6, 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 365.1 [M − H]−. N 1 -(3-Fluoro-4-methylphenyl)-N 2 -(4-methoxyphenethyl)oxalamide (26). The title compound was prepared according to General Procedure 1 to provide 36.0 mg of a white solid, 41% yield. 1 H NMR (400 MHz, DMSO-d6) δ 10.74 (br s, 1H), 9.01 (t, J = 6.0 Hz, 1H), 7.68 (dd, J = 12.2, 1.9 Hz, 1H), 7.56 (dd, J = 8.3, 1.9 Hz, 1H), 7.24 (t, J = 8.6 Hz, 1H), 7.16−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H), 2.19 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 160.1 (d, 1JCF = 241.1 Hz), 159.6, 158.7, 157.7, 137.0 (d, JCF = 10.9 Hz), 131.4 (d, JCF = 6.4 Hz), 130.9, 129.6, 119.9 (d, JCF = 17.2 Hz), 116.0 (d, JCF = 3.1 Hz), 113.8, 106.9 (d, JCF = 27.2 Hz), 55.0, 40.8, 33.7, 13.8. ESI-MS (m/z): 329.1 [M − H]−. N 1 -(4-Chloro-3-fluorophenyl)-N 2 -(4-methoxyphenethyl)oxalamide (27). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 86.0 mg, 82% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.98 (br s, 1H), 9.05 (t, J = 6.0 Hz, 1H), 7.93 (dd, J = 11.9, 2.4 Hz, 1H), 7.72 (ddd, J = 8.9, 2.4, 1.0 Hz, 1H), 7.57 (t, J = 8.7 Hz, 1H), 7.16−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.3, 159.0, 157.7, 156.8 (J = 244.1 Hz), 138.3 (J = 10.1 Hz) 130.9, 130.6, 129.6, 117.3 (J = 3.5 Hz), 114.3 (J = 17.6 Hz), 113.8, 108.5 (J = 25.7 Hz), 55.0, 40.9, 33.7. ESI-MS (m/z): 350.9 [M + H]+. N 1 -(3-Chloro-4-fluorophenyl)-N 2 -(4-methoxyphenethyl)oxalamide (28). The title compound was prepared according to General Procedure 1 to provide 60.7 mg of a white solid, 64% yield. 1 H NMR (400 MHz, DMSO-d6) δ 10.89 (br s, 1H), 9.03 (t, J = 5.9 Hz, 1H), 8.08 (dd, J = 6.9, 2.6 Hz, 1H), 7.85−7.79 (m, 1H), 7.42 (t, J = 9.1 Hz, 1H), 7.16−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.36 (m, 2H), 2.76 (t, J = 7.40 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.4, 158.8, 157.7, 153.8 (d, 1JCF = 243.6 Hz), 135.0 (d, JCF = 3.1 Hz), 130.9, 129.6, 121.9, 120.8 (d, JCF = 7.0 Hz), 119.1 (d, 2JCF = 18.4 Hz), 116.9 (d, 2JCF = 21.7 Hz), 113.8, 55.0, 40.8, 33.7. ESI-MS (m/z): 351.1 [M + H]+. N 1 -(3-Chloro-4-methylphenyl)-N 2 -(4-methoxyphenethyl)oxalamide (29). The title compound was synthesized according to General Procedure 1 to give a white solid, 50.5 mg, 87% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.73 (br s, 1H), 8.98 (t, J = 6.0 Hz, 1H), 7.95 (d, J = 1.7 Hz, 1H), 7.67 (dt, J = 8.4, 1.7 Hz, 1H), 7.31 (d, J = 8.4 Hz, 1H), 7.16−7.12 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.35−3.28 (m, 2H, under H2O peak), 2.75 (t, J = 7.3 Hz, 2H), 2.28 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 157.7, 136.8, 132.9, 131.2, 131.1, 130.9, 129.6, 120.3, 119.0, 113.8, 55.0, 40.8, 33.7, 19.0. ESI-MS (m/z): 347.9 [M + H]+. 5209



Article

N1-(4-Methoxyphenethyl)-N2-(pyrazin-2-yl)oxalamide (37). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel (0 to 100% EtOAc/hexanes) to give a white solid, 26.0 mg, 29% yield, contaminated with 15% dimer 57. 1H NMR (500 MHz, DMSO-d6) δ 10.57 (br s, 1H), 9.24 (d, J = 1.2 Hz, 1H), 9.21 (t, J = 6.1 Hz, 1H), 8.73 (t, J = 6.0 Hz, 0.4H, dimer), 8.50−8.49 (m, 1H), 8.48 (d, J = 2.4 Hz, 1H), 7.16−7.13 (m, 2H), 7.11−7.08 (m, 0.8H, dimer), 6.88−6.85 (m, 2H), 6.84−6.82 (m, 0.5H, dimer), 3.71 (s, 3H), 3.71 (s, 1.3H, dimer), 3.42−3.37 (m, 2H), 3.33−3.28 (m, 0.9H, dimer), 2.76 (t, J = 7.5 Hz, 2H), 2.70 (t, J = 7.4 Hz, 0.8H, dimer). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 158.9, 157.7, 147.1, 143.0, 141.1, 137.0, 130.8, 129.6, 113.8, 55.0, 41.0, 33.6. ESI-MS (m/z): 301.0 [M + H]+. N1-(2-Fluoropyridin-3-yl)-N2-(4-methoxyphenethyl)oxalamide (38). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 102 mg, 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.28 (br s, 1H), 9.09 (t, J = 5.9 Hz, 1H), 8.16 (ddd, J = 10.0, 7.8, 1.8 Hz, 1H), 8.08 (dt, J = 4.9, 1.5 Hz, 1H), 7.40 (ddd, J = 7.8, 4.9, 1.2 Hz, 1H), 7.16−7.12 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.43−3.37 (m, 2H), 2.76 (t, J = 7.7 Hz, 2H). 13C NMR (400 MHz, DMSO-d6) δ 159.01, 158.97, 157.8, 155.8 (d, 1JCF = 237.2 Hz), 143.7 (d, 3JCF = 14.3 Hz), 135.7 (d, 3JCF = 3.2 Hz), 130.9, 129.6, 122.3 (d, 4JCF = 4.2 Hz), 120.0 (d, 2JCF = 27.4 Hz), 113.8, 55.0, 40.9, 33.7. ESI-MS (m/z): 318.0 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(4-methylpyridin-3-yl)oxalamide (39). The title compound was synthesized according to General Procedure 1 to give the final product as a pink solid, 68.3 mg, 73% yield, containing 22% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 10.25 (br s, 1H), 9.01 (t, J = 6.0 Hz, 1H), 8.48 (s, 1H), 8.31 (d, J = 4.9 Hz, 1H), 7.30 (d, J = 4.9 Hz, 1H), 7.17−7.12 (m, 2H), 6.88−6.84 (m, 2H), 3.72 (s, 3H), 3.43−3.37 (m, 2H), 2.77 (t, J = 7.8 Hz, 2H), 2.20 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.4, 159.1, 157.8, 147.0, 146.6, 142.2, 132.4, 130.9, 129.6, 125.4, 113.8, 55.0, 40.9, 33.7, 17.2. ESI-MS (m/z): 314.0 [M + H]+. N1-(5-Chloropyridin-3-yl)-N2-(4-methoxyphenethyl)oxalamide (40). The title compound was synthesized according to General Procedure 1 to give a cream-colored solid, 93.2 mg, 89% yield, contaminated with 26% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 9.09 (t, J = 6.0 Hz, 1H), 8.97 (d, J = 2.1 Hz, 1H), 8.40 (d, J = 2.2 Hz, 1H), 8.35 (t, J = 2.2 Hz, 1H), 7.16−7.12 (m, 2H), 6.87−6.85 (m, 2H), 3.71 (s, 3H), 3.44−3.37 (m, 2H), 2.77 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.4, 159.1, 157.7, 143.5, 140.3, 135.4, 130.9, 130.4, 129.6, 126.6, 113.8, 55.0, 43.7, 33.6. ESI-MS (m/ z): 333.9 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(5-methylpyridin-3-yl)oxalamide (41). The title compound was synthesized according to General Procedure 1 to give a cream-colored solid, 79.8 mg, 87% yield, contaminated with 24% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 9.02 (t, J = 5.9 Hz, 1H), 8.78 (d, J = 2.3 Hz, 1H), 8.19−8.18 (m, 1H), 8.04−8.01 (m, 1H), 7.16−7.12 (m, 2H), 6.87−6.85 (m, 2H), 3.71 (s, 3H), 3.44−3.37 (m, 2H), 2.80−2.73 (m, 2H), 2.29 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.4, 159.1, 157.7, 145.7, 139.4, 134.0, 132.8, 130.9, 129.6, 127.7, 113.8, 55.0, 40.8, 33.7, 18.0. ESI-MS (m/ z): 314.0 [M + H]+. N1-(6-Chloropyridin-3-yl)-N2-(4-methoxyphenethyl)oxalamide (42). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 103.4 mg, 96% yield, contaminated with 20% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 11.05 (br s, 1H), 9.07 (t, J = 6.0 Hz, 1H), 8.85 (dd, J = 2.8, 0.4 Hz, 1H), 8.26 (dd, J = 8.7, 2.8 Hz, 1H), 7.52 (dd, J = 8.7, 0.4 Hz, 1H), 7.16−7.12 (m, 2H), 6.87−6.82 (m, 2H), 3.71 (s, 3H), 3.44−3.37 (m, 2H), 2.76 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 159.2, 157.7, 144.8, 141.8, 134.1, 131.0, 130.9, 129.6, 124.2, 113.8, 55.0, 40.8, 33.6. ESI-MS (m/z): 333.9 [M + H]+. N1-(4-Methoxyphenethyl)-N2-(pyridin-2-yl)oxalamide (43). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel (0 to 100% EtOAc/hexanes) to give a yellow solid, 10.5 mg, 11% yield, contaminated with 16% dimer 57. 1H NMR (500 MHz, DMSO-d6) δ 9.19 (t, J = 6.0 Hz, 1H), 8.39 (ddd, J = 4.9, 1.9, 0.8 Hz, 1H), 8.05

(d, J = 8.3 Hz, 1H), 7.89 (ddd, J = 8.3, 7.5, 1.9 Hz, 1H), 7.23 (ddd, J = 7.5, 4.9, 0.8 Hz, 1H), 7.16−7.12 (m, 2H), 6.87−6.84 (m, 2H), 3.71 (s, 3H), 3.42−3.37 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.2, 158.3, 157.7, 149.9, 148.5, 138.7, 130.8, 129.6, 120.8, 113.8, 113.7, 55.0, 41.0, 33.6. ESI-MS (m/z): 300.0 [M + H]+. N1-(5-Chloropyridin-2-yl)-N2-(4-methoxyphenethyl)oxalamide (44). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel (0 to 100% EtOAc/hexanes) to give an orange solid, 9.1 mg, 9% yield. 1 H NMR (500 MHz, DMSO-d6) δ 9.19 (t, J = 6.0 Hz, 1H), 8.46 (dd, J = 2.6, 0.7 Hz, 1H), 8.06 (d, J = 8.8 Hz, 1H), 8.02 (dd, J = 8.8, 2.6 Hz, 1H), 7.15−7.12 (m, 2H), 6.87−6.84 (m, 2H), 3.71 (s, 3H), 3.41−3.36 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.1, 158.5, 157.7, 148.7, 146.9, 138.3, 130.8, 129.6, 126.6, 115.0, 113.8, 55.0, 41.0, 33.6. ESI-MS (m/z): 333.9 [M + H]+. N1-(5-Bromopyridin-2-yl)-N2-(4-methoxyphenethyl)oxalamide (45). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel (0 to 100% EtOAc/hexanes) to give a white solid, 13.1 mg, 12% yield. 1 H NMR (500 MHz, DMSO-d6) δ 10.22 (br s, 1H), 9.20 (t, J = 6.0 Hz, 1H), 8.53 (dd, J = 2.5, 0.4 Hz, 1H), 8.13 (dd, J = 8.8, 2.5 Hz, 1H), 8.01 (dd, J = 8.8, 0.4 Hz, 1H), 7.15−7.11 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 3.41−3.36 (m, 2H), 2.75 (t, J = 7.7 Hz, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 159.0, 158.5, 157.7, 149.1, 149.0, 141.1, 130.8, 129.6, 115.5, 115.1, 113.8, 55.0, 41.0, 33.6. ESI-MS (m/ z): 377.8 [M + H]+. N1-(6-Bromopyridin-2-yl)-N2-(4-methoxyphenethyl)oxalamide (46). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 37.2 mg, 33% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.23 (br s, 1H), 9.15 (t, J = 5.9 Hz, 1H), 8.02 (d, J = 8.1 Hz, 1H), 7.83 (t, J = 8.0 Hz, 1H), 7.46 (dd, J = 7.8, 0.4 Hz, 1H), 7.16−7.11 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.42−3.35 (m, 2H), 2.76 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 158.9, 158.7, 157.7, 150.2, 141.7, 139.1, 130.8, 129.6, 124.5, 113.8, 113.1, 55.0, 41.0, 33.6. ESI-MS (m/z): 379.8 [M + H]+. N1-(6-Fluoropyridin-2-yl)-N2-(4-methoxyphenethyl)oxalamide (47). The title compound was synthesized according to General Procedure 1 to give a white solid, 35.2 mg, 37% yield, contaminated with 16% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 9.16 (t, J = 5.9 Hz, 1H), 8.06 (dd, J = 8.1, 8.1 Hz, 1H), 7.94 (dd, J = 8.1, 2.2 Hz, 1H), 7.16−7.12 (m, 2H), 6.99 (dd, J = 8.1, 2.2 Hz, 1H), 6.88−6.84 (m, 2H), 3.71 (s, 3H), 3.42−3.35 (m, 2H), 2.76 (t, J = 7.7 Hz, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 161.4 (d, 1JCF = 237.8 Hz), 159.0, 158.7, 157.7, 148.4 (d, 3JCF = 15.2 Hz), 144.3 (d, 3JCF = 7.9 Hz), 130.8, 129.6, 113.8, 111.2 (d, 4JCF = 4.2 Hz), 105.4 (2JCF = 35.5 Hz), 55.0, 41.0, 33.6. ESI-MS (m/z): 318.0 [M + H]+. General Procedure 2, for the Synthesis of Amides with EDC or HOBt, As Exemplified by the Synthesis of N-(4-Methoxyphenethyl)quinoline-2-carboxamide (48). Quinaldic acid (0.7 mmol), 1-(3(dimethylamino)propyl)-3-ethylcarbodiimide HCl (EDC, 0.735 mmol), and 1-hydroxybenzotriazole hydrate (HOBT, 0.735 mmol) were dissolved in DMF (0.1 M), and the solution was stirred for 20 min before 4-methoxyphenethylamine (0.7 mmol) was added. The reaction mixture was stirred overnight. Water was added to the reaction mixture, and a precipitate formed. The precipitate was collected by filtration, washed with EtOAc, and dried to give the final product as a white solid, 139.7 mg, 66% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.95 (t, J = 6.0 Hz, 1H), 8.56 (d, J = 8.5 Hz, 1H), 8.15 (d, J = 8.5 Hz, 1H), 8.12 (d, J = 8.4 Hz, 1H), 8.08 (d, J = 8.2 Hz, 1H), 7.87 (ddd, J = 8.6, 6.8, 1.4 Hz, 1H), 7.72 (ddd, J = 8.4, 6.8, 1.2 Hz, 1H), 7.21−7.17 (m, 2H), 6.89−6.84 (m, 2H), 3.71 (s, 3H), 3.60− 3.53 (m, 2H), 2.85 (t, J = 7.8 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 163.8, 157.7, 150.2, 146.0, 137.9, 131.2, 130.5, 129.6, 129.1, 128.8, 128.1, 128.0, 118.6, 113.8, 55.0, 40.8, 34.3. ESI-MS (m/z): 307.0 [M + H]+. HRMS (ESI) m/z: calcd for C19H18N2O2 (M + H)+ 307.144 10, found 307.1449. N-(4-Methoxyphenethyl)-4-oxo-3,4-dihydroquinazoline-2-carboxamide (49). The title compound was synthesized according to 5210



Article

General Procedure 2 with 4-oxo-3,4-dihydroquinazoline-2-carboxylic acid32 to yield a white solid, 6.4 mg, 9% isolated yield, after triturating in CH2Cl2. 1H NMR (400 MHz, DMSO-d6) δ 8.76 (t, J = 6.0 Hz, 1H), 7.95 (dd, J = 7.9, 1.1 Hz, 1H), 7.56 (d, J = 8.1 Hz, 1H), 7.50 (ddd, J = 8.3, 6.5, 1.4 Hz, 1H), 7.21 (ddd, J = 8.2, 6.6, 1.2 Hz, 1H), 7.19−7.15 (m, 2H), 6.88−6.85 (m, 2H), 3.72 (s, 3H), 3.49−3.42 (m, 2H), 2.78 (t, J = 7.6 Hz, 2H). ESI-MS (m/z): 324.0 [M + H]+. N-(4-Methoxyphenethyl)quinoxaline-2-carboxamide (50). The title compound was synthesized according to General Procedure 2 to give the final product as a peach-colored solid, 144.1 mg, 68% yield. 1 H NMR (400 MHz, DMSO-d6) δ 9.46 (br s, 1H), 9.10 (t, J = 5.9 Hz, 1H), 8.22−8.16 (m, 2H), 8.01−7.96 (m, 2H), 7.21−7.16 (m, 2H), 6.89−6.84 (m, 2H), 3.71 (s, 3H), 3.60−3.53 (m, 2H), 2.85 (t, J = 7.8 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 163.0, 157.7, 144.4, 143.7, 142.9, 139.8, 131.9, 131.3, 131.1, 129.6, 129.4, 129.1, 113.8, 55.0, 40.9, 34.2. ESI-MS (m/z): 308.0 [M + H]+. 8-Hydroxy-N-(4-methoxyphenethyl)quinoline-2-carboxamide (51). The title compound was synthesized according to General Procedure 2 to give the final product as a red oil, 108.5 mg, 49% yield. 1 H NMR (500 MHz, DMSO-d6) δ 10.13 (br s, 1H), 9.74 (t, J = 5.9 Hz, 1H), 8.49 (d, J = 8.5 Hz, 1H), 8.13 (d, J = 8.5 Hz, 1H), 7.56 (t, J = 7.8 Hz, 1H), 7.47 (d, J = 8.2 Hz, 1H), 7.21−7.18 (m, 2H), 7.18− 7.16 (m, 1H), 6.88−6.84 (m, 2H), 3.71 (s, 3H), 3.60−3.54 (m, 2H), 2.86 (t, J = 7.9 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 163.6, 157.7, 153.6, 147.5, 137.7, 136.4, 131.2, 129.6, 129.44, 129.35, 118.8, 117.5, 113.8, 111.5, 55.0, 40.8, 34.5. ESI-MS (m/z): 323.0 [M + H]+. 1-Mydroxy-N-(4-methoxyphenethyl)-2-naphthamide (52). The title compound was synthesized according to General Procedure 2, except no precipitate formed. Water and EtOAc were added to the reaction mixture, and the organic layer was washed with 1 M NaOH (3×), water (3×), 1 M HCl, water (3×), and brine. The organic extracts were dried over Na2SO4, filtered, and concentrated under reduced pressure to give the final product as a brown solid, 57.4 mg, 56% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.05 (t, J = 5.4 Hz, 1H), 8.26 (d, J = 8.3 Hz, 1H), 7.89−7.83 (m, 2H), 7.63 (ddd, J = 8.4, 6.9, 1.2 Hz, 1H), 7.55 (ddd, J = 8.5, 6.7, 1.2 Hz, 1H), 7.37 (d, J = 8.8 Hz, 1H), 7.21−7.16 (m, 2H), 6.89−6.85 (m, 2H), 3.71 (s, 3H), 3.57−3.50 (m, 2H), 3.34 (br s, 1H), 2.85 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 170.4, 159.6, 157.7, 135.8, 131.0, 129.6, 128.8, 127.4, 125.8, 124.7, 123.0, 122.5, 117.6, 113.8, 107.0, 55.0, 41.0, 33.9. ESI-MS (m/z): 322.0 [M + H]+. N-(4-Methoxyphenethyl)-1H-benzo[d]imidazole-2-carboxamide (53). The title compound was synthesized according to General Procedure 2 to give the final product as a white solid, 132.7 mg, 65% yield. 1H NMR (400 MHz, DMSO-d6) δ 13.20 (br s, 1H), 8.95 (t, J = 6.0 Hz, 1H), 7.74−7.67 (m, 1H), 7.56−7.48 (m, 1H), 7.34−7.23 (m, 2H), 7.19−7.14 (m, 2H), 6.87−6.82 (m, 2H), 3.71 (s, 3H), 3.55− 3.48 (m, 2H), 2.82 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 158.6, 157.7, 145.7, 142.5, 134.4, 131.1, 129.6, 124.0, 122.5, 119.8, 113.8, 112.5, 55.0, 40.5, 34.1. ESI-MS (m/z): 296.0 [M + H]+. N-(4-Methoxyphenethyl)benzo[d]oxazole-2-carboxamide (54). The title compound was synthesized according to General Procedure 2, except no precipitate formed. Water and EtOAc were added to the reaction mixture, and the organic layer was washed with 1 M NaOH (3×), water (3×), 1 M HCl, water (3×), and brine. The organic extracts were dried over Na2SO4, filtered, and concentrated under reduced pressure to give the final product as cream-colored solid, 6.3 mg, 7% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.33 (t, J = 5.7 Hz, 1H), 7.91−7.88 (m, 1H), 7.88−7.84 (m, 1H), 7.57 (ddd, J = 8.4, 7.3, 1.3 Hz, 1H), 7.50 (ddd, J = 8.4, 7.8, 1.2 Hz, 1H), 7.19−7.14 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.52−3.46 (m, 2H), 2.81 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 157.7, 155.7, 155.0, 150.1, 139.8, 130.9, 129.6, 127.4, 125.6, 121.0, 113.8, 111.8, 55.0, 40.9, 33.8. ESI-MS (m/z): 297.0 [M + H]+. 2-((4-Methoxyphenethyl)amino)-2-oxoacetic Acid (55). Lithium hydroxide (0.9 mmol) was added to a solution of ethyl (4methoxyphenethyl)carbamate (0.3 mmol, formed according to General Procedure 1) in 3:3:1 THF/MeOH/H2O (2 M). A white precipitate formed, and the reaction was finished in 2 h. HCl (1 M) was added until the reaction mixture became acidic. EtOAc was

added, and the organic layer was washed with water and brine, dried with Na2SO4, and concentrated under reduced pressure to yield a white solid, 52.9 mg, 87% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.81 (t, J = 5.7 Hz, 1H), 7.13−7.08 (m, 2H), 6.87−6.82 (m, 2H), 3.71 (s, 3H), 3.34−3.27 (m, 2H), 2.70 (t, J = 7.7 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 162.2, 158.8, 157.7, 130.9, 129.6, 113.8, 55.0, 40.7, 33.7. ESI-MS (m/z): 224.0 [M + H]+. N1-(4-Chlorobenzyl)-N2-(4-methoxyphenethyl)oxalamide (56). The title compound was prepared according to General Procedure 1 to give a white solid, 12 mg, 23% yield. 1H NMR (500 MHz, DMSO-d6) δ 9.33 (t, J = 6.4 Hz, 1H), 8.79 (t, J = 6.1 Hz, 1H), 7.40− 7.36 (m, 2H), 7.30−7.24 (m, 2H), 7.13−7.09 (m, 2H), 6.86−6.82 (m, 2H), 4.32−4.27 (m, 2H), 3.71 (s, 3H), 3.34−3.30 (m, 2H), 2.71 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ160.2, 159.7, 157.7, 137.8, 131.5, 130.9, 129.5, 129.2, 128.2, 113.8, 55.0, 41.7, 40.6, 33.7. ESI-MS (m/z): 346.9 [M + H]+. N1,N2-Bis(4-methoxyphenethyl)oxalamide (57). The title compound was prepared according to General Procedure 1 to provide 54.0 mg of a white solid in 56% yield. 1H NMR (400 MHz, DMSOd6) δ 8.70 (t, J = 6.1 Hz, 2H), 7.13−7.07 (m, 4H), 6.86−6.82 (m, 4H), 3.71 (s, 6H), 3.33−3.28 (m, 4H), 2.70 (t, J = 7.2 Hz, 4H). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 157.7, 130.9, 129.5, 113.8, 55.0, 40.5, 33.7. ESI-MS (m/z): 357.2 [M + H]+. N1-Cyclopropyl-N2-(4-methoxyphenethyl)oxalamide (58). The title compound was synthesized according to General Procedure 1 to give a white solid, 147 mg, 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.75−8.68 (m, 2H), 7.13−7.08 (m, 2H), 6.86−6.82 (m, 2H), 3.71 (s, 3H), 3.35−3.28 (m, 2H), 2.78−2.67 (m, 3H), 0.65− 0.59 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 161.2, 159.7, 157.7, 130.9, 129.5, 113.8, 55.0, 40.5, 33.7, 22.8, 5.4. ESI-MS (m/z): 263.0 [M + H]+. HRMS (ESI) m/z: calcd for C14H18N2O3 (M + H)+ 263.139 02, found 263.1396. N1-Cyclobutyl-N2-(4-methoxyphenethyl)oxalamide (59). The title compound was synthesized according to General Procedure 1 to give a white solid, 170 mg, 99% yield. 1H NMR (500 MHz, DMSO-d6) δ 8.95 (d, J = 8.6 Hz, 1H), 8.72 (t, J = 6.0 Hz, 1H), 7.12− 7.08 (m, 2H), 6.86−6.82 (m, 2H), 4.27−4.18 (m, 1H), 3.71 (s, 3H), 3.34−3.29 (m, 2H), 2.70 (t, J = 7.3 Hz, 2H), 2.13−2.06 (m, 4H), 1.64−1.56 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.9, 158.9, 157.7, 130.9, 129.5, 113.8, 55.0, 44.0, 40.5, 33.7, 29.5, 14.7. ESI-MS (m/z): 277.0 [M + H]+. N1-Cyclopentyl-N2-(4-methoxyphenethyl)oxalamide (60). The title compound was synthesized according to General Procedure 1 to give a white solid, 287 mg, 92% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.70 (t, J = 6.0 Hz, 1H), 8.55 (d, J = 8.2 Hz, 1H), 7.13− 7.09 (m, 2H), 6.87−6.82 (m, 2H), 4.07−3.97 (m, 1H), 3.71 (s, 3H), 3.35−3.29 (m, 2H), 2.71 (t, J = 7.5 Hz, 2H), 1.84−1.72 (m, 2H), 1.71−1.58 (m, 2H), 1.55−1.44 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 160.0, 159.6, 157.7, 130.9, 129.5, 113.8, 55.0, 50.6, 40.5, 33.7, 31.7, 23.5. ESI-MS (m/z): 291.0 [M + H]+. N1-Cyclohexyl-N2-(4-methoxyphenethyl)oxalamide (61). The title compound was synthesized according to General Procedure 1 to give a white solid, 225 mg, 92% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.75−8.67 (m, 1H), 8.45 (d, J = 8.4 Hz, 1H), 7.11 (d, J = 7.7 Hz, 2H), 6.84 (d, J = 7.7 Hz, 2H), 3.71 (s, 3H), 3.61−3.49 (m, 1H), 2.71 (t, J = 6.7 Hz, 2H), 1.74−1.61 (m, 4H), 1.60−1.51 (m, 2H), 1.40−1.16 (m, 4H), 1.15−1.01 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 160.0, 158.9, 157.7, 130.9, 129.5, 113.8, 55.0, 48.2, 40.6, 33.7, 31.8, 25.0, 24.8. ESI-MS (m/z): 305.0 [M + H]+. N1-Cycloheptyl-N2-(4-methoxyphenethyl)oxalamide (62). The title compound was synthesized according to General Procedure 1 to give a white solid, 225 mg, 94% yield. 1H NMR (500 MHz, DMSO-d6) δ 8.72 (apps, 1H), 8.49 (d, J = 7.2 Hz, 1H), 7.11 (d, J = 6.4 Hz, 2H), 6.84 (d, J = 6.4 Hz, 2H), 3.78−3.65 (m, 4H), 3.32−3.26 (m, 2H), 2.75−2.66 (m, 2H), 1.77−1.29 (m, 12H). 13C NMR (100 MHz, DMSO-d6) δ 160.1, 158.6, 157.7, 130.9, 129.5, 113.8, 55.0, 50.4, 40.6, 33.9, 33.7, 27.6, 23.8. ESI-MS (m/z): 319.0 [M + H]+. N-(4-Methoxyphenethyl)-2-oxo-2-(pyrrolidin-1-yl)acetamide (63). The title compound was synthesized according to General Procedure 1 with purification by flash chromatography over silica gel 5211



Article

(0 to 100% EtOAc/hexanes) to give a cream-colored solid, 161.5 mg, 97% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.61 (t, J = 5.8 Hz, 1H), 7.15−7.09 (m, 2H), 6.87−6.82 (m, 2H), 3.71 (s, 3H), 3.55− 3.50 (m, 2H), 3.35−3.28 (m, 4H), 2.69 (t, J = 7.5 Hz, 2H), 1.85− 1.71 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 162.1, 160.9, 157.7, 131.0, 129.7, 113.7, 55.0, 47.3, 46.1, 40.0, 33.8, 25.9, 23.2. ESI-MS (m/z): 277.0 [M + H]+. N-(4-Methoxyphenethyl)-2-oxo-2-(piperidin-1-yl)acetamide (64). The title compound was synthesized according to General Procedure 1. The crude residue was purified by flash chromatography (silica, 1:3 EtOAc/hex) to give a yellow oil, 103.8 mg, 99% yield. 1H NMR (500 MHz, DMSO-d6) δ 8.65 (t, J = 5.7 Hz, 1H), 7.15−7.11 (m, 2H), 6.86−6.82 (m, 2H), 3.71 (s, 3H), 3.40−3.36 (m, 2H), 3.33−3.30 (m, 2H), 3.16−3.12 (m, 2H), 2.68 (t, J = 7.1 Hz, 2H), 1.59−1.53 (m, 2H), 1.47−1.35 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 159.7, 158.6, 138.2, 136.6, 130.8, 130.6, 128.6, 128.3, 128.2, 122.0, 40.3, 33.8. ESI-MS (m/z): 291.0 [M + H]+. 2-(Indolin-1-yl)-N-(4-methoxyphenethyl)-2-oxoacetamide (65). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 158 mg, 91% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.86 (t, J = 5.7 Hz, 1H), 8.04 (d, J = 8.0 Hz, 1H), 7.29 (d, J = 7.5 Hz, 1H), 7.24−7.18 (m, 1H), 7.18−7.13 (m, 2H), 7.12 7.05 (m, 1H), 6.89−6.84 (m, 2H), 4.08−4.01 (m, 2H), 3.72 (s, 3H), 3.43−3.36 (m, 2H), 3.09 (t, J = 8.4 Hz, 2H), 2.77−2.71 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 162.3, 161.3, 157.8, 142.0, 132.7, 130.9, 129.7, 127.1, 125.1, 124.6, 116.7, 113.7, 55.0, 48.4, 40.1, 33.8, 27.8. ESI-MS (m/z): 325.0 [M + H]+. 2-(Isoindolin-2-yl)-N-(4-methoxyphenethyl)-2-oxoacetamide (66). The title compound was synthesized according to General Procedure 1 to give the final product as a cream-colored solid, 140 mg, 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.77 (t, J = 5.9 Hz, 1H), 7.39−7.34 (m, 1H), 7.33−7.27 (m, 3H), 7.17−7.12 (m, 2H), 6.88−6.82 (m, 2H), 4.94 (s, 2H), 4.73 (s, 2H), 3.71 (s, 3H), 3.41− 3.36 (m, 2H), 2.74 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 161.4, 160.8, 157.7, 137.1, 134.9, 131.0, 129.7, 127.5, 127.4, 122.9, 122.7, 113.7, 55.0, 53.3, 52.6, 40.1, 33.8. ESI-MS (m/z): 325.0 [M + H]+. 2-(3,4-Dihydroisoquinolin-2(1H)-yl)-N-(4-methoxyphenethyl)-2oxoacetamide (67). The title compound was synthesized according to General Procedure 1. The crude residue was purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the final product as a yellow oil, 103.6 mg, 77% yield. 1H NMR (500 MHz, DMSO-d6) (rotamers observed) δ 8.78−8.72 (m, 1H), 7.22− 7.12 (m, 6H), 6.89−6.82 (m, 2H), 4.60 (s, 1H), 4.47 (s, 1H), 3.73− 3.70 (m, 3H), 3.67 (t, J = 6.0 Hz, 1H), 3.49 (t, J = 6.0 Hz, 1H), 3.41− 3.36 (m, 2H), 2.80 (t, J = 5.7 Hz, 1H), 2.75 (t, J = 5.7 Hz, 1H), 2.71 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) (rotamers observed) δ 163.9, 163.5, 163.4, 163.3, 157.8, 157.70, 157.68, 134.2, 134.0, 132.8, 132.4, 131.7, 129.7, 129.6, 128.7, 127.5, 127.0, 126.68, 126.67, 126.5, 126.3, 126.2, 126.1, 113.8, 113.7, 55.02, 54.97, 46.8, 43.0, 42.9, 33.8, 33.7, 28.8, 27.7, 27.5. ESI-MS (m/z): 339.0 [M + H]+. N1-(4-Chlorophenyl)-N2-phenethyloxalamide (68). Step 1. Nitromethane (5.5 mL, 99.75 mmol, 80 equiv) was added to a solution of benzaldehyde (0.12 mL, 1.25 mmol, 1 equiv) and ammonium acetate (62.1 mg, 0.75 mmol, 0.6 equiv) in toluene (9 mL, 0.1 M), and the reaction was heated to 100 °C for 22 h. After the solution cooled to room temperature, water was added, and the solution was extracted with EtOAc (3×). The organic extracts were washed with brine, dried over Na2SO4, filtered, and concentrated under reduced pressure to yield 293.5 mg of (E)-(2-nitrovinyl)benzene as an orange solid in 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.24 (d, J = 13.6 Hz, 1H), 8.13 (d, J = 13.6 Hz, 1H), 7.88−7.83 (m, 2H), 7.56−7.45 (m, 3H). Step 2. Pd/C (10%, 329.7 mg, 3.1 mmol, 1.6 equiv) was added to a solution of the nitro alkene from Step 1 (289.6 mg, 1.9 mmol, 1 equiv) in anhydrous EtOH (30 mL, 0.067 M) and concentrated HCl (0.31 mL, 6.25 M) under nitrogen at 0 °C. The nitrogen was purged and replaced with a H2 balloon. When the reaction was finished as determined by TLC analysis (3.5 h), the reaction was filtered through Celite and washed with EtOH, and the filtrate was concentrated

under reduced pressure. Then, CH3CN was added, and a white precipitate formed, which was collected by filtration to yield 120.6 mg of 2-phenylethan-1-amine hydrochloride as a white solid in 51% yield. 1 H NMR (500 MHz, DMSO-d6) δ 8.11 (br s, 2H), 7.35−7.31 (m, 2H), 7.28−7.23 (m, 3H), 3.03−2.98 (m, 2H), 2.91−2.86 (m, 2H). Step 3. Oxalamide 68 was synthesized according to General Procedure 1 using the amine from Step 2 to give a white solid, 32.6 mg, 77% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.78 (br s, 1H), 9.05 (t, J = 5.9 Hz, 1H), 7.88−7.83 (m, 2H), 7.43−7.38 (m, 2H), 7.32−7.28 (m, 2H), 7.24−7.18 (m, 3H), 3.47−3.40 (m, 2H), 2.83 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 139.1, 136.7, 128.6, 128.4, 128.2, 126.2, 122.0, 40.6, 34.6. ESI-MS (m/z): 302.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(3-methoxyphenethyl)oxalamide (69). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 77.1 mg, 76% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.79 (br s, 1H), 9.03 (t, J = 6.0 Hz, 1H), 7.87−7.83 (m, 2H), 7.43−7.39 (m, 2H), 7.20 (t, J = 8.0 Hz, 1H), 6.81−6.75 (m, 3H), 3.73 (s, 3H), 3.48−3.40 (m, 2H), 2.81 (t, J = 7.6 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 159.3, 158.7, 140.6, 136.7, 129.4, 128.6, 128.2, 121.9, 120.9, 114.2 111.7, 54.9, 40.4, 34.5. ESI-MS (m/z): 333.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(2-methoxyphenethyl)oxalamide (70). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 37.9 mg, 72% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.79 (br s, 1H), 8.99 (t, J = 5.9 Hz, 1H), 7.87−7.83 (m, 2H), 7.43−7.39 (m, 2H), 7.20 (ddd, J = 7.8, 7.8, 1.7 Hz, 1H), 7.13 (dd, J = 7.4, 1.7 Hz, 1H), 6.96 (d, J = 8.0 Hz, 1H), 6.86 (ddd, J = 7.4, 7.4, 0.8 Hz, 1H), 3.78 (s, 3H), 3.44−3.37 (m, 2H), 2.81 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 157.3, 136.7, 130.0, 128.6, 127.7, 126.9, 121.9, 120.3, 110.7, 55.3, 39.9, 29.3. ESI-MS (m/z): 332.9 [M + H]+. N1-(4-Bromophenethyl)-N2-(4-chlorophenyl)oxalamide (71). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 42.2 mg, 74% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.79 (br s, 1H), 9.07 (t, J = 6.0 Hz, 1H), 7.87−7.83 (m, 2H), 7.50−7.45 (m, 2H), 7.43−7.39 (m, 2H), 7.21− 7.17 (m, 2H), 3.46−3.40 (m, 2H), 2.81 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) 159.7, 158.7, 138.6, 136.7, 131.2, 131.0, 128.7, 128.3, 122.0, 119.3, 40.3, 33.9. ESI-MS (m/z): 380.9 [M + H]+. N1-(4-Chlorophenethyl)-N2-(4-chlorophenyl)oxalamide (72). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 36.7 mg, 72% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.79 (br s, 1H), 9.08 (t, J = 6.0 Hz, 1H), 7.87−7.83 (m, 2H), 7.43−7.39 (m, 2H), 7.36−7.32 (m, 2H), 7.26− 7.23 (m, 2H), 3.46−3.40 (m, 2H), 2.82 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.6, 138.2, 136.6, 130.8, 130.6, 128.6, 128.3, 128.2, 121.9, 40.3, 33.8. ESI-MS (m/z): 336.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-fluorophenethyl)oxalamide (73). The title compound was synthesized according to General Procedure 1 to give a white solid, 8.6 mg, 20% yield. 1H NMR (500 MHz, DMSO-d6) 10.79 (br s, 1H), 9.07 (t, J = 6.1 Hz, 1H), 7.87−7.83 (m, 2H), 7.43− 7.39 (m, 2H), 7.27−7.23 (m, 2H), 7.14−7.09 (m, 2H), 3.45−3.39 (m, 2H), 2.82 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 158.4, 136.6, 135.3, 130.4 (d, 3JCF = 7.7 Hz), 128.6, 128.2, 121.9, 115.0 (d, 2JCF = 21.0 Hz), 40.6, 33.6. ESI-MS (m/z): 320.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-ethoxyphenethyl)oxalamide (74). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 43.1 mg, 77% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.78 (br s, 1H), 9.01 (t, J = 5.9 Hz, 1H), 7.88−7.82 (m, 2H), 7.43−7.38 (m, 2H), 7.14−7.09 (m, 2H), 6.86−6.81 (m, 2H), 3.97 (q, J = 7.0 Hz, 2H), 3.43−3.35 (m, 2H), 2.75 (t, J = 7.2 Hz, 2H), 1.30 (t, J = 7.0 Hz, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 157.0, 136.7, 130.8, 129.6, 128.6, 128.2, 121.9, 114.3, 62.9, 40.8, 33.7, 14.7. ESI-MS (m/z): 346.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-methylphenethyl)oxalamide (75). The title compound was synthesized according to General Procedure 5212



Article

1 to give a white solid, 27.4 mg, 53% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.80 (br s, 1H), 9.04 (t, J = 6.0 Hz, 1H), 7.88−7.83 (m, 2H), 7.43−7.39 (m, 2H), 7.12−7.08 (m, 4H), 3.43−3.38 (m, 2H), 2.78 (t, J = 7.3 Hz, 2H), 2.25 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 136.7, 135.9, 135.1, 128.9, 128.6, 128.5, 128.2, 121.9, 40.7, 34.1, 20.7. ESI-MS (m/z): 316.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(3,4-dimethoxyphenethyl)oxalamide (76). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 38.0 mg, 67% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.78 (br s, 1H), 9.00 (t, J = 6.0 Hz, 1H), 7.87−7.82 (m, 2H), 7.43−7.38 (m, 2H), 6.85 (d, J = 8.1 Hz, 1H), 6.81 (s, 1H), 6.72 (d, J = 8.1, 1H), 3.72 (s, 3H), 3.70 (s, 3H), 3.45−3.38 (m, 2H), 2.76 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 148.6, 147.3, 136.7, 131.5, 128.6, 128.2, 122.0, 120.4, 112.4, 111.8, 55.5, 55.3, 40.7, 34.1. ESI-MS (m/ z): 362.9 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-hydroxyphenethyl)oxalamide (77). The title compound was synthesized according to General Procedure 1 to give a white solid, 35.2 mg, 68% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.79 (br s, 1H), 9.20 (br s, 1H), 9.01 (t, J = 6.0 Hz, 1H), 7.88−7.83 (m, 2H), 7.44−7.39 (m, 2H), 7.02−6.98 (m, 2H), 6.69−6.65 (m, 2H), 3.39−3.35 (m, 2H), 2.70 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 155.7, 136.7, 129.5, 129.0, 128.6, 128.2, 121.9, 115.1, 40.9, 33.8. ESI-MS (m/z): 317.0 [M − H]−. N1-(4-Chlorophenyl)-N2-(3-hydroxyphenethyl)oxalamide (78). A solution of analogue 69 (49.7 mg, 0.15 mmol, 1 equiv) in anhydrous CH2Cl2 (0.75 mL, 0.2 M) under nitrogen was cooled to −78 °C for 20 min before 1.5 mL of a 1.0 M solution of BBr3 in CH2Cl2 (1.5 mL, 1 mmol, 10 equiv) was added dropwise at −78 °C. The reaction was stirred at −78 °C until the reaction was complete as judged by TLC analysis (4−5 h). Then, the reaction was warmed to room temperature, poured into ice water, and extracted into EtOAc (3×). The organic extracts were washed with brine, dried over Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by flash chromatography over silica gel (0.5% MeOH in CH2Cl2) to provide 12.7 mg of a white solid in 27% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.79 (br s, 1H), 9.29 (br s, 1H), 9.03 (t, J = 5.9 Hz, 1H), 7.88−7.83 (m, 2H), 7.43−7.38 (m, 2H), 7.07 (t, J = 7.7 Hz, 1H), 6.65−6.57 (m, 3H), 3.43−3.36 (m, 2H), 2.73 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.7, 157.4, 140.4, 136.7, 129.3, 128.6, 128.2, 122.0, 119.2, 115.5, 113.2, 40.6, 34.6. ESIMS (m/z): 317.0 [M − H]−. HRMS (ESI) m/z: calcd for C16H15ClN2O3 (M + H)+ 319.084 40, found 319.0855. N1-(4-Chlorophenyl)-N2-(2-hydroxyphenethyl)oxalamide (79). The title compound was prepared using the synthetic procedures described for the preparation of analogue 78 using analogue 70 (66.3 mg, 0.2 mmol, 1 equiv). The residue was purified by flash chromatography over silica gel (0 to 10% MeOH/CH2Cl2) and triturated with hexanes to provide 19.8 mg of a white solid in 31% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.78 (br s, 1H), 9.44 (br s, 1H), 8.97 (t, J = 5.3 Hz, 1H), 7.88−7.82 (m, 2H), 7.43−7.38 (m, 2H), 7.08−6.97 (m, 2H), 6.81−6.76 (m, 1H), 6.74−6.68 (m, 1H), 3.45−3.37 (m, 2H), 2.81−2.74 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 159.6, 158.8, 155.3, 136.7, 130.2, 128.6, 128.2, 127.4, 125.2, 121.9, 118.9, 114.9, 39.4, 29.3. ESI-MS (m/z): 319.1 [M + H]+. HRMS (ESI) m/z: calcd for C16H15ClN2O3 (M + H)+ 319.084 40, found 319.0853. N1-(4-Chlorophenyl)-N2-(4-methoxybenzyl)oxalamide (80). The title compound was synthesized according to General Procedure 1 to give the final product as a white solid, 32.3 mg, 68% yield. 1H NMR (500 MHz, DMSO-d6) 10.83 (br s, 1H), 9.50 (t, J = 6.4 Hz, 1H), 7.87−7.83 (m, 2H), 7.43−7.39 (m, 2H), 7.25−7.22 (m, 2H), 6.90− 6.86 (m, 2H), 4.31 (d, J = 6.4 Hz, 2H), 3.72 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 159.8, 158.8, 158.3, 136.7, 130.6, 128.9, 128.6, 128.2, 122.0, 113.7, 55.1, 42.1. ESI-MS (m/z): 317.0 [M − H]−. HRMS (ESI) m/z: calcd for C16H15ClN2O3 (M + Na)+ 341.066 34, found 341.0667. N1-(4-Chlorophenyl)-N2-(2-(4-methoxyphenyl)propyl)oxalamide (81). Step 1. (E)-1-Methoxy-4-(2-nitrovinyl)benzene was synthesized

following Step 1 of the procedure to synthesize 68 with panisaldehyde (0.15 mL, 1.25 mmol, 1 equiv) to yield 213.5 mg of (E)-1-methoxy-4-(2-nitrovinyl)benzene as an orange solid in 96% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.17−8.07 (m, 2H), 7.86− 7.81 (m, 2H), 7.07−7.02 (m, 2H), 3.83 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 162.6, 139.4, 135.8, 132.1, 122.7, 114.8, 55.6. LCMS (m/z): 180.1 [M + H]+. Step 2. According to a procedure by Bartoli,28 CeCl3·7H2O was dried by heating to 140 °C under vacuum with stirring overnight. Then, a slurry of CeCl3 (492.7 mg, 2 mmol, 2 equiv) in anhydrous THF (16 mL, 0.0625 M) was cooled to −78 °C under nitrogen for 20 min before a 2.9 M solution of methylmagnesium bromide in Et2O (0.69 mL, 2 mmol, 2 equiv) was added. The reaction was stirred at −78 °C for 1.5 h before it was warmed to −40 °C. Then, the nitro alkene from Step 1 (179.3 mg, 1 mmol, 1 equiv) in anhydrous THF (2.5 mL, 0.4 M) was added slowly to the reaction mixture. The reaction was finished in 10 min as determined by TLC, and the reaction was quenched with acetic acid (0.6 mL, 10 equiv). The solution was stirred for 5 min, and the solution turned bright yellow. The reaction was diluted with water, extracted into hexanes (3×), dried over Na2SO4, filtered, and concentrated under reduced pressure. The solution was azeotroped with toluene to remove acetic acid. The residue was purified by flash chromatography over silica gel (0 to 10% EtOAc in hexanes) to yield 139.3 mg of 1-methoxy-4-(1-nitropropan2-yl)benzene as a yellow oil in 71% yield. 1H NMR (400 MHz, CDCl3) δ 7.17−7.12 (m, 2H), 6.90−6.85 (m, 2H), 4.54−4.42 (m, 2H), 3.79 (s, 3H), 3.64−3.54 (m, 1H), 1.36 (d, J = 7.0 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ 159.0, 133.0, 128.1, 114.5, 82.3, 55.4, 38.1, 19.0. GC-MS m/z = 195.1. Step 3. Pd/C (10%, 295.4 mg) was added to a solution of the nitro alkane from Step 2 (179.2 mg, 0.9 mmol, 1 equiv) in EtOAc (9 mL, 0.1 M). The reaction solution was purged with N2 before the addition of H2 at 50 psi. After 2 h, additional 10% Pd/C (293.7 mg) was added. When the reaction was complete according to TLC analysis (2 h), the solution was filtered through Celite and washed with EtOAc. The filtrate was concentrated under reduced pressure to yield 104.1 mg of 2-(4-methoxyphenyl)propan-1-amine as a cream-colored solid in 69% yield. 1H NMR (400 MHz, CDCl3) δ 7.15−7.10 (m, 2H), 6.89−6.84 (m, 2H), 3.79 (s, 3H), 2.86−2.75 (m, 2H), 2.73−2.67 (m, 1H), 1.22 (d, J = 6.9 Hz, 3H). 13C NMR (100 MHz, CDCl3) 158.2, 137.1, 128.4, 114.1, 55.4, 49.8, 42.8, 19.6. GC-MS m/z = 165.1. Step 4. Oxalamide 81 was synthesized with the amine salt from Step 3 (104.1 mg, 0.63 mmol) according to General Procedure 1 to provide 20.2 mg of a white solid in 20% yield. 1H NMR (400 MHz, DMSO-d6) 10.78 (br s, 1H), 8.90 (t, J = 6.1 Hz, 1H), 7.87−7.81 (m, 2H), 7.43−7.38 (m, 2H), 7.18−7.13 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.39−3.25 (m, 2H, under water), 3.06−2.95 (m, 1H), 1.16 (d, J = 7.0 Hz, 3H). 13C NMR (100 MHz, DMSO-d6) 159.6, 158.7, 157.8, 136.7, 136.3, 128.7, 128.3, 128.1, 122.0, 113.8, 55.0, 46.2, 37.9, 19.5. LC-MS (m/z): 347.1 [M + H]+. N 1 -(4-Chlorophenyl)-N 2 -(1-(4-methoxyphenyl)propan-2-yl)oxalamide (82). Step 1. (E)-1-Methoxy-4-(2-nitroprop-1-en-1-yl)benzene was synthesized according to Step 1 of the procedure to synthesize 68 with nitroethane (7.2 mL, 99.75 mmol, 80 equiv). The crude mixture was purified by flash chromatography over silica gel (0 to 30% EtOAc in hexanes) to yield 465.0 mg of (E)-1-methoxy-4-(2nitroprop-1-en-1-yl)benzene (130) as a yellow solid in 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.08 (s, 1H), 7.63−7.58 (m, 2H), 7.10−7.05 (m, 2H), 3.82 (s, 3H), 2.42 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 160.9, 145.5, 133.3, 132.6, 124.3, 114.5, 55.4, 14.0. ESIMS (m/z): 194.1 [M + H]+. Step 2. Following a procedure by Munoz,33 a solution of the nitro alkene from Step 1 (287.1 mg, 1.5 mmol, 1 equiv) in anhydrous Et2O (15 mL, 0.1 M) was added dropwise to a solution of LiAlH4 in anhydrous Et2O (15 mL, 0.1 M) under a nitrogen atmosphere. The solution was heated at reflux until complete as judged by TLC analysis (3 h). Then, the solution was cooled to 0 °C and quenched slowly with 0.2 mL of water, 0.2 mL of 15% NaOH(aq), and 0.7 mL of water. The solution was stirred for 15 min. MgSO4 was added, and the solution was stirred for 15 min. Then, the solution was filtered to 5213



Article

119.6, 113.7, 113.6, 55.0, 46.8, 40.3, 34.3. ESI-MS (m/z): 318.9 [M + H]+. 1-(4-Chlorophenyl)-3-(4-methoxyphenethyl)urea (86). Step 1. 4Nitrophenyl chloroformate (1.11 mmol) was added to a mixture of 4chloroaniline (1 mmol), saturated NaHCO3 (1.3 mmol), t-butyl methyl ether (0.71 M), and H2O (0.35 M). After being stirred at rt for 1 h, the reaction was washed with 1 M HCl (3 × 5 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to yield 4nitrophenyl (4-chlorophenyl)carbamate (137) as a cream-colored solid, 213.2 mg, 82% yield. 1H NMR (500 MHz, CDCl3) δ 8.32−8.28 (m, 2H), 7.43−7.37 (m, 4H), 7.36−7.32 (m, 2H), 7.01 (br s, 1H). ESI-MS (m/z): 293.8 [M + H]+. Step 2. 4-Methoxyphenethylamine (0.3 mmol) was added to a solution of Et3N (0.3 mmol) and the carbamate from Step 1 (0.3 mmol) in CH2Cl2 (0.125 M), and the yellow reaction mixture was stirred at room temperature for 1 h. The precipitate was collected by filtration and rinsed with water to remove the yellow color, which give urea 86 as a white solid, 55.7 mg, 61% yield. 1H NMR (500 MHz, DMSO-d6) δ 8.62 (br s, 1H), 7.42−7.38 (m, 2H), 7.26−7.23 (m, 2H), 7.16−7.13 (m, 2H), 6.89−6.85 (m, 2H), 6.11 (t, J = 5.6 Hz, 1H), 3.72 (s, 3H), 3.31−3.25 (m, 2H), 2.67 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 157.7, 155.0, 139.6, 131.3, 129.6, 128.5, 124.4, 119.0, 113.8, 55.0, 40.8, 34.9. ESI-MS (m/z): 304.9 [M + H]+. 1-(4-Chlorophenyl)-3-(3-(4-methoxyphenyl)propyl)urea (87). The title compound was synthesized analogously to compound 86 from the known 3-(4-methoxyphenyl)propan-1-amine (prepared in three steps from 3-(4-methoxyphenyl)-1-propanol).34 The crude residue was purified by flash chromatography (silica gel, 0−100% EtOAc in hexanes) to give the product as a white solid, 27.6 mg, 66% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.56 (br s, 1H), 7.44−7.39 (m, 2H), 7.27−7.22 (m, 2H), 7.14−7.10 (m, 2H), 6.86−6.82 (m, 2H), 6.22 (t, J = 5.6 Hz, 1H), 3.71 (s, 3H), 3.10−3.03 (m, 2H), 2.56−2.51 (m, 2H), 1.68 (quint, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 157.4, 155.0, 139.6, 133.5, 129.2, 128.4, 124.3, 119.0, 113.7, 55.0, 38.6, 31.7, 31.6. ESI-MS (m/z): 318.9 [M + H]+. N-(4-Chlorophenyl)-3-(4-methoxyphenyl)propanamide (88). Oxalyl chloride (0.5 mmol) and DMF (1 drop) were added to a solution of 3-(4-methoxyphenyl)propanoic acid (2.5 mmol) in CH2Cl2 (0.1 M). The reaction was stirred at room temperature for 1.5 h, and the solvent was removed under a flow of N2 over 1 h. The acid chloride was dissolved in CH2Cl2 (0.1 M), and Et3N (2 equiv) and 4chloroaniline (0.19 mmol) were added. The reaction mixture was stirred overnight at rt. It was concentrated under reduced pressure, and the residue was purified by flash chromatography (silica gel, 0 to 30% EtOAc in hexanes) to give the product as a white solid, 12.9 mg, 23% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.02 (br s, 1H), 7.63− 7.58 (m, 2H), 7.36−7.31 (m, 2H), 7.17−7.13 (m, 2H), 6.86−6.82 (m, 2H), 3.70 (s, 3H), 2.84 (t, J = 7.3 Hz, 2H), 2.58 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 170.6, 157.5, 138.2, 132.9, 129.2, 128.6, 126.5, 120.5, 113.7, 55.0, 38.3, 29.9. ESI-MS (m/z): 290.0 [M + H]+. N-(4-Chlorophenyl)-4-(4-methoxyphenyl)butanamide (89). The title compound was synthesized analogously to amide 88. After the reaction was finished, the reaction was concentrated under reduced pressure, and then water (3 mL) and EtOAc (5 mL) were added. The residue was extracted with EtOAc (3 × 5 mL), washed with brine, dried over Na2SO4, filtered, and concentrated under reduced pressure to give the product as a white solid, 125.9 mg, 83% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.00 (br s, 1H), 7.64−7.59 (m, 2H), 7.35− 7.31 (m, 2H), 7.14−7.09 (m, 2H), 6.87−6.82 (m, 2H), 3.71 (s, 3H), 2.55 (t, J = 7.5 Hz, 2H), 2.29 (t, J = 7.5 Hz, 2H), 1.84 (quint, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 171.2, 157.4, 138.3, 133.4, 129.3, 128.5, 126.4, 120.5, 113.7, 55.0, 35.7, 33.7, 26.9. ESI-MS (m/z): 304.0 [M + H]+. N-(4-Chlorophenyl)-5-(4-methoxyphenyl)pentanamide (90). The title compound was synthesized analogously to amide 88. The crude reaction mixture was purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the product as a cream-colored solid, 1.9 mg, 1% yield after. 1H NMR (500 MHz, DMSO-d6) δ 10.02 (br s,

remove salts and concentrated under reduced pressure to yield 201.3 mg of 2-(4-methoxyphenyl)propan-1-amine (131) as a clear oil in 82% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.10−7.05 (m, 2H), 6.86−6.81 (m, 2H), 3.71 (s, 3H), 2.97−2.88 (m, 1H), 2.44 (dd, J = 6.7, 1.6 Hz, 2H), 0.92 (d, J = 6.3 Hz, 3H). 13C NMR (100 Hz, DMSO-d6) δ 157.5, 132.0, 130.1, 113.6, 54.9, 48.4, 45.4, 23.2. ESI-MS (m/z): 166.1 [M + H]+. Step 3. Oxalamide 82 was prepared according to General Procedure 1 using the amine from Step 2 (41.3 mg, 2.5 mmol, 3 equiv) to provide 92.5 mg of a white solid in 25% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.72 (br s, 1H), 8.87 (d, J = 8.9 Hz, 1H), 7.86−7.81 (m, 2H), 7.43−7.38 (m, 2H), 7.14−7.09 (m, 2H), 6.85− 6.80 (m, 2H), 4.10−4.00 (m, 1H), 3.69 (s, 3H), 2.82 (dd, J = 13.5, 8.2 Hz, 1H), 2.67 (dd, J = 13.5, 6.2 Hz, 1H), 1.14 (d, J = 6.6 Hz, 3H). 13 C NMR (100 Hz, DMSO-d6) δ 159.1, 158.9, 157.8, 136.7, 131.0, 130.2, 128.8, 128.5, 122.2, 133.8, 55.1, 47.5, 40.5, 20.1. ESI-MS (m/ z): 347.2 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-methoxyphenethyl)-N2-methyloxalamide (83). The title compound was synthesized according to General Procedure 1. Upon cooling the reaction mixture to rt, a precipitate formed and was filtered. The filtrate was concentrated, and the residue was purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the final product as a cream-colored solid, 4.4 mg, 11% yield. 1H NMR (400 MHz, DMSO-d6) (rotamers) δ 10.87 (br s, 1H), 10.71 (br s, 1H), 7.72−7.65 (m, 4H), 7.43−7.38 (m, 4H), 7.22−7.17 (m, 2H), 7.08−7.03 (m, 2H), 6.90−6.85 (m, 2H), 6.80− 6.76 (m, 2H), 3.72 (s, 2H), 3.65 (s, 3H), 3.61−3.55 (m, 2H), 3.55− 3.48 (m, 2H), 2.99 (s, 2H), 2.92 (s, 3H), 2.88−2.75 (m, 4H). 13C NMR (100 Hz, DMSO-d6) (rotamers) δ 163.6, 163.4, 162.1, 161.9, 157.8, 136.91, 136.87, 129.7, 129.6, 128.8, 128.7, 121.4, 113.94, 113.90, 55.0, 54.9, 51.4, 48.3, 35.5, 33.5, 32.4, 31.6. ESI-MS (m/z): 346.9 [M + H]+. 2-(4-Chlorophenoxy)-N-(4-methoxyphenethyl)acetamide (84). Step 1. A solution of 4-methoxyphenethylamine (0.73 mL, 5 mmol, 1 equiv) and Et3N (0.69 mL, 5 mmol, 1 equiv) in anhydrous CH2Cl2 (10 mL, 0.5 M) was stirred at 0 °C for 30 min. Then, bromoacetyl bromide (0.52 mL, 6 mmol, 1.2 equiv) was added dropwise over 20 min. The solution was warmed to room temperature and stirred overnight. Then, the solution was filtered to remove the HBr salts, and the filtrate was washed with water (5 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to yield 1.21 g of 2bromo-N-(4-methoxyphenethyl)acetamide (136) as a brown solid in 88% yield. 1H NMR (500 MHz, CDCl3) δ 7.14−7.10 (m, 2H), 6.88− 6.84 (m, 2H), 6.52 (br s, 1H), 3.86 (s, 2H), 3.80 (s, 3H), 3.53−3.48 (m, 2H), 2.78 (t, J = 7.0 Hz, 2H). ESI-MS (m/z): 273.9 [M + H]+. Step 2. 4-Chlorophenol (1.5 mmol) was added to the bromoacetamide from Step 1 (1.5 mmol) and K2CO3 (1.5 mmol) in acetonitrile (0.5 M). The mixture was headed at reflux overnight until the reaction was complete as judged by TLC analysis. After cooling to rt, the reaction mixture was filtered to remove salts, and the filtrate was concentrated. H2O and diethyl ether (1:1, v/v; 4 mL) were added to the residue with stirring, and a precipitate formed. The precipitate was collected by filtration, rinsed with diethyl ether, and dried to give analogue 84 as a cream-colored solid, 83.1 mg, 17% yield. 1H NMR (500 MHz, DMSO-d6) δ 8.14 (t, J = 5.7 Hz, 1H), 7.36−7.32 (m, 2H), 7.11−7.07 (m, 2H), 6.96−6.92 (m, 2H), 6.85− 6.81 (m, 2H), 4.45 (s, 2H), 3.71 (s, 3H), 3.33−3.27 (m, 2H), 2.67 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.2, 157.7, 156.6, 131.1, 129.6, 129.2, 124.8, 116.5, 113.7, 67.2, 55.0, 39.9, 34.2. ESI-MS (m/z): 319.9 [M + H]+. 2-((4-Chlorophenyl)amino)-N-(4-methoxyphenethyl)acetamide (85). The title compound was synthesized analogously to compound 84. The crude precipitate was purified by flash chromatography (silica gel, 0−100% EtOAc in hexanes) to give an orange solid, 244.7 mg, 50% yield. 1H NMR (500 MHz, DMSO-d6) δ 7.90 (t, J = 5.8 Hz, 1H), 7.11−7.08 (m, 2H), 7.07−7.04 (m, 2H), 6.82−6.79 (m, 2H), 6.51−6.47 (m, 2H), 6.16 (t, J = 5.8 Hz, 1H), 3.71 (s, 3H), 3.57 (d, J = 5.8 Hz, 2H), 3.28−3.23 (m, 2H), 2.62 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO- d6) δ 169.7, 157.6, 147.3, 131.2, 129.6, 128.5, 5214



Article

1H), 7.62−7.59 (m, 2H), 7.35−7.31 (m, 2H), 7.11−7.08 (m, 2H), 6.84−6.81 (m, 2H), 3.70 (s, 3H), 2.55−2.51 (m, 2H), 2.33−2.93 (m, 2H), 1.60−1.52 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 171.3, 157.3, 138.3, 133.9, 129.2, 128.6, 126.4, 120.5, 113.7, 55.0, 36.3, 34.0, 30.9, 24.6. ESI-MS (m/z): 318.0 [M + H]+. 4-Chloro-N-(4-methoxyphenethyl)benzamide (91). 4-Methoxyphenethylamine (0.3 mmol) was added to a solution of 4chlorobenzoyl chloride (0.3 mmol) and Et3N (0.6 mmol) in CH2Cl2 (0.1 M). After TLC indicated the complete consumption of the starting materials (2 d), the reaction mixture was diluted with CH2Cl2 and washed with saturated NaHCO3. The organic layer was dried with Na2SO4, filtered, concentrated under reduced pressure, and purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the final product as a white solid, 18.6 mg, 21% yield. 1 H NMR (400 MHz, DMSO-d6) δ 8.62 (t, J = 5.6 Hz, 1H), 7.85− 7.81 (m, 2H), 7.55−7.51 (m, 2H), 7.17−7.12 (m, 2H), 6.88−6.83 (m, 2H), 3.71 (s, 3H), 3.46−3.39 (m, 2H), 2.77 (t, J = 7.5 Hz, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 165.0, 157.7, 135.9, 133.3, 131.3, 129.6, 129.1, 128.4, 113.8, 55.0, 41.2, 34.2. ESI-MS (m/z): 290.1 [M + H]+. 4-Chloro-N-(3-(4-methoxyphenyl)propyl)benzamide (92). The title compound was synthesized analogously to amide 91 to give the final product as a white solid, 51.2 mg, 54% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.54 (t, J = 5.5 Hz, 1H), 7.88−7.83 (m, 2H), 7.55−7.51 (m, 2H), 7.15−7.11 (m, 2H), 6.86−6.82 (m, 2H), 3.71 (s, 3H), 3.29−3.22 (m, 2H), 2.56 (t, J = 7.5 Hz, 2H), 1.78 (quint, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 165.1, 157.4, 135.8, 133.5, 133.4, 129.2, 129.1, 128.3, 113.7, 55.0, 38.9, 31.7, 31.0. ESI-MS (m/z): 304.1 [M + H]+. 4-Chloro-N-(4-(4-methoxyphenyl)butyl)benzamide (93). Step 1. Oxalyl chloride (5 equiv, 5.2 mmol) was added to a solution of 4-(4methoxyphenyl)butyric acid (1 equiv, 1 mmol) in THF (0.1 M), and then a catalytic amount of DMF was added to initiate the reaction. After 1 h, the volatile components were removed under a positive N2 flow over 1 h. Ammonium hydroxide (1 mL) was added to the residue in CH2Cl2 (2 mL) at 0 °C, and a white precipitate formed. The reaction mixture was stirred for 20 min at rt. Then, the precipitate was collected by filtration, rinsed with water, and dried under vacuum to give 4-(4-methoxyphenyl)butanamide as a white solid, which was used without purification. ESI-MS (m/z): 194.0 [M + H]+. Step 2. A solution of the amide from Step 1 (1 equiv, 1.04 mmol) in anhydrous THF (1 M) was added to a suspension of LiAlH4 (5 equiv, 4.2 mmol) in anhydrous THF (5 M) under N2. The reaction was heated at reflux for 6.5 h. Then, the reaction was cooled to 0 °C and quenched slowly with 0.167 mL of H2O, 0.17 mL of 1 M NaOH, and 0.5 mL of 1 M NaOH, consecutively, and then stirred for 1 h. The mixture was filtered through Celite, and the filtrate was diluted with water and extracted with EtOAc. The organic extracts were washed with brine, dried with Na2SO4, filtered, and concentrated to give 4-(4-methoxyphenyl)butan-1-amine as a yellow oil in 36% yield (67.9 mg). ESI-MS (m/z): 180.1 [M + H]+. Step 3. The amine from Step 2 was coupled with 4-chlorobenzoyl chloride as described for compound 91 to give amide 93 as a white solid in 14% yield (16.7 mg). 1H NMR (400 MHz, DMSO-d6) δ 8.53 (t, J = 5.5 Hz, 1H), 7.86−7.82 (m, 2H), 7.54−7.50 (m, 2H), 7.12− 7.08 (m, 2H), 6.85−6.80 (m, 2H), 3.70 (s, 3H), 3.29−3.23 (m, 2H), 2.56−2.51 (m, 2H), 1.61−1.45 (m, 4H). 4-Chlorobenzoyl chloride: δ 7.96−7.92 (m, 0.7H), 7.59−7.55 (m, 0.7H). 13C NMR (100 MHz, DMSO-d6) δ 165.0, 157.3, 135.8, 134.0, 133.4, 129.2, 129.1, 128.3, 113.7, 55.0, 33.9, 28.73, 28.67. 4-Chlorobenzoyl chloride: δ 131.2 and 128.8. ESI-MS (m/z): 318.1 [M + H]+. General Procedure 3, for the Synthesis of N-Acyl Ureas, As Exemplified by the Synthesis of 4-Chloro-N-((4methoxyphenethyl)carbamoyl)benzamide (94). Oxalyl chloride (4.3 mmol) was added to a solution of 4-chlorobenzamide (0.85 mmol) in CH2Cl2 (0.1 M) and heated at 50 °C to form an intermediate isocyanate (138).30 After 5 h, the volatile solvents were removed under a flow of N2. 4-Methoxyphenethylamine (1 mmol) was added to a solution of the residue in acetonitrile (0.1 M), and a white precipitate formed immediately. The reaction mixture was

stirred until completion as indicated by TLC analysis (12−48 h). The reaction mixture was concentrated under reduced pressure, and the residue was suspended in water (10 mL) and EtOAc (15 mL). The white precipitate was collected by filtration and dried under vacuum to give the final product as a white solid, 234.4 mg, 82% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.74 (br s, 1H), 8.66 (t, J = 5.6 Hz, 1H), 7.97−7.93 (m, 2H), 7.58−7.53 (m, 2H), 7.19−7.14 (m, 2H), 6.89−6.84 (m, 2H), 3.72 (s, 3H), 3.46−3.40 (m, 2H), 2.75 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.3, 157.7, 153.8, 137.4, 131.8, 131.0, 130.1, 129.6, 128.5, 113.8, 55.0, 40.9, 34.4. ESIMS (m/z): 332.9 [M + H]+. N-((4-Chlorophenyl)carbamoyl)-3-(4-methoxyphenyl)propanamide (95). The title compound was synthesized according to General Procedure 3 after purification by flash chromatography over silica gel (0 to 100% EtOAc in hexanes) to give a white solid, 18.7 mg, 36% yield (18.7 mg). 1H NMR (500 MHz, DMSO-d6) δ 10.77 (br s, 1H), 10.60 (br s, 1H), 7.58−7.54 (m, 2H), 7.39−7.35 (m, 2H), 7.16−7.12 (m, 2H), 6.87−6.83 (m, 2H), 3.71 (s, 3H), 2.81 (t, J = 7.4 Hz, 2H), 2.66 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 175.0, 157.6, 150.8, 136.6, 132.3, 129.2, 128.8, 127.3, 121.3, 113.8, 55.0, 37.7, 29.1. ESI-MS (m/z): 332.9 [M + H]+. 4-Bromo-N-((4-methoxyphenethyl)carbamoyl)benzamide (96). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 279.4 mg, 70% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.76 (br s, 1H), 8.62 (t, J = 5.6 Hz, 1H), 7.88−7.85 (m, 2H), 7.73−7.69 (m, 2H), 7.18−7.15 (m, 2H), 6.88−6.85 (m, 2H), 3.72 (s, 3H), 3.45−3.40 (m, 2H), 2.75 (t, J = 7.4 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.4, 157.7, 153.3, 131.8, 131.5, 131.0, 130.2, 129.6, 126.7, 113.8, 55.0, 40.9, 34.3. ESIMS (m/z): 377.0 [M + H]+. N-((4-Methoxyphenethyl)carbamoyl)-[1,1′-biphenyl]-4-carboxamide (97). A solution of 4-bromo-N-((4-methoxyphenethyl)carbamoyl)benzamide (96, 0.15 mmol, 1 equiv), phenylboronic acid (0.3 mmol, 1 equiv), Pd(dppf)Cl2 (0.015 mmol), cesium fluoride (0.39 mmol, 2.6 equiv), isopropanol (0.1 M), and Et3N (0.225 mmol, 1.5 equiv) was heated at 100 °C for 5 h. After the solution cooled to rt, water was added to the reaction solution, and it was extracted into EtOAc. The organic layer was washed with brine, dried with Na2SO4, filtered, and concentrated before purification by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the desired product as a white solid, 20.3 mg, 36% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.74 (br s, 1H), 8.72 (t, J = 5.8 Hz, 1H), 8.07−8.03 (m, 2H), 7.82−7.79 (m, 2H), 7.77−7.73 (m, 2H), 7.53−7.48 (m, 2H), 7.45− 7.41 (m, 1H), 7.20−7.17 (m, 2H), 6.89−6.85 (m, 2H), 3.72 (s, 3H), 3.48−3.42 (m, 2H), 2.77 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.8, 157.8, 153.5, 144.2, 138.8, 131.3, 131.0, 129.7, 129.1, 128.9, 128.4, 127.0, 126.7, 113.9, 55.0, 40.9, 34.4. ESI-MS (m/ z): 375.1 [M + H]+. 4-Methoxy-N-((4-methoxyphenethyl)carbamoyl)benzamide (98). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 63.9 mg, 29% yield, contaminated with 41% dimer 57. 1H NMR (400 MHz, DMSO-d6) δ 10.50 (br s, 1H), 8.74 (t, J = 5.6 Hz, 1H), 7.99−7.94 (m, 2H), 7.19−7.14 (m, 2H), 7.13−7.08 (m, 2H), 6.89−6.86 (m, 2H), 3.82 (s, 3H), 3.72 (s, 3H), 3.46−3.40 (m, 2H), 3.32−3.28 2.75 (t, J = 7.1 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.4, 162.8, 159.8, 153.7, 131.0, 130.3, 129.6, 124.5, 113.81, 113.75, 55.5, 55.0, 40.8, 34.4. ESI-MS (m/z): 329.1 [M + H]+. Methyl 4-(((4-Methoxyphenethyl)carbamoyl)carbamoyl)benzoate (99). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 707.8 mg, 35% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.83 (br s, 1H), 8.60 (t, J = 5.4 Hz, 1H), 8.04−8.02 (m, 4H), 7.19−7.14 (m, 2H), 6.89−6.84 (m, 2H), 3.88 (s, 3H), 3.72 (s, 3H), 3.48−3.41 (m, 2H), 2.76 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.5, 165.5, 157.7, 153.2, 136.7, 132.9, 131.0, 129.6, 129.1, 128.6, 113.8, 55.0, 52.5, 40.9, 34.3. ESI-MS (m/z): 357.1 [M + H]+. 4-(((4-Methoxyphenethyl)carbamoyl)carbamoyl)benzoic Acid (100). Methyl 4-(((4-methoxyphenethyl)carbamoyl)carbamoyl)benzoate (99, 2 mmol, 1 equiv), LiOH (6 mmol, 3 equiv), THF, 5215



Article

mg, 36% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.65 (br s, 1H), 8.68 (t, J = 5.7 Hz, 1H), 7.98−7.93 (m, 2H), 7.58−7.53 (m, 2H), 7.33−7.26 (m, 2H), 7.20−7.14 (m, 2H), 7.04−6.99 (m, 2H), 6.98− 6.92 (m, 1H), 6.89−6.84 (m, 2H), 5.19 (s, 2H), 3.72 (s, 3H), 3.48− 3.41 (m, 2H), 2.76 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 167.9, 158.1, 157.7, 153.5, 142.1, 131.9, 131.0, 129.62, 129.56, 128.3, 127.2, 120.9, 114.8, 113.8, 68.3, 55.0, 40.9, 34.4. ESI-MS (m/ z): 405.1 [M + H]+. HRMS (ESI) m/z: calcd for C24H24N2O4 (M + H)+ 405.180 88, found 405.1811. 4-(Hydroxy(phenyl)methyl)-N-((4-methoxyphenethyl)carbamoyl)benzamide (107). Ti(iPrO)4 (2 equiv, 1 mmol) was added dropwise over 5 min to a cloudy solution of 4-benzoyl-N-((4methoxyphenethyl)carbamoyl)benzamide (106, 1 equiv, 0.5 mmol) in 1,4-dioxane (1 M) under N2. Then, NaBH4 (1.5 equiv, 0.75 mmol) was added; lastly, MeOH (0.64 M) was added, and the reaction solution bubbled and turned clear. The reaction mixture was stirred at rt for 2 h until the reaction was complete as judged by TLC analysis. NH4OH and then water were added to the reaction solution. The reaction mixture was extracted into EtOAc, and the organic extracts were washed with brine, dried with Na2SO4, filtered, and concentrated under reduced pressure to give the alcohol product as a white solid in 35% yield (73.3 mg). 1H NMR (400 MHz, DMSO-d6) δ 10.59 (br s, 1H), 8.68 (t, J = 5.7 Hz, 1H), 7.90−7.86 (m, 2H), 7.51−7.46 (m, 2H), 7.40−7.36 (m, 2H), 7.33−7.27 (m, 2H), 7.24−7.18 (m, 1H), 7.18−7.13 (m, 2H), 6.88−6.83 (m, 2H), 6.06 (d, J = 4.0 Hz, 1H), 5.76 (d, J = 4.0 Hz, 1H), 3.71 (s, 3H), 3.46−3.39 (m, 2H), 2.74 (t, J = 7.1 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 168.0, 157.7, 153.5, 150.5, 145.1, 131.0, 130.9, 129.6, 128.2, 128.1, 127.0, 126.3, 126.1, 113.8, 73.8, 55.0, 40.8, 34.4. ESI-MS (m/z): 405.1 [M + H]+. 4-(Amino(phenyl)methyl)-N-((4-methoxyphenethyl)carbamoyl)benzamide (108). Step 1. Thionyl chloride (5 equiv, 0.58 mmol) was added to a solution of 4-(hydroxy(phenyl)methyl)-N-((4methoxyphenethyl)carbamoyl)benzamide (107, 1 equiv, 0.12 mmol) in CH2Cl2 (0.1 M). After 1.5 h, another 5 equiv of thionyl chloride were added. When the reaction was complete as judged by TLC analysis (3 more hours), the reaction mixture was concentrated and dried under vacuum to give 4-(chloro(phenyl)methyl)-N-((4methoxyphenethyl)carbamoyl)benzamide in 40.7 mg, 83% yield, which was used directly in the next step. 1H NMR (400 MHz, DMSO-d6) δ 10.69 (br s, 1H), 8.64 (t, J = 5.7 Hz, 1H), 7.96−7.92 (m, 2H), 7.61−7.56 (m, 2H), 7.50−7.46 (m, 2H), 7.42−7.37 (m, 2H), 7.35−7.30 (m, 1H), 7.19−7.14 (m, 2H), 6.88−6.84 (m, 2H), 6.60 (s, 1H), 3.72 (s, 3H), 3.46−3.40 (m, 2H), 2.75 (t, J = 7.3 Hz, 2H). ESI-MS (m/z): 422.9 [M + H]+. Step 2. Sodium azide (2 equiv, 0.19 mmol) was added to a solution of the chloride from Step 1 (1 equiv, 0.09 mmol) in DMF (0.1 M) and stirred overnight at rt. Then, water was added to the reaction, and a precipitate formed. The precipitate was collected by filtration, rinsed with water, and dried under vacuum to give 4-(azido(phenyl)methyl)N-((4-methoxyphenethyl)carbamoyl)benzamide in 74% yield (30.6 mg). ESI-MS (m/z): 430.1 [M + H]+. Step 3. Solid 10% Pd/C (1.8 equiv, 0.13 mmol) was added to a solution of the azide from Step 2 (1 equiv, 0.07 mmol) in anhydrous MeOH (0.1 M). After purging with N2, the suspension was stirred under H2 at 50 psi for 2.5 h. Then, the solution was filtered through Celite to remove Pd/C, and the filtrate was concentrated. The residue was purified by flash chromatography (silica gel, 0 to 20% MeOH in CH2Cl2) to give amine 108 as a white solid in 37% yield (10.7 mg). 1 H NMR (400 MHz, DMSO-d6) δ 10.58 (br s, 1H), 8.69 (t, J = 5.6 Hz, 1H), 7.89−7.85 (m, 2H), 7.54−7.49 (m, 2H), 7.42−7.37 (m, 2H), 7.31−7.25 (m, 2H), 7.20−7.17 (m, 1H), 7.17−7.14 (m, 2H), 6.88−6.84 (m, 2H), 5.14 (s, 1H), 3.71 (s, 3H), 3.46−3.39 (m, 2H), 2.74 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.9, 157.7, 153.5, 151.9, 146.2, 131.0, 130.5, 129.6, 128.2, 128.1, 126.8, 126.7, 126.6, 113.8, 59.0, 55.0, 40.8, 34.4. ESI-MS (m/z): 402.0 [M − H]−. 3-Bromo-N-((4-methoxyphenethyl)carbamoyl)benzamide (109). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 144.2 mg, 40% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.77 (br s, 1H), 8.58 (t, J = 5.6 Hz,

H2O, and MeOH (3:1:3 ratio, 0.3 M) were combined in a vial. Additional LiOH (6 mmol) was added after 1 h. After 1 h, 1 M HCl was added until the reaction mixture became acidic. A precipitate was collected by filtration, rinsed with water, and dried under vacuum to give the acid as a white solid, 561.0 mg, 82% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.74 (br s, 1H), 8.65 (br s, 1H), 7.99−7.88 (m, 4H), 7.19−7.15 (m, 2H), 6.89−6.85 (m, 2H), 3.72 (s, 3H), 3.47− 3.41 (m, 2H), 2.76 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 167.7, 166.7, 157.8, 153.3, 136.0, 131.0, 129.6, 129.4, 129.2, 128.3, 113.8, 55.0, 40.9, 34.4. ESI-MS (m/z): 342.9 [M + H]+. N-((4-Methoxyphenethyl)carbamoyl)-4-(morpholine-4carbonyl)benzamide (101). Carboxylic acid 100 (0.88 mmol, 1 equiv), EDC (0.92 mmol, 1.05 equiv), and HOBT (0.92 mmol, 1.05 equiv) were dissolved in DMF (0.1 M) and stirred for 20 min before the addition of morpholine (0.88 mmol, 1 equiv). The reaction mixture was stirred overnight. Water and EtOAc were added to the reaction mixture, and the organic layer was washed with 1 M NaOH (3×), water (3×), 1 M HCl, water (3×), and brine. Then, the organic layer was dried with Na2SO4, filtered, and concentrated to give the final product as a white solid, 3.8 mg, 12% yield. 1H NMR (400 MHz, CDCl3) δ 8.84 (br s, 1H), 8.65 (t, J = 5.6 Hz, 1H), 7.97−7.92 (m, 2H), 7.54−7.49 (m, 2H), 7.19−7.13 (m, 2H), 6.89−6.84 (m, 2H), 3.85−3.62 (m, 9H), 3.62−3.55 (m, 2H), 3.38 (br s, 2H), 2.86 (t, J = 7.2 Hz, 2H). ESI-MS (m/z): 412.1 [M + H]+. N-((4-Methoxyphenethyl)carbamoyl)-4-(piperidine-1-carbonyl)benzamide (102). The title compound was synthesized analogously to amide 101 to give the final product as a white solid, 15.7 mg, 38% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.75 (br s, 1H), 8.65 (t, J = 5.7 Hz, 1H), 8.00−7.95 (m, 2H), 7.48−7.43 (m, 2H), 7.20−7.15 (m, 2H), 6.89−6.85 (m, 2H), 3.72 (s, 3H), 3.59 (br s, 2H), 3.48−3.41 (m, 2H), 3.22 (br s, 2H), 2.76 (t, J = 7.3 Hz, 2H), 1.65−1.51 (m, 4H), 1.45 (br s, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.8, 167.7, 157.7, 153.4, 140.5, 133.1, 131.0, 129.6, 128.3, 126.5, 113.8, 55.0, 47.9, 40.9, 34.4, 25.9, 24.0. ESI-MS (m/z): 410.0 [M + H]+. 4-Benzoyl-N-((4-methoxyphenethyl)carbamoyl)benzamide (103). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 17.7 mg, 18% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.85 (br s, 1H), 8.64 (t, J = 5.6 Hz, 1H), 8.11−8.06 (m, 2H), 7.83−7.78 (m, 2H), 7.78−7.74 (m, 2H), 7.74−7.69 (m, 1H), 7.62−7.56 (m, 2H), 7.20−7.15 (m, 2H), 6.90−6.85 (m, 2H), 3.73 (s, 3H), 3.49−3.42 (m, 2H), 2.77 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 195.3, 167.6, 159.8, 157.7, 153.3, 140.4, 136.4, 135.8, 133.2, 130.9, 129.7, 129.5, 128.7, 128.4, 113.8, 55.0, 40.5, 33.7. ESI-MS (m/z): 403.0 [M + H]+. N-((4-Methoxyphenethyl)carbamoyl)-4-phenoxybenzamide (104). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 23.8 mg, 24% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.58 (br s, 1H), 8.69 (t, J = 5.7 Hz, 1H), 8.02−7.97 (m, 2H), 7.49−7.43 (m, 2H), 7.28−7.22 (m, 1H), 7.19−7.14 (m, 2H), 7.14−7.10 (m, 2H), 7.03−6.99 (m, 2H), 6.89−6.84 (m, 2H), 3.72 (s, 3H), 3.47−3.40 (m, 2H), 2.75 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 167.2, 160.9, 157.7, 155.0, 153.5, 131.0, 130.6, 130.3, 129.6, 126.8, 124.8, 120.0, 117.0, 113.8, 55.0, 40.8, 34.4. ESI-MS (m/z): 391.0 [M + H]+. HRMS (ESI) m/z: calcd for C23H22N2O4 (M + H)+ 391.165 23, found 391.1653. 4-(Benzyloxy)-N-((4-methoxyphenethyl)carbamoyl)benzamide (105). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 35.3 mg, 36% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.50 (br s, 1H), 8.73 (t, J = 5.7 Hz, 1H), 7.98−7.93 (m, 2H), 7.48−7.43 (m, 2H), 7.43−7.37 (m, 2H), 7.37−7.31 (m, 1H), 7.19−7.14 (m, 2H), 7.12−7.07 (m, 2H), 6.89−6.84 (m, 2H), 5.19 (s, 2H), 3.72 (s, 3H), 3.47−3.40 (m, 2H), 2.75 (t, J = 7.2 Hz, 2H). 13C NMR (400 MHz, DMSO-d6) δ 167.4, 161.9, 157.7, 153.7, 136.5, 131.0, 130.3, 129.6, 128.5, 128.0, 127.8, 124.7, 114.5, 113.8, 69.5, 55.0, 40.8, 34.4. ESI-MS (m/z): 405.1 [M + H]+. HRMS (ESI) m/z: calcd for C24H24N2O4 (M + H)+ 405.180 88, found 405.1815. N-((4-Methoxyphenethyl)carbamoyl)-4-(phenoxymethyl)benzamide (106). The title compound was synthesized according to General Procedure 3 to give the final product as a white solid, 26.5 5216



Article

4-Benzoyl-N-((4-methoxyphenethyl)(methyl)carbamoyl)-Nmethylbenzamide (113). Methyl iodide (0.37 mmol) was added to a solution of 4-benzoyl-N-((4-methoxyphenethyl)carbamoyl)benzamide (103, 0.07 mmol) and NaH (60% in mineral oil, 0.15 mmol) in DMF (0.2 M). When the reaction was complete as judged by TLC analysis (3 h), EtOAc was added, and the organic layer was washed with water and brine, dried with Na2SO4, filtered, and concentrated. The residue was purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give a colorless oil, 11.0 mg, 35% yield. 1H NMR (400 MHz, CDCl3) δ 7.82−7.78 (m, 2H), 7.78−7.74 (m, 2H), 7.74−7.63 (m, 2H), 7.63−7.57 (m, 1H), 7.50− 7.43 (m, 2H), 7.06−7.01 (m, 2H), 6.86−6.80 (m, 2H), 3.77 (s, 3H), 3.43−3.29 (m, 2H), 3.21 (s, 3H), 2.77 (s, 3H), 2.65 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, CDCl3) δ 196.0, 158.6, 158.0, 139.9, 138.8, 137.1, 133.0, 130.21, 130.17, 130.0, 129.8, 128.6, 128.5, 127.6, 114.3, 55.4, 51.2, 33.4, 32.1, 29.8. ESI-MS (m/z): 431.1 [M + H]+. N1-Cyclopropyl-N2-(4-hydroxyphenethyl)oxalamide (114). The title compound was prepared according to General Procedure 1 to provide 143.3 mg of a white solid in 96% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.18 (s, 1H), 8.77−8.65 (m, 2H), 6.98 (d, J = 8.0 Hz, 2H), 6.66 (d, J = 8.0 Hz, 2H), 3.32−3.24 (m, 2H), 2.79−2.70 (m, 1H), 2.65 (t, J = 7.2 Hz, 2H), 0.68−0.55 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 161.3, 159.7, 155.7, 129.5, 129.1, 115.1, 40.6, 33.9, 22.8, 5.4. ESI-MS (m/z): 249.1 [M + H]+. N1-(4-Chlorophenyl)-N2-(4-hydroxybenzyl)oxalamide (115). The title compound was prepared using the synthetic procedures described for the preparation of phenol 78 using methyl ether 80 (78.3 mg, 0.25 mmol, 1 equiv). The residue was purified by flash chromatography over silica gel (0 to 100% EtOAc in hexanes) to provide 33.0 mg of a white solid in 44% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.78 (s, 1H), 9.41 (t, J = 6.3 Hz, 1H), 7.85−7.80 (m, 2H), 7.43−7.38 (m, 2H), 7.13−7.08 (m, 2H), 6.72−6.67 (m, 2H), 4.26 (d, J = 6.3 Hz, 2H). 13C NMR (100 Hz, DMSO-d6) δ 159.8, 159.0, 156.5, 136.7, 129.1, 129.0, 128.8, 128.4, 122.2, 115.2, 42.3. ESIMS (m/z): 303.0 [M − H]−. N-(4-Chlorophenyl)-3-(4-hydroxyphenyl)propanamide (116). The title compound was prepared according to General Procedure 2, using 3-(4-hydroxyphenyl)propionic acid (50.0 mg, 0.3 mmol) and 4-chloroaniline (38.2 mg, 0.3 mmol). When water was added, a precipitate formed upon cooling to 0 °C. The precipitate was filtered and washed with cold water to yield 9.7 mg of a white solid in 12% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.01 (br s, 1H), 9.16 (br s, 1H), 7.62−7.57 (m, 2H), 7.36−7.31 (m, 2H), 7.04−6.99 (m, 2H), 6.68−6.63 (m, 2H), 2.81−2.75 (m, 2H), 2.58−2.52 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 170.7, 155.5, 138.2, 131.1, 129.1, 128.6, 126.5, 120.5, 115.1, 38.5, 30.0. ESI-MS (m/z): 276.1 [M + H]+. N1-(4-Chlorophenyl)-N2-(2-(4-hydroxyphenyl)propyl)oxalamide (117). The title compound was prepared using the synthetic procedures described for the preparation of phenol 78 using methyl ether 81 (11.8 mg, 0.34 mmol). The residue was purified by flash chromatography over silica gel (0% to 100% EtOAc) to yield 5.8 mg of a white solid in 51% yield. 1H NMR (400 MHz, DMSO-d6) 10.77 (br s, 1H), 9.19 (br s, 1H), 8.86 (t, J = 6.0 Hz, 1H), 7.87−7.82 (m, 2H), 7.42−7.38 (m, 2H), 7.05−7.00 (m, 2H), 6.70−6.66 (m, 2H), 3.32−3.21 (m, 2H), 3.00−2.90 (m, 1H), 1.16−1.11 (d, J = 7.0 Hz, 3H). 13C NMR (100 MHz, DMSO-d6) 159.6, 158.7, 155.7, 136.7, 134.4, 128.6, 128.2, 127.9, 121.9, 115.1, 46.3, 37.8, 19.5. ESI-MS (m/ z): 333.1 [M + H]+. N 1 -(4-Chlorophenyl)-N 2 -(1-(4-hydroxyphenyl)propan-2-yl)oxalamide (118). The title compound was prepared using the synthetic procedures described for the preparation of phenol 78 using methyl ether 82 (42.2 mg, 0.12 mmol). The crude product was purified by flash chromatography over silica gel (0 to 20% MeOH in CH2Cl2) to provide 9.3 mg of a white solid in 23% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.72 (br s, 1H), 9.16 (br s, 1H), 8.83 (d, J = 8.9 Hz, 1H), 7.86−7.81 (m, 2H), 7.42−7.38 (m, 2H), 7.01−6.96 (m, 2H), 6.66−6.62 (m, 2H), 4.07−3.95 (m, 1H), 2.76 (dd, J = 13.6, 8.0 Hz, 1H), 2.61 (dd, J = 13.6, 6.4 Hz, 1H), 1.12 (d, J = 6.6 Hz, 3H). 13 C NMR (100 Hz, DMSO-d6) δ 158.9, 158.8, 155.6, 136.7, 129.9,

1H), 8.11 (t, J = 1.8 Hz, 1H), 7.92 (ddd, J = 7.8, 1.6, 0.9 Hz, 1H), 7.81 (ddd, J = 8.0, 1.9, 0.9 Hz, 1H), 7.46 (t, J = 7.9 Hz, 1H), 7.19− 7.14 (m, 2H), 6.89−6.84 (m, 2H), 3.72 (s, 3H), 3.47−3.40 (m, 2H), 2.75 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 166.8, 157.7, 153.2, 135.3, 134.8, 131.0, 130.8, 130.7, 129.6, 127.2, 121.7, 113.8, 55.0, 40.9, 34.3. ESI-MS (m/z): 377.0 [M + H]+. N-((4-Methoxyphenethyl)carbamoyl)-[1,1′-biphenyl]-3-carboxamide (110). The title compound was synthesized from bromide 109 analogously to the synthesis of 97 to provide the product as a white solid, 3.0 mg, 5% yield. 1H NMR (500 MHz, DMSO-d6) δ 10.89 (br s, 1H), 8.73 (t, J = 5.7 Hz, 1H), 8.26 (t, J = 1.6 Hz, 1H), 7.93−7.89 (m, 2H), 7.82−7.79 (m, 2H), 7.60 (t, J = 7.8 Hz, 1H), 7.52−7.48 (m, 2H), 7.43−7.39 (m, 1H), 7.20−7.17 (m, 2H), 6.89−6.85 (m, 2H), 3.72 (s, 3H), 3.49−3.43 (m, 2H), 2.77 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 168.1, 157.7, 153.5, 140.3, 139.2, 133.2, 131.0, 130.8, 129.6, 129.2, 129.0, 127.9, 127.3, 127.0, 126.2, 113.8, 55.0, 40.9, 34.4. ESI-MS (m/z): 375.1 [M + H]+. 4-Benzoyl-N-((4-methoxyphenethyl)(methyl)carbamoyl)benzamide (111). Step 1. Ethyl chloroformate (1 equiv, 0.5 mmol) was added to a solution of 4-methoxyphenethylamine (1 equiv, 0.5 mmol) and pyridine (2 equiv, 1 mmol) in CH2Cl2 (0.5 M) at 0 °C. The solution warmed to rt. When the reaction was complete as judged by TLC analysis, the reaction was filtered to remove the solid. The filtrate was washed with 1 M HCl and brine, dried with Na2SO4, filtered, and concentrated to give ethyl (4-methoxyphenethyl)carbamate as a cream-colored solid, which was used without further purification, 147.4 mg, 99% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.12−7.07 (m, 2H), 6.86−6.82 (m, 2H), 3.95 (q, J = 7.1 Hz, 2H), 3.71 (s, 3H), 3.17−3.10 (m, 2H), 2.65−2.59 (m, 2H), 1.13 (t, J = 7.0 Hz, 3H). ESI-MS (m/z): 224.0 [M + H]+. Step 2. A solution of the carbamate from Step 1 (1 equiv, 0.5 mmol) in anhydrous THF (1.3 M) was added dropwise to a solution of LiAlH4 (6 equiv, 3 mmol) in anhydrous THF (0.4 M) at 0 °C under a nitrogen atmosphere. The reaction mixture warmed to rt after 45 min. After 4 h, the solution was heated to 50 °C overnight. Then, the reaction mixture was cooled to 0 °C and quenched by the slow addition of 0.1 mL of water, 0.1 mL of 1 M NaOH, and 0.3 mL of water. The reaction mixture was stirred for 1 h before an additional 0.5 mL of 1 M NaOH was added. The solid material was removed by filtration through Celite, and the filtrate was concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 0 to 10% MeOH in CH2Cl2) to give 2-(4-methoxyphenyl)N-methylethan-1-amine as a cream-colored solid in 27% yield (29.6 mg). 1H NMR (400 MHz, CDCl3) δ 7.15−7.10 (m, 2H), 6.86−6.82 (m, 2H), 3.79 (s, 3H), 2.87−2.81 (m, 2H), 2.81−2.75 (m, 2H), 2.00 (br s, 3H). ESI-MS (m/z): 166.1 [M + H]+. Step 3. The amine from Step 2 was used to prepare N-acyl urea 111 according to General Procedure 3. The crude reaction mixture was purified by flash chromatography (silica gel, 0 to 100% EtOAc in hexanes) to give the final product as a solid, 1.7 mg, 3% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.99−7.85 (m, 2H), 7.83−7.78 (m, 2H), 7.78−7.75 (m, 2H), 7.74−7.69 (m, 1H), 7.62−7.57 (m, 2H), 7.20−7.13 (m, 2H), 6.89−6.83 (m, 2H), 3.71 (s, 3H), 3.53−3.47 (m, 2H), 2.93 (s, 3H), 2.79 (t, J = 7.7 Hz, 2H). ESI-MS (m/z): 417.1 [M + H]+. 4-Benzoyl-N-((4-methoxyphenethyl)carbamoyl)-N-methylbenzamide (112). Methyl iodide (0.37 mmol) was added to a solution of 4-benzoyl-N-((4-methoxyphenethyl)carbamoyl)benzamide (103, 0.07 mmol) and K2CO3 (0.22 mmol) in DMF (0.2 M). When the reaction was complete as judged by TLC analysis (3 h), EtOAc was added, and the organic layer was washed with water and brine, dried with Na2SO4, and concentrated to give a white solid, 23.3 mg, 73% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.54 (t, J = 5.4 Hz, 1H), 7.78−7.72 (m, 4H), 7.72−7.67 (m, 1H), 7.64−7.59 (m, 2H), 7.59− 7.53 (m, 2H), 7.13−7.08 (m, 2H), 6.87−6.82 (m, 2H), 3.69 (s, 3H), 3.32−3.25 (m, 2H), 3.12 (s, 3H), 2.61 (t, J = 7.3 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 195.2, 171.2, 157.7, 155.3, 139.9, 138.4, 136.6, 133.0, 130.9, 129.7, 129.6, 129.4, 128.7, 127.0, 113.8, 55.0, 41.9, 33.9, 33.7. ESI-MS (m/z): 417.1 [M + H]+. 5217



Article

to N-acyl urea 120 to yield 30.9 mg of a white solid in 34% yield. 1H NMR (400 MHz, DMSO-d6) δ 8.62 (t, J = 5.6 Hz, 1H), 8.08−8.03 (m, 2H), 7.79−7.74 (m, 2H), 7.52−7.47 (m, 1H), 7.39 (d, J = 7.7 Hz, 1H), 7.35−7.30 (m, 2H), 7.20−7.14 (m, 2H), 6.89−6.84 (m, 2H), 3.72 (s, 3H), 3.48−3.41 (m, 2H), 2.76 (t, J = 7.3 Hz, 2H), 2.25 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 197.5, 167.8, 157.9, 153.5, 140.3, 137.7, 136.8, 136.4, 131.3, 131.13, 131.07, 129.8, 129.7, 128.8, 128.7, 125.8, 114.0, 55.2, 41.1, 34.5, 19.8. ESI-MS (m/z): 415.1 [M − H]−. N-((4-Hydroxyphenethyl)carbamoyl)-4-(phenoxymethyl)benzamide (122). The title compound was synthesized according to General Procedure 3, except the addition of tyramine was carried out at 80 °C. The final product was isolated by filtration as a white solid, 111.9 mg, 65% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.66 (br s, 1H), 9.23 (br s, 1H), 8.67 (t, J = 5.6 Hz, 1H), 7.96 (d, J = 8.2 Hz, 2H), 7.55 (d, J = 8.2 Hz, 2H), 7.33−7.26 (m, 2H), 7.06−6.99 (m, 4H), 6.97−6.92 (m, 1H), 6.72−6.67 (m, 2H), 5.19 (s, 2H), 3.45− 3.38 (m, 2H), 2.70 (t, J = 7.2 Hz, 2H). 13C NMR (100 MHz, DMSOd6) δ 167.9, 158.1, 155.7, 153.5, 142.1, 131.9, 129.6, 129.5, 129.1, 128.3, 127.2, 120.9, 115.2, 114.8, 68.3, 41.0, 34.4. ESI-MS (m/z): 391.1 [M + H]+. HRMS (ESI) m/z: calcd for C23H22N2O4 (M + H)+ 391.165 23, found 391.1655. N1-(4-Hydroxyphenethyl)-N2-(4-(4-(prop-2-yn-1-yloxy)benzoyl)phenyl)oxalamide (123). Step 1. To a solution of (4-aminophenyl)(4-(prop-2-yn-1-yloxy)phenyl)methanone (135, 654 mg, 2.60 mmol, 1 equiv) and triethylamine (0.72 mL, 5.2 mmol, 2 equiv) dissolved in anhydrous methylene chloride (26 mL) was slowly added a solution of ethyl chlorooxoacetate (0.29 mL, 2.60 mmol, 1 equiv) in anhydrous methylene chloride (2.9 mL). The reaction was stirred for 16 h at room temperature and then washed with saturated aqueous sodium bicarbonate. The organic layer was dried over sodium sulfate and concentrated under vacuum to yield the intermediate ethyl 2-oxo2-((4-(4-(prop-2-yn-1-yloxy) benzoyl)phenyl)amino)acetate, which was used immediately in the next step without purification. 1H NMR (400 MHz, CDCl3-d) δ 9.09 (s, 1H), 7.85−7.74 (m, 7H), 7.07−7.03 (m, 2H), 4.78 (d, J = 2.4 Hz, 2H), 4.41 (dq, J = 21.1, 7.2 Hz, 4H), 2.57 (t, J = 2.4 Hz, 1H), 1.42 (dt, J = 17.9, 7.1 Hz, 6H). Step 2. The ethyl ester from Step 1 (93 mg, 0.3 mmol) was dissolved in THF (1 mL) and treated with aqueous 1 M lithium hydroxide (1 mL). The resulting mixture was stirred for 16 h at room temperature. The volatiles were removed by passing a stream of dry nitrogen over the mixture, and the resulting residue was dissolved in water (1 mL) and acidified with 1 M HCl, whereupon a precipitate formed. The precipitate was filtered and dried by aspiration to give 2oxo-2-((4-(4-(prop-2-yn-1-yloxy)benzoyl)phenyl)amino)acetic acid as a tan powder, 48 mg, 55% yield. 1H NMR (400 MHz, DMSOd6) δ = 10.97 (s, 1H), 7.94 (d, J = 8.4 Hz, 1H), 7.78−7.68 (m, 2H), 7.66−7.57 (m, 2H), 7.53−7.45 (m, 2H), 7.16−7.03 (m, 3H), 6.64− 6.54 (m, 2H), 4.90 (dd, J = 12.3, 2.4 Hz, 3H), 3.63 (dt, J = 6.9, 2.3 Hz, 1H); 13C NMR (100 MHz, DMSO-d6) δ 193.77, 192.74, 162.34, 161.05, 160.03, 153.87, 141.90, 133.53, 132.80, 132.75, 132.34, 131.70, 131.54, 131.00, 130.74, 124.58, 119.97, 115.09, 114.75, 112.94, 79.35, 79.22, 79.18, 79.07, 56.16, 56.04, 40.58, 40.37, 40.22, 40.16, 39.95, 39.74, 39.54, 39.33; ESI-MS (m/z): 322.0 [M − H]−. Step 3. The acid from Step 2 (26 mg, 0.08 mmol) was combined with HATU (37 mg, 0.1 mmol) and tyramine (12 mg, 0.09 mmol) and dissolved in DMF (300 μL) and triethylamine (33 μL, 0.24 mmol). The resulting mixture was stirred for 16 h at room temperature and diluted with water, and the resulting precipitate was filtered, triturated with ether/methanol, and purified by flash chromatography over silica gel to give N-acyl urea 123, 3.2 mg, 9% yield. 1H NMR (400 MHz, DMSO-d6) δ 10.96 (s, 1H), 9.19 (s, 1H), 9.05 (t, J = 6.0 Hz, 1H), 8.03−7.98 (m, 2H), 7.78−7.70 (m, 2H), 7.16−7.11 (m, 2H), 7.04−6.99 (m, 2H), 6.70−6.66 (m, 2H), 4.93 (d, J = 2.2 Hz, 2H), 3.66 (t, J = 2.2 Hz, 1H), 3.42−3.35 (m, 2H), 2.72 (t, J = 7.5 Hz, 2H). 13C NMR (100 MHz, DMSO-d6) δ 161.06, 159.94, 159.49, 156.14, 141.70, 133.63, 132.35, 130.94, 130.72, 129.92, 129.49, 120.17, 115.59, 115.10, 103.70, 79.22, 79.18, 56.16, 41.42,

129.0, 128.6, 128.2, 121.9, 115.0, 47.3, 40.4, 19.9. ESI-MS (m/z): 331.0 [M − H]−. N-(4-Hydroxyphenethyl)quinoline-2-carboxamide (119). The title compound was prepared according to General Procedure 2 using quinaldic acid (56.0 mg, 0.3 mmol) and tyramine (41.1 mg, 0.3 mmol) to yield 34.6 mg of a white solid in 40% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.20 (br s, 1H), 8.94 (t, J = 6.1 Hz, 1H), 8.56 (d, J = 8.5 Hz, 1H), 8.15 (d, J = 8.5 Hz, 1H), 8.13−8.06 (m, 2H), 7.90− 7.84 (m, 1H), 7.75−7.69 (m, 1H), 7.09−7.04 (m, 2H), 6.72−6.67 (m, 2H), 3.58−3.49 (m, 2H), 2.83−2.76 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 163.8, 155.7, 150.2, 146.0, 137.9, 130.5, 129.5, 129.4, 129.1, 128.8, 128.1, 128.0, 118.6, 115.2, 40.9, 34.4. ESI-MS (m/z): 293.1 [M + H]+. N-((4-Hydroxyphenethyl)carbamoyl)-4-(2-methylbenzoyl)benzamide (120). Step 1. A 1.9 M solution of o-tolylmagnesium bromide in THF (2.4 mmol, 1.2 equiv) was added dropwise to a solution of 4-formylbenzonitrile (2 mmol, 1 equiv) in anhydrous THF (0.1 M) at −78 °C. After 2 h at −78 °C, the solution was warmed to room temperature, and the solution was stirred at room temperature overnight. Then, the solution was washed with NH4Cl(aq) and extracted into EtOAc (3×). The organic extracts were washed with brine, dried over Na2SO4, filtered, and concentrated under reduced pressure. The crude residue was purified by flash chromatography over silica gel (1:1 EtOAc/hexanes) to yield 425.8 mg of 4(hydroxy(o-tolyl)methyl)benzonitrile as a yellow oil in 95% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.79−7.75 (m, 2H), 7.50−7.46 (m, 2H), 7.37−7.34 (m, 1H), 7.22−7.11 (m, 3H), 6.05 (d, J = 4.4 Hz, 1H), 5.92 (d, J = 4.4 Hz, 1H), 2.22 (s, 3H). 13C NMR (100 Hz, DMSO-d6) δ 150.1, 142.1, 135.0, 132.1, 130.3, 127.7, 127.2, 126.9, 125.9, 118.9, 109.5, 71.0, 19.1. Step 2. The nitrile from Step 1 (425.8 mg, 1.9 mmol, 1 equiv) and KOH (2.6 g, 34 mmol, 18 equiv) were dissolved in anhydrous tbutanol (6.4 mL, 0.3 M), and the solution was heated at 60 °C for 4 h. The solution was then cooled, and water was added. The solution was extracted into EtOAc (3×), dried over Na2SO4, filtered, and concentrated under reduced pressure to yield 340.5 mg of 4(hydroxy(o-tolyl)methyl)benzamide in 80% yield. 1H NMR (400 MHz, DMSO-d6) δ 7.90 (br s, 1H), 7.81−7.77 (m, 2H), 7.43−7.39 (m, 1H), 7.37−7.32 (m, 2H), 7.30 (br s, 1H), 7.22−7.09 (m, 3H), 5.88 (s, 2H), 2.21 (s, 3H). Step 3. Pyridinium chlorochromate (PCC, 4.6 mmol, 3 equiv) was added to a solution of the alcohol from Step 2 (1.5 mmol, 1 equiv) in anhydrous CH2Cl2 (15 mL, 0.1 M), and the reaction was stirred at room temperature. After 1.5 h, more PCC (3 equiv) was added, and the reaction was finished in 1.5 h as determined by TLC. The reaction was filtered through Celite and washed with CH2Cl2, and the filtrate was concentrated under reduced pressure. The residue was purified by flash chromatography over silica gel (3:4 EtOAc/hexanes) to yield 51.8 mg of 4-(2-methylbenzoyl)benzamide as a white solid in 14% yield. ESI-MS (m/z): 240.1 [M + H]+. Step 4. N-((4-Hydroxyphenethyl)carbamoyl)-4-(2methylbenzoyl)benzamide (120) was prepared according to General Procedure 3. To form the N-acyl urea, tyramine (55.4 mg, 0.39 mmol) was dissolved in acetonitrile (1 mL, 0.1 M) by heating to 80 °C, and then a solution of the isocyanate in CH3CN (1 mL, 0.1 M) was added. The reaction mixture was stirred overnight at 80 °C before being concentrated. The residue was extracted with water (3 mL) and EtOAc (3 × 3 mL), and the organic extracts were washed with brine, dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by flash chromatography over silica gel (0 to 40% EtOAc in hexanes) to provide 33.4 mg of a yellow oil in 43% yield. 1H NMR (400 MHz, CDCl3) δ 7.84−7.79 (m, 2H), 7.77−7.72 (m, 2H), 7.45−7.39 (m, 1H), 7.34−7.27 (m, 3H), 7.11−7.07 (m, 2H), 6.83−6.78 (m, 2H), 6.22 (t, J = 5.6 Hz, 1H), 3.73−3.66 (m, 2H), 2.87 (t, J = 6.8 Hz, 2H), 2.33 (s, 3H). 13C NMR (100 MHz, CDCl3) δ 198.0, 166.8, 154.9, 140.2, 138.6, 138.0, 137.2, 131.4, 130.9, 130.7, 130.43, 130.41, 130.0, 128.9, 127.1, 125.5, 115.8, 41.6, 34.8, 20.3. ESI-MS (m/z): 403.0 [M − H]+. N-((4-Methoxyphenethyl)carbamoyl)-4-(2-methylbenzoyl)benzamide (121). The title compound was synthesized analogously 5218



Article

Treated or untreated cell lysate (250 μL) was aliquoted into Eppendorf tubes. Blank lysates were spiked with varying concentrations of each compound to create a standard curve. Each sample was mixed with 0.5 mL of methanol, vortexed for 15 s, incubated for 10 min at RT, and then centrifuged twice at 16 100g. The supernatant was dried, resuspended in 175 μL of a 1:1 mixture of dH20/methanol containing 0.1% formic acid and 100 ng/mL tolbutamide, and analyzed by LC-MS/MS as described above. The length and width of trypsinized H1155 cells was measured by light microscopy, and an estimate of the intracellular volume was calculated as volume = π /6 × L × W 2 . The total compound in lysate was then divided by the total intracellular volume represented by the cells in that lysate to estimate the final intracellular compound concentration.

40.57, 40.36, 40.15, 39.94, 39.73, 39.53, 39.32, 34.23. ESI-MS (m/z): 441.1 [M − H]−. Cell Culture and Toxicity Testing. All NSCLC cell lines were obtained from the Minna and Gazdar laboratories at UT Southwestern Medical Center. They were screened for mycoplasma by PCR and authenticated by short-tandem-repeat (STR) analysis through the McDermott Core at UT Southwestern. All NSCLC cell lines were cultured in RPMI 1640 (Sigma) supplemented with 5% FBS (Sigma) and 2 mM L-glutamine (Sigma). For the dose−response analyses, cells were plated in 96-well plates at 15% confluence in 100 μL of the above medium and allowed to adhere overnight. On the next day, this medium was removed, and new medium containing 1 of 10 concentrations of the compound was added, starting from 50 μM and decreasing in 3-fold serial dilutions in DMSO. The final concentration of DMSO in each well was 0.5%. Each dose of compound was tested in duplicate, and the values displayed represent the averages of these duplicates. In Vitro SCD Assay. Mouse liver were collected from female CD1 mice sacrificed by inhalation overdoses of CO2 followed by cervical dislocation. Livers were diced and incubated on ice for 30 min in a buffer containing 50 mM HEPES (pH 8.0), 2 mM MgCl2, and 10 mM KCl supplemented with the SIGMAFAST protease-inhibitor cocktail (Sigma). The liver was then homogenized with 10 strokes in a dounce homogenizer. The resulting homogenate was centrifuged at 12 000g to remove the heavy membrane fraction, and the supernatant was used for reactions. Either DMSO or a compound was added to a 100 μL reaction containing 250 μg of protein, 2 mM NADPH (Sigma, from a 100 mM stock solution solubilized in 10 mM NaOH), and 50 nM Stearoyl [9,10-3H] CoA (American Radiolabeled Chemical, from a 2 μM stock solution in a 1:1 EtOH/0.1 M sodium acetate solution). Reactions were incubated for 2 h at 37 °C and then transferred to a 40% charcoal solution in water (JT Baker) and incubated for another 20 min. The charcoal−reaction mixture was filtered through a Spin-XCentrifuge filter (#8160), and the filtrate was transferred to scintillation vials containing 10 mL of scintillation fluid (RPI). Counts per minute were measured by a Beckman LS 6000IC scintillation counter. Cellular and S9-Stability Assay. Methylated-prodrug and demethylated-drug compound levels for metabolic stability and cellaccumulation studies were monitored by LC-MS/MS using an AB Sciex 4000 Qtrap mass spectrometer coupled to a Shimadzu Prominence LC. Analytes were detected with the mass spectrometer in positive multiple-reaction-monitoring (MRM) mode by following the precursor to the fragment-ion transitions indicated here: 58: 263.152/135.163, 114: 249.162/121.000, 80: 317.03/125.9, 115: 303.001/125.970, 48: 307.147/135.098, 119: 293.130/121.109, 160: 405.162/228.123, 122: 391.140/228.121, 15: 457.099/135.046, and 123: 443.169/146.040. An Agilent C18 XDB column (5 μm, 50 × 4.6 mm) was used for chromatography for all compounds with the following conditions: Buffer A was dH20 with 0.1% formic acid, and Buffer B was methanol with 0.1% formic acid. The gradient was 0−1.0 min, 3% B; 1.0−1.5 min, gradient to 100% B; 1.5−3.0 min, 100% B; 3.0−3.1 min, gradient to 3% B; and 3.1−4.0 min, 3% B. Tolbutamide (Sigma) was used as an internal standard (IS, transition of 269.1/ 169.9 for negative mode and 271.2/91.2 for positive mode). Ammonium acetate was added at 2−5 mM as necessary to buffers to prevent the formation of sodium adducts. Cell culture and S9 studies were conducted as described in ref 21. Methylated prodrug was incubated with H1155 cells expressing Cyp4F11 or the vector only at a concentration of 500 nM. Compound Accumulation in H1155 Cells. H1155 cells were seeded in 6 cm dishes at 500 000 cells per well. After 18 h, the media was replaced with fresh media, and cells were treated with a 500 nM concentration of a compound. After 4 h of incubation with a compound, cells were washed three times with ice-cold PBS, harvested using trypsin, and counted on ice. Each sample was then centrifuged, resuspended at a million cells per milliliter, and snap frozen. All samples were stored at −80 °C until analyses were performed.

■

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jmedchem.8b00052. 1 H and 13C NMR spectra for synthetic compounds, 95% confidence intervals for EC50 values, and time-course studies for conversion from drug to prodrug forms (CSV) Molecular-formula strings (PDF)

■

AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected] (D.N.). *E-mail: [email protected] (J.M.R.). ORCID

Joseph M. Ready: 0000-0003-1305-9581 Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS Funding was provided by CPRIT (RP110708-P3), NIH (R01 CA216863, R01 CA217333), and the Welch Foundation (I1612). This research was also supported by a Harold C. Simmons Cancer Center Startup Award, a Disease Oriented Clinical Scholar (DOCS) award, a Damon Runyon Clinical Investigator award (CI-68-13), and a grant from the Welch Foundation (I-1879) to D.N. Pano Theodoropoulos is acknowledged for helpful discussions, and Meghan Campbell and Vo Lam are acknowledged for synthetic work.

■

ABBREVIATIONS USED Ac, acetate;; ADME, adsorption, distribution, metabolism, excretion; ATP, adenosine triphosphate; CYP, cytochrome P450; DMF, dimethylformamide; DME, 1,2-dimethoxyethane; DMSO, dimethyl sulfoxide; Dppf, 1,1′-bis(diphenylphosphino)ferrocene; EDC, 1-(3-(dimethylamino)propyl)-3-ethylcarbodiimide HCl; FBS, fetal bovine serum; HATU, 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate; HEPES, 4-(2hydroxyethyl)-1-piperazineethanesulfonic acid; HOBt, 1-hydroxybenzotriazole hydrate; HTS, high-throughput screening; NADPH, nicotinamide adenine dinucleotide phosphate; NSCLC, non-small-cell lung cancer; SAR, structure−activity relationship; PCC, pyridinium chlorochromate; SCD, stearoylCoA desaturase; SDS-PAGE, sodium dodecyl sulfate−polyacrylamide-gel electrophoresis; THF, tetrahydrofuran; TLC, thin-layer chromatography; UFA, unsaturated fatty acid 5219



■

Article

thiazolylpyridinone scd1 inhibitors with preferential liver distribution and reduced mechanism-based adverse effects. Bioorg. Med. Chem. Lett. 2014, 24, 526−531. (15) Oballa, R. M.; Belair, L.; Black, W. C.; Bleasby, K.; Chan, C. C.; Desroches, C.; Du, X.; Gordon, R.; Guay, J.; Guiral, S.; Hafey, M. J.; Hamelin, E.; Huang, Z.; Kennedy, B.; Lachance, N.; Landry, F.; Li, C. S.; Mancini, J.; Normandin, D.; Pocai, A.; Powell, D. A.; Ramtohul, Y. K.; Skorey, K.; Sørensen, D.; Sturkenboom, W.; Styhler, A.; Waddleton, D. M.; Wang, H.; Wong, S.; Xu, L.; Zhang, L. Development of a liver-targeted stearoyl-CoA desaturase (SCD) inhibitor (MK-8245) to establish a therapeutic window for the treatment of diabetes and dyslipidemia. J. Med. Chem. 2011, 54, 5082−5096. (16) Zhang, Z.; Sun, S.; Kodumuru, V.; Hou, D.; Liu, S.; Chakka, N.; Sviridov, S.; Chowdhury, S.; McLaren, D. G.; Ratkay, L. G.; Khakh, K.; Cheng, X.; Gschwend, H. W.; Kamboj, R.; Fu, J.; Winther, M. D. Discovery of piperazin-1-ylpyridazine-based potent and selective stearoyl-CoA desaturase-1 inhibitors for the treatment of obesity and metabolic syndrome. J. Med. Chem. 2013, 56, 568−583. (17) (a) Miyazaki, M.; Man, W. C.; Ntambi, J. M. Targeted disruption of stearoyl-CoA desaturase1 gene in mice causes atrophy of sebaceous and meibomian glands and depletion of wax esters in the eyelid. J. Nutr. 2001, 131, 2260−2268. (b) Meingassner, J. G.; Aschauer, H.; Winiski, A. P.; Dales, N.; Yowe, D.; Winther, M. D.; Zhang, Z.; Stütz, A.; Billich, A. Pharmacological inhibition of stearoylCoA desaturase in the skin induces atrophy of the sebaceous glands. J. Invest. Dermatol. 2013, 133, 2091−2094. (18) Lee, S.-H.; Dobrzyn, A.; Dobrzyn, P.; Rahman, S. M.; Miyazaki, M.; Ntambi, J. M. Lack of stearoyl-CoA desaturase 1 upregulates basal thermogenesis but causes hypothermia in a cold environment. J. Lipid Res. 2004, 45, 1674−1682. (19) Miyazaki, M.; Flowers, M. T.; Sampath, H.; Chu, K.; Otzelberger, C.; Liu, X.; Ntambi, J. M. Hepatic stearoyl-CoA desaturase-1 deficiency protects mice from carbohydrate-induced adiposity and hepatic steatosis. Cell Metab. 2007, 6, 484−496. (20) Merck Sharp & Dhome Corporation. Pharmacokinetics and Pharmacodynamics of MK-8245 in Participants With Type 2 Diabetes (MK-8245-012). https://clinicaltrials.gov/ct2/show/results/ NCT00972322 (accessed May 1, 2018). (21) Theodoropoulos, P. C.; Gonzales, S. S.; Winterton, S. E.; Rodriguez-Navas, C.; McKnight, J. S.; Morlock, L. K.; Hanson, J. M.; Cross, B.; Owen, A. E.; Duan, Y.; Moreno, J. R.; Lemoff, A.; Mirzaei, H.; Posner, B. A.; Williams, N. S.; Ready, J. M.; Nijhawan, D. Discovery of tumor-specific irreversible inhibitors of stearoyl CoA desaturase. Nat. Chem. Biol. 2016, 12, 218−225. (22) The results published here are fully or partially based upon data generated by the Cancer Target Discovery and Development (CTD2) Network established by the National Cancer Institute’s Office of Cancer Genomics. See ‘Lung Cancer Oncogenotype-Selective Drug Target Discovery’ at https://ctd2.nci.nih.gov/dataPortal/ (accessed May 1, 2018). (b) McMillan, E. A.; Ryu, M.-J.; Diep, C. H.; Mendiratta, S.; Clemenceau, J. R.; Vaden, R. M.; Kim, J.-H.; Motoyaji, T.; Covington, K. R.; Peyton, M.; Huffman, K.; Wu, X.; Girard, L.; Sung, Y.; Chen, P.-H.; Mallipeddi, P. L.; Lee, J. Y.; Hanson, J.; Voruganti, S.; Yu, Y.; Park, S.; Sudderth, J.; DeSevo, C.; Muzny, D. M.; Doddapaneni, H.; Gazdar, A.; Gibbs, R. A.; Hwang, T.-H.; Heymach, J. V.; Wistuba, I.; Coombes, K. R.; Williams, N. S.; Wheeler, D. A.; MacMillan, J. B.; Deberardinis, R. J.; Roth, M. G.; Posner, B. A.; Minna, J. D.; Kim, H. S.; White, M. A. Chemistry-first approach for nomination of personalized treatment in lung cancer. Cell 2018, 173, 864−878. (23) Rodriguez-Antona, C.; Ingelman-Sundberg, M. Cytochrome P450 pharmacogenetics and cancer. Oncogene 2006, 25, 1679−1691. (24) The 95% confidence intervals for the EC50 values are provided in the Supporting Information. (25) We speculate that the two bands vary in terms of posttranslational modifications. (26) Intracellular compound concentration was determined after a 24 h incubation with 500 nM drug in H1155 cells to avoid

REFERENCES

(1) (a) Ackerman, D.; Simon, M. C. Hypoxia, lipids, and cancer: Surviving the harsh tumor microenvironment. Trends Cell Biol. 2014, 24, 472−478. (b) Listenberger, L. L.; Han, X.; Lewis, S. E.; Cases, S.; Farese, R. V.; Ory, D. S.; Schaffer, J. E. Triglyceride accumulation protects against fatty acid-induced lipotoxicity. Proc. Natl. Acad. Sci. U. S. A. 2003, 100, 3077−3082. (2) Paton, C. M.; Ntambi, J. M. Biochemical and physiological function of stearoyl-coA desaturase. Am. J. Physiol. Endocrinol. Metab. 2009, 297, E28−E37. (3) Zhang, L.; Ge, L.; Parimoo, S.; Stenn, K.; Prouty, S. M. Human stearoyl-CoA desaturase: Alternative transcripts generated from a single gene by usage of tandem polyadenylation sites. Biochem. J. 1999, 340, 255−264. (4) Peck, B.; Schug, Z. T.; Zhang, Q.; Dankworth, B.; Jones, D. T.; Smethurst, E.; Patel, R.; Mason, S.; Jiang, M.; Saunders, R.; Howell, M.; Mitter, R.; Spencer-Dene, B.; Stamp, G.; McGarry, L.; James, D.; Shanks, E.; Aboagye, E. O.; Critchlow, S. E.; Leung, H. Y.; Harris, A. L.; Wakelam, M. J. O.; Gottlieb, E.; Schulze, A. Inhibition of fatty acid desaturation is detrimental to cancer cell survival in metabolically compromised environments. Cancer Metab. 2016, 4, 1−18. (5) Kim, H.; Rodriguez-Navas, C.; Kollipara, R. K.; Kapur, P.; Pedrosa, I.; Brugarolas, J.; Kittler, R.; Ye, J. Unsaturated fatty acids stimulate tumor growth through stabilization of β-catenin. Cell Rep. 2015, 13, 495−503. (6) Zhang, Z.; Dales, N. A.; Winther, M. D. Opportunities and challenges in developing stearoyl-coenzyme A desaturase-1 inhibitors as novel therapeutics for human disease. J. Med. Chem. 2014, 57, 5039−5056. (7) Xin, Z.; Zhao, H.; Serby, M. D.; Liu, B.; Liu, M.; Szczepankiewicz, B. G.; Nelson, L. T. J.; Smith, H. T.; Suhar, T. S.; Janis, R. S.; Cao, N.; Camp, H. S.; Collins, C. A.; Sham, H. L.; Surowy, T. K.; Liu, G. Discovery of piperidine-aryl urea-based stearoyl-coa desaturase 1 inhibitors. Bioorg. Med. Chem. Lett. 2008, 18, 4298−4302. (8) Roongta, U. V.; Pabalan, J. G.; Wang, X.; Ryseck, R.-P.; Fargnoli, J.; Henley, B. J.; Yang, W.-P.; Zhu, J.; Madireddi, M. T.; Lawrence, R. M.; Wong, T. W.; Rupnow, B. A. Cancer cell dependence on unsaturated fatty acids implicates stearoyl-CoA desaturase as a target for cancer therapy. Mol. Cancer Res. 2011, 9, 1551−1561. (9) von Roemeling, C. A.; Marlow, L. A.; Wei, J. J.; Cooper, S. J.; Caulfield, T. R.; Wu, K.; Tan, W. W.; Tun, H. W.; Copland, J. A. Stearoyl-CoA desaturase 1 is a novel molecular therapeutic target for clear cell renal cell carcinoma. Clin. Cancer Res. 2013, 19, 2368−2380. (10) Liu, G.; Lynch, J. K.; Freeman, J.; Liu, B.; Xin, Z.; Zhao, H.; Serby, M. D.; Kym, P. R.; Suhar, T. S.; Smith, H. T.; Cao, N.; Yang, R.; Janis, R. S.; Krauser, J. A.; Cepa, S. P.; Beno, D. W. A.; Sham, H. L.; Collins, C. A.; Surowy, T. K.; Camp, H. S. Discovery of potent, selective, orally bioavailable stearoyl-CoA desaturase 1 inhibitors. J. Med. Chem. 2007, 50, 3086−3100. (11) Mason, P.; Liang, B.; Li, L.; Fremgen, T.; Murphy, E.; Quinn, A.; Madden, S. L.; Biemann, H.-P.; Wang, B.; Cohen, A.; Komarnitsky, S.; Jancsics, K.; Hirth, B.; Cooper, C. G. F.; Lee, E.; Wilson, S.; Krumbholz, R.; Schmid, S.; Xiang, Y.; Booker, M.; Lillie, J.; Carter, K. SCD1 inhibition causes cancer cell death by depleting mono-unsaturated fatty acids. PLoS One 2012, 7, No. e33823. (12) Koltun, D. O.; Parkhill, E. Q.; Vasilevich, N. I.; Glushkov, A. I.; Zilbershtein, T. M.; Ivanov, A. V.; Cole, A. G.; Henderson, I.; Zautke, N. A.; Brunn, S. A.; Mollova, N.; Leung, K.; Chisholm, J. W.; Zablocki, J. Novel, potent, selective, and metabolically stable stearoylCoA desaturase (SCD) inhibitors. Bioorg. Med. Chem. Lett. 2009, 19, 2048−2052. (13) Hess, D.; Chisholm, J. W.; Igal, R. A. Inhibition of stearoyl-CoA desaturase activity blocks cell cycle progression and induces programmed cell death in lung cancer cells. PLoS One 2010, 5, No. e11394. (14) Sun, S.; Zhang, Z.; Raina, V.; Pokrovskaia, N.; Hou, D.; Namdari, R.; Khakh, K.; Ratkay, L. G.; McLaren, D. G.; Mork, M.; Fu, J.; Ferreira, S.; Hubbard, B.; Winther, M. D.; Dales, N. Discovery of 5220



Article

complications from toxicity. Although differences in accumulation were observed, the most highly accumulated compounds in this study showed little toxicity to H1155 cells, ruling out poor permeability as an explanation for lack of toxicity. 58: 80 ± 40 nM, 80: 340 ± 100 nM, 106: 1670 ± 560 nM, 15: 870 ± 200. (27) Lam, S. S.; Martell, J. D.; Kamer, K. J.; Deerinck, T. J.; Ellisman, M. H.; Mootha, V. K.; Ting, A. Y. Directed evolution of APEX2 for electron microscopy and proximity labeling. Nat. Methods 2015, 12, 51−54. (28) Bartoli, G.; Bosco, M.; Sambri, L.; Marcantoni, E. Cerium(III) chloride mediated Michael addition of RMgX to nitroenes: A very efficient access to complex nitroalkanes. Tetrahedron Lett. 1994, 35, 8651−8654. (29) Dehmlow, H.; Aebi, J. D.; Jolidon, S.; Ji, Y.-H.; von der Mark, E. M.; Himber, J.; Morand, O. H. Synthesis and structure−activity studies of novel orally active non-terpenoic 2,3-oxidosqualene cyclase inhibitors. J. Med. Chem. 2003, 46, 3354−3370. (30) Qiao, C.; Gupte, A.; Boshoff, H. I.; Wilson, D. J.; Bennett, E. M.; Somu, R. V.; Barry, C. E.; Aldrich, C. C. 5‘-O-[(Nacyl)sulfamoyl]adenosines as antitubercular agents that inhibit MbtA: An adenylation enzyme required for siderophore biosynthesis of the mycobactins. J. Med. Chem. 2007, 50, 6080−6094. (31) Shani, J.; Gazit, A.; Livshitz, T.; Biran, S. Synthesis and receptor-binding affinity of fluorotamoxifen, a possible estrogenreceptor imaging agent. J. Med. Chem. 1985, 28, 1504−1511. (32) Zhou, H.-B.; Liu, G.-S.; Yao, Z.-J. Short and efficient total synthesis of luotonin a and 22-hydroxyacuminatine using a common cascade strategy. J. Org. Chem. 2007, 72, 6270−6272. (33) Munoz, L.; Rodriguez, A. M.; Rosell, G.; Bosch, M. P.; Guerrero, A. Enzymatic enantiomeric resolution of phenylethylamines structurally related to amphetamine. Org. Biomol. Chem. 2011, 9, 8171−8177. (34) Hirsh, A. J.; Molino, B. F.; Zhang, J.; Astakhova, N.; Geiss, W. B.; Sargent, B. J.; Swenson, B. D.; Usyatinsky, A.; Wyle, M. J.; Boucher, R. C.; Smith, R. T.; Zamurs, A.; Johnson, M. R. Design, synthesis, and structure−activity relationships of novel 2-substituted pyrazinoylguanidine epithelial sodium channel blockers: Drugs for cystic fibrosis and chronic bronchitis. J. Med. Chem. 2006, 49, 4098− 4115.

5221


Discovery of Cytochrome P450 4F11 Activated Inhibitors of Stearoyl

Recommend Documents