Supplementary Note 1: Synthetic Chemistry Procedures

Transcription

1 Supplementary Note 1: Synthetic Chemistry Procedures Structure-inspired design of β-arrestin-biased ligands for aminergic GPCRs John D. McCorvy 1 *, Kyle V. Butler 2 *, Brendan Kelly 3*, Katie Rechsteiner 1, Joel Karpiak 4, Robin M. Betz 3, Bethany L. Kormos 5, Brian K. Shoichet 4, Ron O. Dror 3, Jian Jin 2, Bryan L. Roth 1 1 National Institute of Mental Health Psychoactive Drug Screening Program, Department of Pharmacology and Division of Chemical Biology and Medicinal Chemistry, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina 27599, USA. 2 Center for Chemical Biology and Drug Discovery, Departments of Pharmacological Sciences and Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA. 3 Departments of Computer Science and Molecular and Cellular Physiology, Institute for Computational and Mathematical Engineering, and Biophysics Program, Stanford University, Stanford, California, 94305, USA. 4 Department of Pharmaceutical Chemistry, University of California at San Francisco, Byers Hall, th Street, San Francisco, California, 94158, USA. 5 Neuroscience and Pain Medicinal Chemistry, Pfizer Worldwide R&D, 610 Main Street, Cambridge, Massachusetts, 02139, USA * These authors contributed equally Correspondence and requests for materials should be addressed to: B.L.R. (bryan_roth@med.unc.edu), R.O.D. (ron.dror@stanford.edu ) J.J. (jian.jin@mssm.edu).

2 Synthesis Scheme for indole-aripiprazole hybrids

3 General procedure for alkylation of indole. 7-(4-(4-(1H-Indol-4-yl)piperazin-1- yl)butoxy)-3,4-dihydroquinolin-2(1h)-one (1) (39 mg, mmol), was dissolved in DMF (1 ml), followed by addition of NaH (60 wt% in mineral oil, 27 mg, 1.11 mmol). The mixture was stirred for 30 min at room temperature, followed by addition of the alkyl halide (0.094 mmol). This was stirred for 2 hours, quenched with water, and purified by HPLC. tert-butyl 4-(piperazin-1-yl)-1H-indole-1-carboxylate (8) A sealed tube was filled with dioxane (25 ml), tert-butyl 4-bromo-1H-indole-1-carboxylate (3.00 g, 10.1 mmol), piperazine (1.747 g, 20.3 mmol), cesium carbonate (6.613 g, 20.3 mmol), palladium acetate (45 mg, 0.20 mmol), and BINAP (318 mg, 0.51 mmol). The tube was filled with argon, sealed, and heated at 100 o C for 18h. The reaction mixture was diluted with EtOAc, filtered through celite, and washed with water. Purification by MPLC (0 to 20% MeOH in DCM, 1% NH 3 ) gave the product. 1 H NMR (Methanol-d 4 ): δ 7.91 (d, 1H), 7.62 (d, 1H), 7.27 (t, 1H), 6.86 (d, 1H), 6.72 (d, 1H), 3.48 (m, 4H), 3.40 (m, 4H), 1.69 (s, 9H). MS (ESI) m/z [M + H] + Calcd for [C 17 H 23 N 3 O 2 + H] + : 302; Found: 302. Yield: g, 4.52 mmol, 45%. 7-(4-(4-(1H-Indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)-one (1) tert- Butyl 4-(piperazin-1-yl)-1H-indole-1-carboxylate (8) (1.30 g, 4.31 mmol), 7-(4- bromobutoxy)-3,4-dihydroquinolin-2(1h)-one (1.672 g, 5.61 mmol), and potassium carbonate (1.191 g, 8.62 mmol) were stirred in dimethylformamide (DMF) (12 ml) overnight at 70 o C. Purification by MPLC (0 to 20% MeOH in DCM, 1% NH 3 ) gave tertbutyl 4-(4-(4-((2-oxo-1,2,3,4-tetrahydroquinolin-7-yl)oxy)butyl)piperazin-1-yl)-1H-indole- 1-carboxylate (1.690 g, MS (ESI) m/z: 519). This intermediate was stirred for 2h in DCM (25 ml) and trifluoroacetic acid (TFA) (25 ml). The reaction mixture was concentrated to an oil and then made basic by addition of saturated aqueous sodium bicarbonate, and the product was extracted with ethyl acetate. The product was purified by MPLC (0 to 10% MeOH in dichloromethane). 1 H NMR (400 MHz, Methanol-d 4 ) δ 7.21 (d, J = 3.2 Hz, 1H), 7.14 (d, J = 8.2, 1H), (m, 2H), 6.58 (m, 2H), (m, 2H), 4.05 (t, J = 5.8 Hz, 2H), 3.83 (d, J = 13.4 Hz, 2H), 3.72 (d, J = 12.1 Hz, 2H), 3.40 (t, J = 12.0 Hz,

4 2H), (m, 3H), 3.10 (t, J = 12.7 Hz, 2H), (m, 2H), (m, 2H), 2.01 (m, 2H), 1.91 (q, J = 6.8, 6.3 Hz, 2H). 13 C NMR (101 MHz, Methanol-d 4 ) δ , , , , , , , , , , , , , , , 98.87, 66.70, 56.47, 52.29, 30.48, 25.98, 23.98, MS (ESI) m/z: [M + H] + Calcd for [C 25 H 30 N 4 O 2 + H] + : 419; Found: 419. Yield: 1.23 g, 2.30 mmol, 53%. HRMS (ESI-TOF) m/z: [M + H] + Calcd for [C 25 H 31 N 4 O 2 + H] + : ; Found: HPLC Purity: Method A, >95%, t R = min. 7-(4-(4-(1-Methyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)- one, trifluoroacetic acid salt (2) Synthesized according to general procedure for alkylation of indole. Methyl iodide was the alkyl halide source. 1 H NMR (400 MHz, Chloroform-d) δ 8.43 (s, 1H), (m, 2H), 7.06 (d, J = 3.0 Hz, 1H), 7.05 (d, J = 2.3 Hz, 1H), 6.70 (d, J = 7.3, 1H), 6.53 (dd, J = 8.3, 2.4 Hz, 1H), 6.39 (d, J = 3.2, 1H), 6.34 (d, J = 2.4 Hz, 1H), 3.97 (d, J = 5.6 Hz, 2H), 3.79 (s, 7H), 3.48 (d, J = 9.0 Hz, 2H), (m, 4H), (m, 2H), 2.63 (dd, J = 8.5, 6.6 Hz, 2H), (m, 2H), (m, 2H). 13 C NMR (101 MHz, Chloroform-d) δ , , , , , , , , , , , , , 98.30, 66.94, 57.01, 52.05, 48.53, 33.11, 30.75, 26.25, 24.30, MS (ESI) m/z: [M + H] + Calcd for [C 26 H 32 N 4 O 2 + H] + : 433; Found: 433. Yield: 13 mg, mmol, 25%. HRMS (ESI-TOF) m/z: [M + H] + Calcd for [C 26 H 33 N 4 O 2 + H] + : ; Found: HPLC Purity: Method A, >95%, t R = min. 7-(4-(4-(1-Propyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)- one, trifluoroacetic acid salt (3) Synthesized according to general procedure for alkylation of indole. 1-Bromopropane was the alkyl halide source. 1 H NMR (400 MHz, Chloroform-d) δ 8.48 (s, 1H), (m, 2H), 7.09 (dd, J = 3.2, 0.7 Hz, 1H), (m, 1H), (m, 1H), 6.51 (dt, J = 8.3, 1.5 Hz, 1H), 6.38 (d, J = 3.2 Hz, 1H), 6.35 (d, J = 2.4 Hz, 1H), 4.07 (t, J = 7.0 Hz, 2H), (m, 2H), (m, 4H), (m, 2H), (m, 4H), 2.88 (t, J = 7.5 Hz, 2H), 2.61 (t, J = 7.5 Hz, 2H), 2.02 (q, J = 7.7 Hz, 2H), 1.86 (p, J = 7.0 Hz, 4H), 0.92 (t, J = 7.4, 3H). 13 C NMR (101 MHz, Chloroform-d) δ , , , , , , ,

5 121.60, , , , , , , 98.24, 66.96, 56.94, 52.12, 48.47, 48.31, 30.88, 26.28, 24.40, 23.50, 20.87, MS (ESI) m/z: [M + H] + Calcd for [C 28 H 36 N 4 O 2 + H] + : 461; Found: 461.Yield: 30 mg, mmol, 56%. HRMS (ESI- TOF) m/z: [M + H] + Calcd for [C 28 H 37 N 4 O 2 + H] + : ; Found: HPLC Purity: Method A, >95%, t R = min. 7-(4-(4-(1-Isopropyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)- one, trifluoroacetic acid salt (4) Synthesized according to general procedure for alkylation of indole. 2-Bromopropane was the alkyl halide source. 1 H NMR (600 MHz, Methanol-d 4 ) δ 7.35 (s, 1H), 7.21 (d, J = 8.1 Hz, 1H), (m, 2H), 6.64 (d, J = 7.2 Hz, 1H), 6.60 (d, J = 8.0 Hz, 1H), 6.50 (d, J = 13.1 Hz, 2H), 4.74 (s, 1H), 4.07 (t, J = 5.5 Hz, 2H), 3.84 (d, J = 13.4 Hz, 2H), 3.74 (d, J = 11.9 Hz, 2H), 3.42 (s, 2H), 3.36 (d, J = 11.1 Hz, 2H), 3.12 (t, J = 12.9 Hz, 2H), 2.90 (d, J = 7.6 Hz, 2H), 2.56 (d, J = 7.6 Hz, 2H), 2.03 (s, 2H), 1.92 (s, 2H), 1.52 (d, J = 6.5 Hz, 6H). 13 C NMR (151 MHz, Methanold 4 ) δ , , , , , , , , , , , , , , 98.59, 66.68, 56.44, 52.23, 49.53, 48.40, 30.44, 25.93, 23.92, 21.52, MS (ESI) m/z: [M + H] + Calcd for [C 28 H 36 N 4 O 2 + H] + : 461; Found: 461. HRMS (ESI-TOF) m/z: [M + H] + Calcd for C 28 H 37 N 4 O 2 + H: ; Found: Yield: 19 mg, mmol, 37%. HPLC Purity: Method A, >95%, t R = min. 7-(4-(4-(1-Benzyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)- one, trifluoroacetic acid salt (5) Synthesized according to general procedure for alkylation of indole. Bromobenzene was the alkyl halide source. 1 H NMR (400 MHz, Chloroform-d) δ (m, 1H), 7.38 (d, J = 8.6 Hz, 1H), (m, 2H), (m, 4H), (m, 1H), 6.79 (d, J = 2.4 Hz, 1H), 6.66 (d, J = 7.3, 1H), 6.44 (d, J = 3.2, 1H), 6.26 (d, J = 9.5 Hz, 1H), 5.31 (s, 2H), 4.07 (t, J = 5.7 Hz, 2H), (m, 4H), (m, 2H), (m, 4H), 2.64 (s, 4H), 2.07 (dd, J = 10.2, 5.9 Hz, 2H), 1.94 (q, J = 7.1, 6.5 Hz, 2H). 13 C NMR (101 MHz, Chloroform-d) δ 13 C NMR (101 MHz, Chloroform-d) δ , , , , , , , , , , , , , , , , ,

6 103.85, 98.90, 64.61, 56.48, 52.25, 49.53, 48.41, 30.27, 25.80, 22.44, MS (ESI) m/z: [M + H] + Calcd for [C 32 H 36 N 4 O 2 + H] + : 509; Found: 509. Yield: 32 mg, mmol, 56%. HRMS (ESI-TOF) m/z: [M + H] + Calcd for [C 32 H 37 N 4 O 2 + H] + : ; Found: HPLC Purity: Method A, >95%, t R = min. 7-(4-(4-(2-methyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)- one (6) 2-Methyl-4-(piperazin-1-yl)-1H-indole (11) (HCl salt, 50 mg, 0.2 mmol) and 7-(4- bromobutoxy)-3,4-dihydroquinolin-2(1h)-one (89 mg, 0.2 mmol) and potassium carbonate (55 mg, 0.4 mmol) were stirred in 1.5 ml DMF for 24 hours. The product was purified by HPLC, giving the product as the TFA salt (36 mg, 33%). HRMS (ESI-TOF) [M + H] + calcd for [C 26 H 32 N 4 O 2 + H] +, ; found, H NMR (600 MHz, DMSO-d 6 ) δ (s, 1H), (s, 1H), 9.50 (s, 1H), 7.07 (d, J = 8.3 Hz, 1H), 6.91 (t, J = 7.8 Hz, 1H), 6.52 (dd, J = 8.3, 2.5 Hz, 1H), 6.45 (m, 2H), 6.14 (s, 1H), 3.97 (t, J = 5.9 Hz, 2H), 3.70 (d, J = 13.1 Hz, 4H), 3.63 (d, J = 12.1 Hz, 4H), 2.97 (t, J = 12.4 Hz, 2H), 2.79 (t, J = 7.6 Hz, 2H), 2.42 (t, J = 7.6 Hz, 2H), 2.37 (s, 3H), 1.85 (m, 2H), 1.78 (d, J = 8.2 Hz, 2H). 13 C NMR (151 MHz, DMSO-d 6 ) δ , , , , , , , , , , , , , , 98.07, 67.01, 55.66, 51.86, 48.11, 31.17, 26.23, 24.40, 20.81, HPLC Purity: Method B, >95%, t R = 3.80 min. tert-butyl 4-(2-methyl-1H-indol-4-yl)piperazine-1-carboxylate (10) 4-Bromo-2-methyl-1H-indole (210 mg, 1 mmol), 1-Boc-piperazine (279 mg, 1.5 mmol), tbubrettphos (10 mg, 0.02 mmol), and tbubrettphos Precatalyst G1 (16 mg, 0.02 mmol) were dissolved in a sealed tube in 3 ml THF. This was treated with 1.2 ml of 1M LiHMDS in THF, and the tube was purged with argon and sealed. The tube was heated at 65 o C for 4 hours. Column chromatography gave the product (155 mg, 49% yield). HRMS (ESI-TOF) [M + H] + calcd for [C 18 H 25 N 3 O 2 + H] +, ; found, H NMR (Chloroform-d): δ 7.93 (s, 1H), 7.06 (t, J = 7.7 Hz, 1H), 7.02 (d, J = 8.0 Hz, 1H), 6.57 (d, J = 7.4 Hz, 1H), 6.23 (s, 1H), 3.68 (t, J = 4.9 Hz, 4H), 3.18 (s, 4H), 2.47 (s, 3H), 1.52 (s, 9H)

7 2-Methyl-4-(piperazin-1-yl)-1H-indole (11) tert-butyl 4-(2-methyl-1H-indol-4-yl)piperazine-1-carboxylate (10) (140 mg, 0.44 mmol) was stirred in 4 ml of 4N HCl in dioxane for two hours. The solid product was collected by filtration and dried under vacuum. Yield: 91 mg of HCl salt, 82%. HRMS (ESI-TOF) [M + H] + calcd for [C 13 H 17 N 3 + H, ] + ; found, C NMR (151 MHz, DMSO-d 6 ) δ , , , , , , 98.03, 66.77, 48.19, 43.31, H NMR (600 MHz, DMSO-d 6 ) δ (s, 1H), 9.31 (s, 2H), 7.03 (d, J = 8.1 Hz, 1H), 6.93 (t, J = 7.8 Hz, 1H), 6.58 (s, 1H), 6.24 (s, 1H), 3.35 (d, 8H), 2.37 (s, 3H). 7-(4-(4-(1,2-Dimethyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin- 2(1H)-one (7) 1,2-Dimethyl-4-(piperazin-1-yl)-1H-indole, trifluoroacetic acid salt (9) was converted to 7-(4-(4-(1,2-dimethyl-1H-indol-4-yl)piperazin-1-yl)butoxy)-3,4-dihydroquinolin-2(1H)-one according to the procedure described for 1. Yield: 41 mg (36%) of the TFA salt. HRMS (ESI-TOF) calcd for [C 27 H 34 N 4 O 2 + H] +, ; found, H NMR (600 MHz, Methanol-d 4 ) δ (m, 3H), (m, 2H), 6.49 (s, 1H), 4.07 (t, J = 5.8 Hz, 2H), 3.82 (d, J = 13.5 Hz, 2H), 3.73 (d, J = 12.0 Hz, 2H), 3.69 (s, 3H), 3.40 (t, J = 11.7 Hz, 2H), 3.36 (m, integration obscured by solvent), 3.09 (t, J = 12.4 Hz, 2H), (m, 2H), (m, 2H), 2.45 (d, J = 5.2 Hz, 3H), 2.03 (s, 2H), 1.92 (t, J = 7.0 Hz, 2H). 13 C NMR (151 MHz, Methanol-d4) δ , , , , , , , , , , , , , , 96.72, 66.68, 56.42, 52.23, 48.27, 30.45, 28.33, 25.94, 23.94, 20.82, HPLC Purity: Method B, >95%, t R = 1.69 min.