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Nov 18, 2015 - Pharmacological property optimization for allosteric ligands: A medicinal chemistry perspective. Shawn Johnstone , Jeffrey S. Albert. B...
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Systematic Study of the Glutathione (GSH) Reactivity of N‑Arylacrylamides: 1. Effects of Aryl Substitution Victor J. Cee,* Laurie P. Volak, Yuping Chen, Michael D. Bartberger,* Chris Tegley, Tara Arvedson, John McCarter, Andrew S. Tasker, and Christopher Fotsch Departments of Medicinal Chemistry, Pharmacokinetics and Drug Metabolism, Molecular Structure, Discovery Technologies, and Oncology Research, Amgen, Inc., One Amgen Center Drive, Thousand Oaks, California 91320, United States S Supporting Information *

ABSTRACT: Success in the design of targeted covalent inhibitors depends in part on a knowledge of the factors influencing electrophile reactivity. In an effort to further develop an understanding of structure−reactivity relationships among N-arylacrylamides, we determined glutathione (GSH) reaction rates for a family of N-arylacrylamides independently substituted at ortho-, meta-, and para-positions with 11 different groups common to inhibitor design. We find that substituent effects on reaction rates show a linear Hammett correlation for ortho-, meta-, and para-substitution. In addition, we note a correlation between 1H and 13C NMR chemical shifts of the acrylamide with GSH reaction rates, suggesting that NMR chemical shifts may be a convenient surrogate measure of relative acrylamide reactivity. Density functional theory calculations reveal a correlation between computed activation parameters and experimentally determined reaction rates, validating the use of such methodology for the screening of synthetic candidates in a prospective fashion.

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INTRODUCTION Recent FDA approvals of covalent inhibitors targeting noncatalytic cysteine residues of kinases BTK (ibrutinib) and EGFR (afatinib) for the treatment of mantle cell lymphoma/chronic lymphocytic leukemia and non-small-cell lung cancer, respectively, have fueled a renewed interest in targeted covalent inhibitors of cysteine-containing proteins.1 To fully achieve the potential of targeted covalent inhibitors, strategies to maximize target protein engagement and minimize off-target reactions are necessary. Key to such strategies will be an understanding of the factors influencing electrophile reactivity. Although the acrylamide functional group has become a widely used electrophile in cysteine-targeted covalent inhibitor design,2 the relative reactivity of structurally diverse acrylamides had not been described until very recently.3 N-Arylacrylamides are featured in a number of targeted covalent inhibitors undergoing clinical testing including AZD9291,3b rociletinib (CO-1686),4 and spebrutinib (AVL-292/CC-292).5 In an effort to further develop an understanding of structure−reactivity relationships among Narylacrylamides, we determined glutathione (GSH) reaction rates for a family of N-arylacrylamides independently substituted at ortho-, meta-, and para-positions with 11 different groups common to inhibitor design.

Figure 1. N-Arylacrylamides. Indicated substituents are independently o (ortho), m (meta), and p (para).

ortho-, meta-, and para-positions (2o−12o, 2m−12m, and 2p−12p, respectively). The acrylamides were either procured from commercial sources or synthesized from the corresponding anilines and acryloyl chloride. NMR spectra (1H and 13C) were obtained for all acrylamides, and for determining correlations between GSH reactivity and chemical shift, we found it convenient to focus on the chemical shifts of the proton defined as Hb1 (the most upfield doublet of doublets in the acrylamide system) and the carbon defined as Cb (easily identified in a distortionless enhancement by polarization transfer6 (DEPT) experiment as the only carbon bound to two hydrogen atoms). Experimental GSH Reaction Rates and Correlations. For determination of rates of reaction with GSH, the acrylamide reactant (1 μM) was incubated at 37 °C with 5 mM GSH in 67

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RESULTS AND DISCUSSION The acrylamides chosen for study are shown in Figure 1 and include the parent N-phenylacrylamide (1) as well as Narylacrylamides containing a wide-range of substituents at © XXXX American Chemical Society

Received: June 29, 2015

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Table 1. Half-Lives and Reaction Rates for Benchmark Electron-Rich and Electron-Deficient N-Arylacrylamides

a

compd

R

Na

GSH t1/2 (min)

kpseudo1st (min−1 × 10−3)

log kGSH (M−1 s−1)b

1 11p 12p

H para-NMe2 para-NO2

4 2 3

179 ± 18 247 ± 18 7.90 ± 0.20

3.86 ± 0.43 2.80 ± 0.21 87.7 ± 2.3

−1.89 ± 0.05 −2.03 ± 0.03 −0.534 ± 0.01

Number of independent determinations. bkGSH is defined as in ref 3b: kpseudo1st/[GSH], with [GSH] = 5 mM.

Table 2. Reaction Rates, Half-Lives, and NMR Chemical Shifts for N-Arylacrylamidesa compd 1 2o 3o 4o 5o 6o 7o 8o 9o 10o 11o 12o 2m 3m 4m 5m 6m 7m 8m 9m 10m 11m 12m 2p 3p 4p 5p 6p 7p 8p 9p 10p 11p 12p

R H F Cl Me CF3 Ph CO2Me CN OMe SMe NMe2 NO2 F Cl Me CF3 Ph CO2Me CN OMe SMe NMe2 NO2 F Cl Me CF3 Ph CO2Me CN OMe SMe NMe2 NO2

Hammettb σm or σp−

GSH t1/2 (min)

kpseudo1st (min−1 × 10−3)

d

−0.03 0.19 −0.17 0.65 0.02 0.64 1.00 −0.26 0.06 −0.12 1.27 0.337 0.373 −0.069 0.43 0.06 0.36 0.56 0.115 0.15 −0.15 0.71 −0.03 0.19 −0.17 0.65 0.02 0.64 1.00 −0.26 0.06 −0.12 1.27

d

3.86 6.29 14.4 2.23 10.9 3.11 23.2 26.9 3.58e 9.55 8.38e 115e 9.68 10.2 3.25 11.2 6.74 5.99 16.7 6.00 7.94 4.56f 27.2e 4.36 7.49 2.62 17.5 5.90 29.7 40.1 2.03 5.09 2.80e 87.7f

179 110 48.3 311 63.5 223 29.9 25.8 194e 72.6 82.7e 6.00e 71.6 68.1 213 61.8 103 116 41.4 116 87.3 152f 25.5e 159 92.5 265 39.5 117 23.3 17.3 342 136 247e 7.90f

log kGSH (M−1 s−1)c

Hb1 δ (ppm)

Cb δ (ppm)

−1.89 −1.68 −1.32 −2.13 −1.44 −1.98 −1.11 −1.05 −1.92e −1.50 −1.55e −0.416e −1.49 −1.47 −1.97 −1.43 −1.65 −1.70 −1.25 −1.70 −1.58 −1.82f −1.04e −1.84 −1.60 −2.06 −1.23 −1.71 −1.00 −0.874 −2.17 −1.77 −2.03e −0.534f

5.75 5.79 5.78 5.74 5.78 5.65 5.83 5.84 5.71 5.74 5.72 5.83 5.79 5.79 5.73 5.81 5.78 5.79 5.82 5.75 5.76 5.71 5.84 5.75 5.77 5.72 5.82 5.77 5.81 5.83 5.71 5.74 5.67 5.86

126.8 127.5 127.4 126.4 127.2 126.4 127.4 128.1 126.5 126.7 126.4 128.2 127.4 127.5 126.6 127.6 126.9 127.3 127.9 126.9 127 126.4 128 126.9 127.2 126.5 127.8 126.9 127.8 128.2 126.2 126.7 125.6 128.7

d

Data represent single determinations unless otherwise indicated. bFor ortho- and para-, σp− is listed; for meta, σm is listed; values were obtained from ref 6. ckGSH is defined as in ref 3b: kpseudo1st/[GSH], with [GSH] = 5 mM. dAverage of four determinations. eAverage of two determinations. f Average of three determinations. For statistical information, see the Supporting Information. a

compounds, and three controls were routinely included in each pool. Compounds were added as 10 mM DMSO solutions, and the final DMSO content varied from 1.0% to 1.5% depending on the number of compounds pooled. From the mass spectrum, peak area ratios (peak area analyte/peak area of a stable internal standard) were calculated and the percent compound remaining was determined relative to time zero. Rates were calculated by fitting to a pseudo-first-order kinetic equation to natural log transformed percent remaining data (eq 1). Half-lives were calculated according to eq 2.

mM potassium phosphate buffer, pH 7.4 (1.0−1.5% DMSO), and the samples were analyzed with an LC/MS system that consisted of a Leap autosampler and an AB Sciex TripleTOF 5600 mass spectrometer. The concentration of GSH was chosen to approximate intracellular concentrations of GSH (reported7 range 0.5−10 mM). Samples were directly injected from the incubation immediately after GSH was added to the mixture (less than 2 min after the acrylamide reactants were dosed into buffer) and then at predetermined intervals over 120 min. The analytes were separated with a Waters Xselect HSS T3 2.5 μm (2.1 mm × 30 mm) column, and the mobile phase consisted of 0.1% formic acid in water/acetonitrile with a linear gradient of organic phase increasing from 5% to 95% over the 1 min run time. To increase throughput, compounds were tested in pools of 10−15 B

ln([acrylamide]) = −k pseudo1stt + ln([acrylamide0])

(1)

t1/2 = 0.693/k pseudo1st

(2) DOI: 10.1021/acs.jmedchem.5b01018 J. Med. Chem. XXXX, XXX, XXX−XXX

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Figure 2. Rate of GSH addition is correlated with Hammett sigma values. (a) Ortho-substituted 2o−12o, plot of log kGSH (M−1 s−1) vs σp−. (b) Metasubstituted 2m−12m, plot of log kGSH (M−1 s−1) vs σm. (c) Para-substitued 2p−12p, plot of log kGSH (M−1 s−1) vs σp−.

Figure 3. GSH reaction rate increases with increasing chemical shift. (a) Acrylamide proton (Hb1) chemical shift (400 MHz, DMSO-d6). (b) Acrylamide carbon (Cb) chemical shift (100 MHz, DMSO-d6). Red, green, and purple coloration correspond to ortho-, meta-, and para-substitution patterns, respectively. Parent denotes N-phenylacrylamide 1.

meta-position (GSH t1/2 range of 25.5−213 min). The impact of the individual substituents on GSH reactivity was evaluated by fitting the data to the Hammett equation (eq 3) and determining the set of Hammett σ values (also known as substituent constants) that provided the best linear fit to the data (Figure 2). From the best linear fit to the data, the reaction constant ρ, a measure of the sensitivity of the reaction to substituent effects relative to the substituent constants, is obtained from the slope, while the intercept provides the constant c, which is equivalent to the log of the rate of the unsubstituted reference compound.

The maximum half-life that could be determined with a 120 min incubation was set at 512 min based on the ability to confidently detect 15% parent depletion, taking all uncertainty in the measurement into consideration. A control experiment in which GSH was not present was simultaneously conducted to confirm that the observed disappearance of acrylamide reactant was GSH-dependent. All compounds reported herein showed only GSH-dependent disappearance. The reproducibility and stability of the assay were confirmed by multiple determinations on different days for GSH reaction with N-phenylacrylamide (1) as well as the electron-rich N-(4-(dimethylamino)phenyl)acrylamide (11p) and electron-poor N-(4-nitrophenyl)acrylamide (12p) (Table 1). Low variability was observed across different determinations, as evidenced by the low standard deviations. An additional confirmatory experiment was performed in which the rate of formation of the GSH-adduct was measured for N-phenylacrylamide (1). The rate of GSH-adduct formation was observed to closely match the rate of Nphenylacrylamide disappearance (Figure SI-1). The mean GSH half-life for N-phenylacrylamide (1, 179 min) under these conditions is between that reported by Flanagan et al.3a (53 min, 1 mM electrophile, 10 mM GSH, 100 mM, pH 7.4 phosphate buffer, 10% acetonitrile) and Ward et al.3b (299 min, 50 μM electrophile, 4.6 mM GSH, pH 7.4 phosphate buffer; phosphate concentration not indicated). We suspect that differences in experimental parameters such as concentration, cosolvent, and ionic strength are responsible for differences in the measured value for N-phenylacrylamide and stress that the relative trends observed under a given set of conditions will have the most value. The full set of N-arylacrylamides (Figure 1) was then evaluated, and the data are presented in Table 2. The aryl-ring substituents are observed to have the largest impact on GSH t1/2 at the ortho- and para-positions (GSH t1/2 range of 6.00−311 and 7.90−342 min, respectively) and a less dramatic impact at the

log k = ρσ + c

(3)

We find that the best fit is obtained by using σp− for both ortho (Figure 2a, R2 = 0.80) and para (Figure 2c, R2 = 0.98) substituents and σm for meta-substituents (Figure 2b, R2 = 0.83). The derived values for ρ are near unity (ortho, 0.85; meta, 0.89; para, 1.0), and the intercept c (ortho, −1.7 M−1 s−1; meta, −1.7 M−1 s−1; para, −1.8 M−1 s−1) is nearly identical to log kGSH of the unsubstituted N-phenylacrylamide (compound 1, Table 2, log kGSH − 1.89 M−1 s−1). The ρ values presented here are lower than that found in a study published recently by Ward et al.3b (5.56), and we believe the discrepancy may be due to the smaller data set used in the latter analysis (one meta- and five para-substituents were studied), as well as the use of a single substituent constant for all substituents, regardless of position. The observation that σp− (derived from ionization of phenols) provides an improved correlation relative to σp (derived from ionization of benzoic acids) indicates that through-resonance effects are significant in the addition of GSH to ortho- and paraN-arylacrylamides; through resonance is generally not considered for meta-substituents.8 It is also notable that orthosubstituted N-phenylacrylamides are well-modeled by a simple Hammett correlation, and consideration of steric and proximity polar effects unique to ortho-substituents as advanced by the C

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Fujita−Nishioka equations9 appears to be unnecessary in this case. The reason for this may be that in the case of Narylacrylamides, the reacting center is separated from the aromatic system by three atoms and is therefore less affected by steric and proximity polar effects. We observe a reasonable correlation between proton and carbon chemical shift (Hb1 and Cb, respectively, as defined in Figure 1) and GSH reaction rate values (Figure 3) for this group of closely related N-arylacrylamides, with R2 values of 0.71 and 0.80, respectively. Examination of the ortho-, meta-, and parasubsets indicates that the correlation between chemical shift and reaction rate is somewhat better for the meta- and para-subsets than the ortho-subset, an outcome that was also observed in the correlation to Hammett σ values (Figure 2).10 The finding that downfield chemical shifts are associated with increased reactivity suggests that the relative electron deficiency of the acrylamide is responsible for both effects.11 While the correlation observed among the structurally similar acrylamide electrophiles reported in this study does not appear to hold as well as substrate diversity increases (i.e., substituted and/or non-arylacrylamides),3a our results suggest that NMR chemical shifts may still provide utility as a rapid screening metric within a homologous series of substrates. To more readily visualize these data, we present a heat map of substituent effects on GSH reaction rates relative to Nphenylacrylamide (Table 3). Values are the fold-change in Table 3. Heatmap of Fold-Change in kGSH Relative to NPhenylacrylamide (1)a

Figure 4. Transition structures and relevant geometric parameters for methanethiolate addition to N-phenylacrylamide 1 at the IEFPCMB3LYP/6-311+G(d,p) level of theory: (a) top and side views of TS-syn1; (b) top and side views of TS-syn-2; (c) single view of TS-syn-3; (d) top and side views of TS-anti-1; (e) single view of TS-anti-2. See Table 4 for comparison of relative energies and geometric parameters determined by the IEFPCM versus SMD solvation models. Interatomic distances are in ångstroms, and angles are in degrees.

for a syn-type orientation of the methanethiolate nucleophile is in accord with high-level CBS-QB3 calculations by Krenske and Houk on model enone−methanethiolate systems.16 A prior study reporting a theoretical analysis of methanethiolate addition to acrylamides at the same level of theory appeared to utilize an anti-type transition structure to the s-cis conformer (Figure 4d, TS-anti-1 (s-cis)).3a Activation parameters (determined in the typical fashion, utilizing the lowest-energy conformation of the TS compared to the lowest-energy s-cis conformer of the reactant) were calculated for the family of acrylamides in Table 2. In all cases, a TS-syn-1like TS was found to be most favored. These results are reported in Table 5. The computed activation parameters Ea and ΔG⧧, determined in this fashion, exhibit a reasonably good correlation (R2 = 0.83 and 0.75, respectively) with experimental reaction rates (Figure 5). Similar methodology has been successfully applied to the reaction of nitriles with cysteine, suggesting that the determination of activation parameters with density functional theory will have broad utility in the prediction of reaction rates of sulfur nucleophiles with covalent warheads.17 Inspection of the individual data points for ortho-, meta-, and para-substitution indicates a somewhat tighter correlation for the last two substitution patterns, as was found for both Hammett constants and chemical shift values (Figures 2 and 3, respectively).18

a

Values are the fold-change in reaction rate relative to N-phenylacrylamide. Color scale is shown as a gradient with green = 0.5 (minimum) and red = 26 (maximum).

reaction rate relative to N-phenylacrylamide (1), with values of 1 indicating a faster rate of reaction with GSH. For polysubstituted N-arylacrylamides, we expect the substituent effects presented in Table 3 to be additive provided that the substituents are able to adopt conformations similar to the simple monosubstituted N-arylacrylamides.12 Theoretical Analysis: Transition Structures and Activation Parameters. The addition of methanethiolate anion to N-phenylacrylamide (1) was studied at the B3LYP/6-311+G(d,p) level of theory with the inclusion of solvent effects via the IEFPCM-SMD13 method, and a number of transition structures for methanethiolate addition to both s-cis and s-trans Nphenylacrylamide and aryl-substituted analogs have been located (Figure 4 and Table 4).14,15 A syn-like orientation of the nucleophile with respect to the acrylamide CC bond is preferred (Figure 4a, TS-syn-1 (s-cis)). The general preference D

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Table 4. Comparison of IEFPCM and SMD-IEFPCM Energetic and Geometric Data for Methanethiolate Addition Transition Structures of 1a TS

r(CC) (Å)

r(C−S) (Å)

∠CCS (deg)

φ(C−C−S−C) (deg)

Erel (kcal/mol)b

ΔGrel (kcal/mol)b

1.420 (1.394) 1.422 e 1.430 (1.400) 1.402 (1.388) 1.411 (1.392)

2.176 (2.376) 2.171 e 2.144 (2.347) 2.291 (2.429) 2.248 (2.410)

115.9 (114.0) 116.2 e 116.4 (113.8) 116.3 (117.7) 115.9 (115.8)

−20.9 (−16.6) 23.3 e −33.1 (−22.8) 177.0 (175.1) 168.1 (160.1)

0 (0) 0.2 e 0.4 (0.6) 1.4 (1.8) 1.9 (2.4)

0 (0) 0.7 e 1.5 (0.4) 0.2 (0.7) 1.9 (0.8)

TS-syn-1c,d TS-syn-2c,d TS-syn-3c,d TS-anti-1c,d,f TS-anti-2c,d a

Data for fully optimized transition structures independently optimized at the B3LYP/6-311+G(d,p) level of theory using both implicit solvation models. All transition structures characterized by Hessian and internal reaction coordinate (IRC) analysis. bTemperature and frequency scale factors of 310.15 K and 0.9877, respectively, were utilized for zero-point energy and entropic contributions in order to attempt consistency with ref 3a. c Data for IEFPCM solvation model in top row (see Figure 4). dData for IEFPCM-SMD solvent model parenthesized in italics in bottom row. eTSsyn-2 converges to TS-syn-1 on the IEFPCM-SMD-B3LYP/6-311+G(d,p) potential energy surface. fTransition structure reported for methanethiolate addition to α,β-unsubstituted acrylamides in ref 3a.

Table 5. Calculated Activation Energies (Ea) and Free Energies (ΔG⧧) in Implicit Water Solvent (B3LYP/6-311+G(d,p)a compd 1 2o 3o 4o 5o 6o 7o 8o 9o 10o 11o 12o 2m 3m 4m 5m 6m

R H F Cl Me CF3 Ph CO2Me CN OMe SMe NMe2 NO2 F Cl Me CF3 Ph

Ea b (kcal/mol) 8.9 (4.4) 7.6 7.3 8.9 6.7 8.8 7.6 5.7 9.4 8.4 9.9 4.4 7.5 7.6 9.0 7.2 8.7

c

ΔG⧧ b (kcal/mol) 19.2 (15.5) 18.2 18.0 20.3 17.7 19.8 18.8 17.3 20.2 19.4 21.3 15.4 18.3 18.5 19.7 17.8 19.8

c

compd

R

Ea b (kcal/mol)

ΔG⧧ b (kcal/mol)

7m 8m 9m 10m 11m 12m 2p 3p 4p 5p 6p 7p 8p 9p 10p 11p 12p

CO2Me CN OMe SMe NMe2 NO2 F Cl Me CF3 Ph CO2Me CN OMe SMe NMe2 NO2

8.0 6.9 8.6 8.4 9.6 6.7 8.7 8.1 9.7 6.8 8.5 6.6 6.0 10.0 8.4 11.1 4.5

19.1 17.5 19.5 19.3 20.7 17.6 19.8 19.1 20.7 18.0 19.7 17.9 17.1 20.8 19.0 22.4 16.0

a

Data for fully optimized transition structures independently optimized at the IEFPCM-B3LYP/6-311+G(d,p) level of theory. IEFPCM utilized due to favorable geometric convergence behavior. bTemperature and frequency scale factors of 310.15 K and 0.9877, respectively, were utilized for zeropoint energy and entropic contributions in order to attempt consistency with ref 3a. cData for IEFPCM-SMD evaluation of 1 parenthesized in italics.

Figure 5. Graph of log kGSH versus computed (a) activation energy (Ea) and (b) activation free energy (ΔG⧧) for systems in Table 5 at the IEFPCMB3LYP/6-311+G(d,p) level of theory. Red, green, and purple coloration correspond to ortho-, meta-, and para-substitution patterns, respectively. Parent denotes N-phenylacrylamide 1.

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CONCLUSIONS In an effort to further develop an understanding of structure− reactivity relationships among N-arylacrylamides, we determined glutathione (GSH) reaction rates for a family of N-

arylacrylamides independently substituted at ortho-, meta-, and para-positions with 11 different groups common to inhibitor design. We find that substituent effects on reaction rates show a linear Hammett correlation for ortho-, meta-, and paraE

DOI: 10.1021/acs.jmedchem.5b01018 J. Med. Chem. XXXX, XXX, XXX−XXX

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substitution. In addition, we note a correlation between 1H and 13 C NMR chemical shifts of the acrylamide with GSH reaction rates, suggesting that NMR chemical shifts may indeed be a convenient surrogate measure of relative acrylamide reactivity. Our density functional theory calculations characterize the nature of the transition structures as involving a preferential synlike orientation of the methanethiolate nucleophile upon addition to the α,β-unsubstituted N-arylacrylamides utilized in this study. A correlation between computed activation parameters and experimentally determined reaction rates was observed, validating the use of such methodology for the screening of synthetic candidates in a prospective fashion. We believe that this comprehensive study elucidates the key structure−reactivity relationships for N-arylacrylamides and will allow for more informed strategies for targeted covalent inhibitor design in three ways: (1) the experimental reaction rates presented here may be used to select N-aryl substituents that will provide a desired level of reactivity; (2) proton and carbon NMR chemical shifts may be used to quickly estimate the reactivity of compounds that have already been prepared; (3) DFT-computed activation energies for methanethiolate addition, still quite modest in terms of CPU demand (on the order of a few hours per system on a typical workstation), can be used to prospectively estimate the reactivity of designed compounds containing N-aryl substituents that have not been addressed by this study. Additional studies involving α- and/or β-substituted acrylamides will be reported in due course.

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127.5, 115.1, 115.0, 110.0, 109.8, 106.2, 106.0. m/z (ESI, +ve ion) 166.1 (M + H)+. N-(3-Fluorophenyl)acrylamide (2m). 1H NMR (400 MHz, DMSO-d6) δ 10.33 (br s, 1H), 7.68 (d, J = 11.74 Hz, 1H), 7.32−7.39 (m, 2H), 6.86−6.93 (m, 1H), 6.42 (dd, J = 9.98, 17.02 Hz, 1H), 6.28 (dd, J = 2.05, 16.92 Hz, 1H), 5.79 (dd, J = 1.96, 9.98 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.4, 163.3, 160.9, 140.7, 140.6, 131.5, 130.4, 130.3, 127.4, 115.1, 115.0, 110.0, 109.8, 106.2, 106.0. m/z (ESI, +ve ion) 166.1 (M + H)+. N-(4-Fluorophenyl)acrylamide (2p). 1H NMR (400 MHz, DMSO-d6) δ 10.17 (br s, 1H), 7.65−7.72 (m, 2H), 7.16 (t, J = 8.41 Hz, 2H), 6.41 (dd, J = 10.07, 16.92 Hz, 1H), 6.26 (dd, J = 2.05, 16.92 Hz, 1H), 5.75 (dd, J = 2.15, 9.98 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.0, 159.3, 156.9, 135.4, 135.4, 131.7, 126.9, 121.1, 121.0, 115.4, 115.2. m/z (ESI, +ve ion) 166.1 (M + H)+. N-(2-Chlorophenyl)acrylamide (3o). 1H NMR (400 MHz, DMSO-d6) δ 9.71 (br s, 1H), 7.78 (d, J = 7.62 Hz, 1H), 7.51 (dd, J = 1.37, 8.02 Hz, 1H), 7.34 (dt, J = 1.57, 7.73 Hz, 1H), 7.21 (dt, J = 1.76, 7.73 Hz, 1H), 6.62 (dd, J = 10.17, 17.02 Hz, 1H), 6.28 (dd, J = 1.96, 17.02 Hz, 1H), 5.78 (dd, J = 1.96, 10.37 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.4, 134.7, 131.3, 129.4, 127.3, 126.6, 126.4, 126.3. m/z (ESI, +ve ion) 182 (M + H)+. N-(3-Chlorophenyl)acrylamide (3m). 1H NMR (400 MHz, DMSO-d6) δ 10.31 (s, 1H), 7.90 (t, J = 1.96 Hz, 1H), 7.51 (d, J = 7.86 Hz, 1H), 7.35 (t, J = 8.12 Hz, 1H), 7.13 (ddd, J = 0.88, 2.10, 7.97 Hz, 1H), 6.42 (dd, J = 9.98, 17.02 Hz, 1H), 6.28 (dd, J = 1.96, 17.02 Hz, 1H), 5.79 (dd, J = 2.05, 10.07 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.4, 140.4, 133.0, 131.5, 130.4, 127.5, 123.1, 118.8, 117.7. m/z (ESI, +ve ion) 182.1 (M + H)+. N-(4-Chlorophenyl)acrylamide (3p). 1H NMR (400 MHz, DMSO-d6) δ 10.26 (s, 1H), 7.64−7.75 (m, 2H), 7.31−7.45 (m, 2H), 6.42 (dd, J = 10.17, 17.02 Hz, 1H), 6.27 (dd, J = 2.05, 16.92 Hz, 1H), 5.77 (dd, J = 2.05, 10.07 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 137.9, 131.6, 128.6, 127.2, 127.0, 120.8. m/z (ESI, +ve ion) 182.1 (M + H)+. N-(2-Methylphenyl)acrylamide (4o). 1H NMR (400 MHz, DMSO-d6) δ 9.46 (br s, 1H), 7.47 (d, J = 7.82 Hz, 1H), 7.07−7.24 (m, 3H), 6.54 (dd, J = 10.27, 17.12 Hz, 1H), 6.24 (dd, J = 1.96, 17.02 Hz, 1H), 5.74 (dd, J = 2.15, 10.17 Hz, 1H), 2.21 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 136.0, 131.8, 131.6, 130.3, 126.4, 125.9, 125.2, 124.9, 124.8, 17.9. m/z (ESI, +ve ion) 162.1 (M + H)+. N-(3-Methylphenyl)acrylamide (4m). 1H NMR (500 MHz, DMSO-d6) δ ppm 10.03 (br s, 1 H), 7.50 (s, 1 H), 7.45 (d, J = 8.0 Hz, 1 H), 7.19 (t, J = 7.9 Hz, 1 H), 6.88 (d, J = 7.4 Hz, 1 H), 6.43 (dd, J = 17.0, 10.2 Hz, 1 H), 6.24 (dd, J = 17.0, 1.9 Hz, 1 H), 5.73 (dd, J = 10.1, 1.9 Hz, 1 H), 2.28 (s, 3 H). 13C NMR (101 MHz, DMSO-d6) δ 163.0, 138.9, 137.9, 131.9, 128.6, 126.6, 124.1, 119.8, 116.5, 21.2. m/z (ESI, +ve ion) 162.1 (M + H)+. N-(4-Methylphenyl)acrylamide (4p). 1H NMR (500 MHz, DMSO-d6) δ ppm 10.02 (br s, 1 H), 7.55 (m, J = 8.3 Hz, 2 H), 7.12 (m, J = 8.2 Hz, 2 H), 6.42 (dd, J = 17.0, 10.2 Hz, 1 H), 6.23 (dd, J = 17.0, 1.9 Hz, 1 H), 5.72 (dd, J = 10.1, 1.9 Hz, 1 H), 2.25 (s, 3 H). 13C NMR (101 MHz, DMSO-d6) δ 162.9, 136.5, 132.4, 131.9, 129.1, 126.5, 119.3, 20.4. m/z (ESI, +ve ion) 162.1 (M + H)+. N-(2-Trifluoromethylphenyl)acrylamide (5o). 1H NMR (400 MHz, DMSO-d6) δ 9.76 (s, 1H), 7.75 (t, J = 7.70 Hz, 1H), 7.69 (d, J = 7.71 Hz, 1H), 7.54 (d, J = 7.71 Hz, 1H), 7.47 (t, J = 7.70 Hz, 1H), 6.53 (dd, J = 10.17, 17.02 Hz, 1H), 6.25 (dd, J = 1.96, 17.02 Hz, 1H), 5.78 (dd, J = 1.96, 10.17 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 164.2, 135.1, 132.9, 131.0, 130.1, 127.2, 126.8, 126.3, 126.3, 124.9, 124.7, 122.2. m/z (ESI, +ve ion) 216.1 (M + H)+. N-(3-Trifluoromethylphenyl)acrylamide (5m). 1H NMR (400 MHz, DMSO-d6) δ 10.46 (s, 1H), 8.17 (s, 1H), 7.85 (d, J = 8.41 Hz, 1H), 7.57 (t, J = 7.92 Hz, 1H), 7.42 (d, J = 7.82 Hz, 1H), 6.43 (dd, J = 9.98, 17.02 Hz, 1H), 6.27−6.34 (m, 1H), 5.81 (dd, J = 2.05, 10.07 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.6, 139.7, 131.4, 130.0, 129.5 (q, J = 32 Hz), 127.6, 124.0 (q, J = 271 Hz), 122.8, 119.7, 115.3. m/z (ESI, +ve ion) 216.1 (M + H)+. N-(4-Trifluoromethylphenyl)acrylamide (5p). 1H NMR (400 MHz, DMSO-d6) δ 10.47 (s, 1H), 7.85−7.90 (m, J = 8.41 Hz, 2H),

EXPERIMENTAL SECTION

Chemistry. Unless otherwise noted, all materials were obtained from commercial suppliers and used without further purification. Anhydrous solvents were obtained from Aldrich and used directly. All reactions involving air- or moisture-sensitive reagents were performed under a nitrogen or argon atmosphere. Silica gel chromatography was performed using medium pressure liquid chromatography (MPLC) on a CombiFlash Companion (Teledyne Isco) with RediSep normal-phase silica gel (35−60 μm) columns and UV detection at 254 nm. All compounds were purified to ≥95% purity as determined by Agilent 1100 series high performance liquid chromatography (HPLC) with UV detection at 254 nm (Phenomenex Synergi, 2 mm × 50 mm, 3 min, 1.0 mL/min flow rate, 0−95% 0.1% TFA in CH3CN/0.1% TFA in H2O). NMR spectra were determined with a Bruker AV-400 (400 MHz) at ambient temperature. Low-resolution mass spectral (MS) data were determined on an Agilent 1100 series LCMS with UV detection at 254 nm and a low resonance electrospray mode (ESI). Chemical shifts are reported in ppm from the solvent resonance (DMSO-d6, 2.50 ppm). Data are reported as follows: chemical shift, multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, br = broad, m = multiplet), coupling constants, and number of protons. General Procedure for Synthesis of Noncommercial Acrylamides. To a solution of aniline (1 mmol) in 4 mL DCM at 0 °C was added N,N-diisopropylethylamine (2 mmol) and acryloyl chloride (1 mmol). Upon completion, the reaction was concentrated and was purified by silica gel chromatography. The product-containing fractions were combined and concentrated to give the desired acrylamides. N-Phenylacrylamide (1). 1H NMR (400 MHz, DMSO-d6) δ 10.12 (br s, 1H), 7.67 (d, J = 7.82 Hz, 2H), 7.33 (t, J = 7.92 Hz, 2H), 7.07 (t, J = 7.16 Hz, 1H), 6.45 (dd, J = 9.98, 17.02 Hz, 1H), 6.23−6.30 (m, 1H), 5.76 (dd, J = 1.86, 10.07 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.1, 139.0, 131.9, 128.7, 126.8, 123.5, 119.3. m/z (ESI, +ve ion) 148.1 (M + H)+. N-(2-Fluorophenyl)acrylamide (2o). 1H NMR (400 MHz, DMSO-d6) δ 10.33 (br s, 1H), 7.68 (d, J = 11.76 Hz, 1H), 7.32−7.39 (m, 2H), 6.86−6.94 (m, 1H), 6.42 (dd, J = 9.98, 17.02 Hz, 1H), 6.25− 6.32 (m, 1H), 5.79 (dd, J = 1.96, 9.98 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.4, 163.3, 160.9, 140.7, 140.6, 131.5, 130.4, 130.3, F

DOI: 10.1021/acs.jmedchem.5b01018 J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

J = 7.24 Hz, 1H), 6.43 (dd, J = 10.07, 16.92 Hz, 1H), 6.25 (dd, J = 1.66, 16.92 Hz, 1H), 5.75 (dd, J = 1.76, 10.17 Hz, 1H), 3.73 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 163.1, 159.5, 140.1, 131.8, 129.5, 126.8, 111.6, 108.9, 105.1, 54.9. m/z (ESI, +ve ion) 178.1 (M + H)+. N-(4-Methoxyphenyl)acrylamide (9p). 1H NMR (400 MHz, DMSO-d6) δ 9.98 (s, 1H), 7.54−7.61 (m, J = 9.00 Hz, 2H), 6.87−6.93 (m, J = 8.80 Hz, 2H), 6.40 (dd, J = 10.17, 17.02 Hz, 1H), 6.22 (dd, J = 1.96, 16.82 Hz, 1H), 5.71 (dd, J = 1.96, 9.98 Hz, 1H), 3.73 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 162.6, 155.3, 132.1, 131.9, 126.2, 120.7, 113.9, 55.1. m/z (ESI, +ve ion) 178.2 (M + H)+. N-(2-(Methylthio)phenyl)acrylamide (10o). 1H NMR (500 MHz, DMSO-d6) δ ppm 9.51 (br s, 1 H), 7.43 (d, J = 7.4 Hz, 1 H), 7.35 (d, J = 7.8 Hz, 1 H), 7.12−7.29 (m, 2 H), 6.40−6.61 (m, 1 H), 6.23 (dd, J = 17.0, 1.8 Hz, 1 H), 5.74 (d, J = 10.1 Hz, 1 H), 2.41 (s, 3 H). 13C NMR (101 MHz, DMSO-d6) δ 163.5, 134.9, 133.4, 131.5, 126.7, 126.3, 126.1, 125.2, 15.1. m/z (ESI, +ve ion) 194.0 (M + H)+. N-(3-(Methylthio)phenyl)acrylamide (10m). 1H NMR (500 MHz, DMSO-d6) δ ppm 10.14 (br s, 1 H), 7.66 (s, 1 H), 7.40 (d, J = 7.5 Hz, 1 H), 7.26 (t, J = 7.9 Hz, 1 H), 6.96 (d, J = 7.2 Hz, 1 H), 6.42 (dd, J = 17.0, 10.1 Hz, 1 H), 6.26 (dd, J = 17.0, 1.8 Hz, 1 H), 5.76 (dd, J = 10.1, 1.8 Hz, 1 H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 139.5, 138.6, 131.7, 129.2, 127.0, 120.8, 116.3, 115.8, 40.4, 14.6. m/z (ESI, +ve ion) 194.0 (M + H)+. N-(4-(Methylthio)phenyl)acrylamide (10p). 1H NMR (400 MHz, DMSO-d6) δ 10.13 (s, 1H), 7.60−7.66 (m, J = 8.61 Hz, 2H), 7.20−7.29 (m, J = 8.61 Hz, 2H), 6.42 (dd, J = 9.98, 17.02 Hz, 1H), 6.21− 6.30 (m, 1H), 5.74 (dd, J = 1.86, 10.07 Hz, 1H), 2.45 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 163.0, 136.4, 132.1, 131.8, 127.0, 126.7, 119.9, 15.4. m/z (ESI, +ve ion) 194.1 (M + H)+. N-(2-(Dimethylamino)phenyl)acrylamide (11o). 1H NMR (400 MHz, DMSO-d6) δ 9.28 (br s, 1H), 7.88−8.01 (m, 1H), 6.99−7.17 (m, 3H), 6.73 (dd, J = 10.17, 17.02 Hz, 1H), 6.24 (dd, J = 1.96, 17.02 Hz, 1H), 5.72 (dd, J = 2.15, 10.17 Hz, 1H), 2.61 (s, 6H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 144.8, 132.2, 131.7, 126.4, 124.5, 122.8, 122.6, 119.1, 43.6. m/z (ESI, +ve ion) 191.1 (M + H)+. N-(3-(Dimethylamino)phenyl)acrylamide (11m). 1H NMR (500 MHz, DMSO-d6) δ ppm 9.92 (br s, 1 H), 7.06−7.13 (m, 2 H), 6.99 (d, J = 7.8 Hz, 1 H), 6.39−6.48 (m, 2 H), 6.23 (dd, J = 17.0, 1.9 Hz, 1 H), 5.71 (dd, J = 10.1, 1.9 Hz, 1 H), 2.87 (s, 6 H). 13C NMR (101 MHz, DMSO-d6) δ 162.9, 150.7, 139.7, 132.1, 129.0, 126.4, 108.0, 107.6, 103.4, 40.1. m/z (ESI, +ve ion) 191.1 (M + H)+. N-(4-(Dimethylamino)phenyl)acrylamide (11p). 1H NMR (400 MHz, DMSO-d6) δ 9.83 (s, 1H), 7.45−7.52 (m, J = 9.00 Hz, 2H), 6.66− 6.74 (m, J = 9.19 Hz, 2H), 6.39 (dd, J = 10.07, 16.92 Hz, 1H), 6.19 (dd, J = 2.05, 16.92 Hz, 1H), 5.67 (dd, J = 2.05, 10.07 Hz, 1H), 2.85 (s, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 162.3, 147.1, 132.1, 128.8, 125.6, 120.6, 112.6, 40.4. m/z (ESI, +ve ion) 191.1 (M + H)+. N-(2-Nitrophenyl)acrylamide (12o). 1H NMR (400 MHz, DMSO-d6) δ 10.47 (br s, 1H), 7.96 (d, J = 7.80 Hz, 1H), 7.65−7.74 (m, 2H), 7.39 (ddd, J = 1.76, 6.90, 8.36 Hz, 1H), 6.46 (dd, J = 10.17, 17.02 Hz, 1H), 6.24−6.31 (m, 1H), 5.83 (dd, J = 1.96, 10.17 Hz, 1H). 13 C NMR (101 MHz, DMSO-d6) δ 163.3, 142.7, 133.9, 130.8, 128.2, 125.4, 125.4, 124.9. m/z (ESI, +ve ion) 193.1 (M + H)+. N-(3-Nitrophenyl)acrylamide (12m). 1H NMR (400 MHz, DMSO-d6) δ 10.61 (s, 1H), 8.70 (t, J = 2.15 Hz, 1H), 7.98 (d, J = 7.80 Hz, 1H), 7.92 (d, J = 7.79 Hz, 1H), 7.62 (t, J = 8.22 Hz, 1H), 6.44 (dd, J = 9.98, 17.02 Hz, 1H), 6.33 (dd, J = 1.96, 17.02 Hz, 1H), 5.84 (dd, J = 2.05, 9.88 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.7, 147.9, 140.1, 131.2, 130.2, 128.0, 125.2, 117.9, 113.4. m/z (ESI, +ve ion) 193.1 (M + H)+. N-(4-Nitrophenyl)acrylamide (12p). 1H NMR (400 MHz, DMSO-d6) δ 10.73 (s, 1H), 8.21−8.28 (m, J = 9.19 Hz, 2H), 7.90− 7.95 (m, J = 9.19 Hz, 2H), 6.48 (dd, J = 9.98, 17.02 Hz, 1H), 6.35 (dd, J = 1.66, 16.92 Hz, 1H), 5.85−5.90 (m, 1H). 13C NMR (101 MHz, DMSOd6) δ 163.8, 145.2, 142.4, 131.2, 128.5, 125.0, 119.1. m/z (ESI, +ve ion) 193.1 (M + H)+.

7.66−7.71 (m, J = 8.61 Hz, 2H), 6.46 (dd, J = 9.98, 17.02 Hz, 1H), 6.28− 6.35 (m, 1H), 5.82 (dd, J = 1.86, 10.07 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.6, 142.5, 131.4, 127.8, 126.0, 125.7, 123.6, 123.3, 123.0, 119.2. m/z (ESI, +ve ion) 216.1 (M + H)+. N-([1,1′-Biphenyl]-2-yl)acrylamide (6o). 1H NMR (500 MHz, DMSO-d6) δ ppm 9.44 (br s, 1 H), 7.53 (d, J = 7.4 Hz, 1 H), 7.29−7.45 (m, 7 H), 6.23−6.39 (m, 1 H), 6.15 (d, J = 15.6 Hz, 1 H), 5.65 (d, J = 10.1 Hz, 1 H). 13C NMR (101 MHz, DMSO-d6) δ 163.8, 138.8, 136.6, 134.4, 131.6, 130.3, 128.7, 128.3, 127.6, 127.2, 126.4, 126.1. m/z (ESI, +ve ion) 224.0 (M + H)+. N-([1,1′-Biphenyl]-3-yl)acrylamide (6m). 1H NMR (500 MHz, DMSO-d6) δ ppm 10.23 (br s, 1 H), 8.00 (s, 1 H), 7.58−7.68 (m, 3 H), 7.33−7.51 (m, 5 H), 6.47 (dd, J = 17.0, 10.2 Hz, 1 H), 6.29 (dd, J = 16.9, 1.8 Hz, 1 H), 5.78 (dd, J = 10.2, 1.8 Hz, 1 H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 140.8, 140.1, 139.5, 131.8, 129.3, 128.9, 127.5, 126.9, 126.6, 121.9, 118.3, 117.6. m/z (ESI, +ve ion) 224.0 (M + H)+. N-([1,1′-Biphenyl]-4-yl)acrylamide (6p). 1H NMR (400 MHz, DMSO-d6) δ 10.23 (s, 1H), 7.77 (d, J = 7.98 Hz, 2H), 7.62−7.68 (m, 4H), 7.44 (t, J = 7.16 Hz, 2H), 7.27−7.38 (m, 1H), 6.46 (dd, J = 10.07, 16.92 Hz, 1H), 6.28 (dd, J = 1.96, 17.02 Hz, 1H), 5.75−5.80 (m, 1H). 13 C NMR (101 MHz, DMSO-d6) δ 163.1, 139.6, 138.4, 135.1, 131.8, 128.8, 127.0, 126.9, 126.9, 126.2, 119.7. m/z (ESI, +ve ion) 224.4 (M + H)+. Methyl 2-Acrylamidobenzoate (7o). 1H NMR (400 MHz, DMSO-d6) δ 10.77 (s, 1H), 8.30 (dd, J = 1.37, 8.41 Hz, 1H), 7.92 (dd, J = 1.66, 7.92 Hz, 1H), 7.62 (t, J = 7.81 Hz, 1H), 7.22 (t, J = 7.49 Hz, 1H), 6.44 (dd, J = 10.17, 17.02 Hz, 1H), 6.27 (dd, J = 1.56, 17.02 Hz, 1H), 5.83 (dd, J = 1.76, 10.17 Hz, 1H), 3.85 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 167.5, 163.3, 139.2, 139.2, 133.8, 132.1, 130.5, 127.4, 123.5, 121.4, 118.2, 52.4. m/z (ESI, +ve ion) 206.1 (M + H)+. Methyl 3-Acrylamidobenzoate (7m). 1H NMR (400 MHz, DMSO-d6) δ 10.36 (s, 1H), 8.34 (s, 1H), 7.93 (dd, J = 1.08, 8.12 Hz, 1H), 7.66 (d, J = 7.39 Hz, 1H), 7.47 (t, J = 7.92 Hz, 1H), 6.43 (dd, J = 9.98, 17.02 Hz, 1H), 6.29 (dd, J = 2.05, 16.92 Hz, 1H), 5.79 (dd, J = 1.96, 9.98 Hz, 1H), 3.86 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 166.0, 163.3, 139.3, 131.6, 130.1, 129.2, 127.3, 124.0, 123.7, 119.8, 52.2. m/z (ESI, +ve ion) 206.1 (M + H)+. Methyl 4-Acrylamidobenzoate (7p). 1H NMR (400 MHz, DMSO-d6) δ 10.46 (s, 1H), 7.91−7.96 (m, J = 8.61 Hz, 2H), 7.78− 7.83 (m, J = 8.80 Hz, 2H), 6.46 (dd, J = 10.17, 17.02 Hz, 1H), 6.30 (dd, J = 1.76, 17.02 Hz, 1H), 5.81 (dd, J = 1.86, 10.07 Hz, 1H), 3.83 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 165.7, 163.5, 143.3, 131.5, 130.2, 127.8, 124.1, 118.7, 51.9. m/z (ESI, +ve ion) 206.1 (M + H)+. N-(2-Cyanophenyl)acrylamide (8o). 1H NMR (400 MHz, DMSO-d6) δ 10.36 (s, 1H), 7.83 (dd, J = 1.66, 7.73 Hz, 1H), 7.62− 7.74 (m, 2H), 7.37 (t, J = 7.28 Hz, 1H), 6.52 (dd, J = 10.17, 17.21 Hz, 1H), 6.32 (dd, J = 1.86, 17.12 Hz, 1H), 5.84 (dd, J = 1.76, 10.17 Hz, 1H). 13 C NMR (101 MHz, DMSO-d6) δ 163.5, 139.9, 133.8, 133.2, 130.8, 128.1, 125.9, 125.6, 116.8, 107.3. m/z (ESI, +ve ion) 173.1 (M + H)+. N-(3-Cyanophenyl)acrylamide (8m). 1H NMR (400 MHz, DMSO-d6) δ 10.47 (s, 1H), 8.17 (s, 1H), 7.86 (t, J = 3.93 Hz, 1H), 7.54 (s, 1H), 7.55 (d, J = 5.94 Hz, 2H), 6.43 (dd, J = 9.88, 16.92 Hz, 1H), 6.28−6.34 (m, 1H), 5.82 (dd, J = 1.96, 9.98 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 163.6, 139.7, 131.3, 130.3, 127.9, 127.0, 123.9, 122.0, 118.6, 111.6. m/z (ESI, +ve ion) 173.1 (M + H)+. N-(4-Cyanophenyl)acrylamide (8p). 1H NMR (400 MHz, DMSO-d6) δ 10.55 (s, 1H), 7.76−7.87 (m, 4H), 6.45 (dd, J = 10.07, 16.92 Hz, 1H), 6.28−6.36 (m, 1H), 5.83 (dd, J = 1.86, 10.07 Hz, 1H). 13 C NMR (101 MHz, DMSO-d6) δ 163.7, 143.2, 133.2, 131.3, 128.2, 119.4, 119.0, 105.2. m/z (ESI, +ve ion) 173.1 (M + H)+. N-(2-Methoxyphenyl)acrylamide (9o). 1H NMR (400 MHz, DMSO-d6) δ 9.34 (br s, 1H), 8.03 (d, J = 8.02 Hz, 1H), 7.03−7.12 (m, 2H), 6.92 (t, J = 7.61 Hz, 1H), 6.70 (dd, J = 10.17, 17.02 Hz, 1H), 6.23 (dd, J = 2.15, 17.02 Hz, 1H), 5.71 (dd, J = 2.15, 10.17 Hz, 1H), 3.84 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 163.2, 149.7, 149.7, 132.1, 127.0, 126.5, 124.6, 122.1, 120.2, 111.1, 55.6. m/z (ESI, +ve ion) 178.1 (M + H)+. N-(3-Methoxyphenyl)acrylamide (9m). 1H NMR (400 MHz, DMSO-d6) δ 10.10 (br s, 1H), 7.38 (s, 1H), 7.17−7.25 (m, 2H), 6.65 (d, G

DOI: 10.1021/acs.jmedchem.5b01018 J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

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Article

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jmedchem.5b01018. Figure SI-1; GSH assay protocol; statistical analysis of experimental GSH data for acrylamides 1, 9o, 11o, 12o, 11m, 12m, 11p, 12p (multiple determinations); NMR spectra of acrylamides 1−12; coordinates of optimized transition structures for acrylamides 1−12 (PDF)

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AUTHOR INFORMATION

Corresponding Authors

*V.J.C.: e-mail, [email protected]; phone, 805-313-5500. *M.D.B.: e-mail, [email protected]; phone, 805-447-3446. Author Contributions

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

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS Useful discussions with Dr. Hua Gao are acknowledged with thanks. ABBREVIATIONS USED B3LYP, Becke three-parameter Lee−Yang−Parr hybrid density functional; BTK, Bruton’s tyrosine kinase; DEPT, distortionless enhancement by polarization transfer; DMSO, dimethylsulfoxide; EGFR, epidermal growth factor receptor; GSH, glutathione; IEFPCM, integral-equation-formalism polarizable continuum model; SMD, solvation model density; TS, transition structure

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REFERENCES

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DOI: 10.1021/acs.jmedchem.5b01018 J. Med. Chem. XXXX, XXX, XXX−XXX