4.1.2. General Procedure for the Ugi Reaction (Method A)

The ketone (1.32 mmol) was dissolved in methanol (2 mL) and the amine, amino acid and isocyanide were successively added (2 equivalents of each) to this solution. After 4 days at room temperature with stirring, 1.2 M hydrochloric acid in methanol was added, and the mixture was further stirred for an additional 30 minutes at room temperature. After removal of the solvents, the residue was treated with ethyl acetate, washed with saturated sodium bicarbonate (3 × 10 mL), then washed with brine (3 × 10 mL). The organic phase was filtered after drying with magnesium sulphate and finally evaporated to dryness. Flash column chromatography (hexane:ethyl acetate, 4:1 to 0:1) was used to purify the final residue to yield the 1,4,4-trisubstituted piperidine analogues 15, 16, 49–53 and 57–59. Details are given below for each compound.

Methyl(S)-4-(benzyl(1-benzyl-4-tert-butylcarbamoylpiperidin-4-yl)amino)-3-tert-butoxycarbonylamino-4-oxobutanoate (15): Reaction of 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-Asp(OMe)-OH (0.44 mmol, 122 mg) and tert-butyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) yielded 15 (98 mg, 73%) as a white foam. ¹H NMR (500 MHz, DMSO-d6) δ 7.51 (d, J = 7.6 Hz, 2H), 7.36 (t, J = 7.6 Hz, 3H), 7.25 (m, 3H), 7.18 (m, 3H), 6.17 (s, 1H), 4.79 (d, J = 18.1 Hz, 1H), 4.69 (q, J = 7.0 Hz, 1H), 4.62 (d, J = 18.1 Hz, 1H), 3.56 (s, 3H), 3.43 (m, 1H), 2.84 (dd, J = 16.4, 6.8 Hz, 1H), 2.50 (m, 1H), 2.41 (m, 4H), 2.26–2.12 (m, 2H), 1.98 (d, J = 10.9 Hz, 1H), 1.81 (m, 1H), 1.58 (m, 1H), 1.30 (s, 9H), 1.16 (s, 9H). ¹³C NMR (126 MHz, DMSO-d6) δ 171.9, 171.6, 171.4, 155.5, 139.6, 138.9, 129.1, 128.6, 128.5, 127.6, 127.2, 78.9, 78.8, 64.7, 62.3, 55.3, 51.9, 50.3, 49.6, 47.4, 36.5, 31.0, 28.6, 28.5. MS (ES+) m/z: 609.49 (M + 1)⁺. HPLC retention time = 8.31 min (96% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₃₄H₄₈N₄O₆ (C, H, N): C, 67.08; H, 7.95; N, 9.20. Found: C, 67.32; H, 7.84; N, 9.53.

Methyl(S)-4-(benzyl(1-benzyl-4-cyclohexylcarbamoylpiperidin-4-yl)amino)-3-tert-butoxycarbonylamino-4-oxobutanoate (16): Following the general procedure, a solution of 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-Asp(OMe)OH (0.44 mmol, 122 mg) and cyclohexyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) was reacted. Compound 16 (81 mg, 58%) was obtained as a white foam after purification. ¹H NMR (500 MHz, DMSO-d6) δ 7.46 (d, J = 7.6 Hz, 2H), 7.36 (t, J = 7.6 Hz, 2H), 7.27 (m, 1H), 7.23 (dd, J = 8.0, 6.5 Hz, 3H), 7.17 (m, 1H), 7.13 (m, 2H), 6.77 (d, J = 8.0 Hz, 1H), 4.76 (d, J = 18.4 Hz, 1H), 4.68 (m, 2H), 3.55 (s, 3H), 3.42 (m, 1H), 3.26 (d, J = 5.6 Hz, 2H), 3.16 (d, J = 5.1 Hz, 1H), 2.84 (dd, J = 16.5, 5.6 Hz, 1H), 2.45 (m, 2H), 2.39 (m, 2H), 2.31 (m, 1H), 2.15 (m, 2H), 1.78 (td, J = 12.3, 4.1 Hz, 1H), 1.62 (m, J = 9.2, 4.2 Hz, 4H), 1.52 (m, 2H), 1.29 (s, 9H), 1.19 (m, 2H), 1.09 (m, 2H). ¹³C NMR (126 MHz, DMSO-d6) δ 171.9, 171.4, 171.4, 155.5, 139.2, 138.9, 129.2, 129.1, 128.9, 128.5, 127.5, 127.3, 127.2, 78.8, 64.1, 62.2, 51.9, 50.3, 49.9, 49.5, 47.9, 47.8, 36.6, 32.8, 32.6, 32.1, 28.5, 25.8, 25.2. MS (ES+) m/z 635.59 (M + 1)⁺. HPLC retention time = 8.53 min (99% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₃₆H₅₀N₄O₆ (C, H, N): C, 68.11; H, 7.94; N, 8.83. Found: C, 68.33; H, 7.76; N, 8.64.

Tert-butyl(S)-(1-(benzyl(1-benzyl-4-benzylcarbamoylpiperidin-4-yl)amino)-3-(4-fluorophenyl)-1-oxopropan-2-yl)carbamate (49): Following the general procedure, a solution of 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-4-fluorophenylalanine (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) was reacted. After purification of the final residue, compound 49 (109 mg, 73%) was obtained as a white foam. ¹H NMR (500 MHz, DMSO-d6) δ 7.84 (t, J = 6.0 Hz, 1H), 7.35 (m, 17H), 7.02 (m, 2H), 6.98 (m, 1H), 5.84 (m, 2H), 4.28 (m, 3H), 3.30 (m, 2H), 2.89 (dd, J = 14.0, 4.2 Hz, 1H), 2.68 (dd, J = 14.0, 9.8 Hz, 1H), 2.52 (m, 1H) 2.42 (m, 4H), 2.26 (m, 1H), 1.80 (m, 1H), 1.55 (m, 1H), 1.21 (s, 9H). ¹³C NMR (126 MHz, DMSO-d6) δ 173.6, 172.8, 161.4 (d, J_C-F = 241.7 Hz), 155.8, 140.4, 139.5, 138.8, 134.6 (d, J_C-F = 2.9 Hz), 131.5 (d, J_C-F = 8.0 Hz), 129.3, 129.2, 129.07, 128.9, 128.8, 128.5, 128.5, 127.6, 127.5, 127.45, 127.3, 127.2, 126.9, 126.9, 126.6, 115.1 (d, J_C-F = 21.0 Hz), 78.5, 63.9, 62.2, 55.5, 55.4, 54.5, 50.1, 49.8, 48.1, 42.8, 38.3, 37.0, 32.7, 32.2, 28.4, 27.7. MS (ES+) m/z: 679.5 (M + 1)⁺. HPLC retention time = 6.98 min (98% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₁H₄₇FN₄O₄ (C, H, F, N): C, 72.54; H, 6.98; F, 2.80; N, 8.25. Found: C, 72.26; H, 6.90; F, 2.91; N, 8.7.

Tert-butyl(1-(benzyl(1-benzyl-4-benzylcarbamoylpiperidin-4-yl)amino)-3-(4-chlorophenyl)-1-oxopropan-2-yl)carbamate (50): According to the general procedure, a solution of 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-4-chlorophenylalanine (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) was reacted. Compound 50 (142 mg, 93%) was obtained as a white foam. ¹H NMR (500 MHz, DMSO-d6) δ 7.85 (t, J = 6.0 Hz, 1H), 7.23 (m, 16H), 7.13 (d, J = 8.0 Hz, 2H), 7.02 (d, J = 8.0 Hz, 2H), 4.88 (d, J = 18.7 Hz, 1H), 4.79 (d, J = 18.7 Hz, 1H), 4.31 (m,1H), 4.26 (m,1H), 3.28 (s, 2H), 2.90 (dd, J = 13.9, 4.2 Hz, 1H), 2.67 (dd, J = 13.9, 9.8 Hz, 1H), 2.53 (m, 1H), 2.40 (m, 4H), 2.37 (m, 1H), 2.25 (t, J = 11.9 Hz, 1H), 1.79 (m, 1H), 1.53 (m, 1H), 1.20 (s, 9H). ¹³C NMR (126 MHz, DMSO-d6) δ 173.5, 172.9, 172.8, 172.6, 155.7, 140.4, 139.5, 138.8, 137.5, 131.7, 131.5, 131.4, 129.3, 129.2, 129.1, 128.9, 128.8, 128.5, 128.5, 128.4, 127.7, 127.5, 127.4, 127.2, 126.9, 126.9, 126.6, 78.6, 78.5, 63.9, 63.9, 62.2, 54.3, 50.1, 49.7, 48.1, 42.8, 37.2, 32.7, 32.2, 28.4. MS (ES+) m/z: 695.46 (M)⁺, 697.45 (M+2)⁺. HPLC retention time = 9.45 min (95% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₁H₄₇ClN₄O₄ (C, H, Cl, N): C, 70.83; H, 6.81; Cl, 5.10; N, 8.06. Found: C, 70.55; H, 6.93; Cl, 5.24; N, 8.34.

Tert-butyl(1-(benzyl(1-benzyl-4-benzylcarbamoylpiperidin-4-yl)amino)-3-(4-methoxyphenyl)-1-oxopropan-2-yl)carbamate (51): Reactants: 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-4-methoxylphenylalanine (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL). 51 was obtained (140 mg, 92%) as a white foam. ¹H NMR (500 MHz, DMSO-d6) δ 7.81 (t, J = 6.0 Hz, 1H), 7.25 (m, 16H), 6.93 (d, J = 8.4 Hz, 2H),6.73 (d, J = 8.4 Hz, 2H), 4.91 (d, J = 18.5 Hz, 1H), 4.78 (d, J = 18.5 Hz, 1H), 4.28 (m, 3H), 3.66 (s, 3H), 3.28 (m, 3H), 2.85 (dd, J = 14.0, 4.1 Hz, 1H), 2.62 (dd, J = 14.0, 9.6 Hz, 1H), 2.52 (m, 1H), 2.40 (m, 3H), 2.26 (t, J = 11.5 Hz, 1H), 1.77 (m, 1H), 1.55 (bt, J = 9.5 Hz, 1H), 1.24 (s, 9H). ¹³C NMR (126 MHz, DMSO-d6) δ 173.9, 172.8, 158.2, 155.8, 140.4, 139.5, 138.9, 130.7, 130.3, 129.3, 129.2, 129.0, 129.0, 128.5, 128.5, 127.7, 127.6, 127.5, 127.4, 127.3, 127.2, 126.9, 126.9, 126.6, 113.9, 78.5, 78.4, 63.9, 62.2, 55.4, 54.8, 50.3, 50.1, 49.8, 48.1, 42.8, 36.9, 32.129, 28.5, 27.8, 19.1. MS (ES+) m/z: 691.57 (M + 1)⁺. HPLC retention time = 8.92 min (95% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₂H₅₀N₄O₅ (C, H, N): C, 73.02; H, 7.29; N, 8.11. Found: C, 73.22; H, 7.05; N, 8.38.

Tert-butyl(1-(benzyl(1-benzyl-4-benzylcarbamoylpiperidin-4-yl)amino)-1-oxo-3-(p-tolyl)propan-2-yl)carbamate (52): Reaction of 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Boc-4-methylphenylalanine (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) gave 52 (119 mg, 80%) as a white foam. ¹H NMR (500 MHz, DMSO-d6) δ 7.82 (t, J = 6.0 Hz, 1H), 7.26 (m, 16H), 6.97 (d, J = 7.6 Hz, 2H), 6.89 (d, J = 7.6 Hz, 2H), 4.92 (d, J = 18.5 Hz, 1H), 4.78 (d, J = 18.5 Hz, 1H), 4.27 (m, 3H), 3.28 (s, 3H), 2.88 (dd, J = 14.0, 4.0 Hz, 1H), 2.64 (dd, J = 14.0, 9.6 Hz, 1H), 2.51 (m, 1H), 2.40 (m, 3H), 2.26 (t, J = 11.4 Hz, 1H), 2.20 (s, 3H), 1.79 (m, 1H), 1.55 (m, 1H), 1.24 (s, 9H). ¹³C NMR (126 MHz, DMSO-d6) δ 173.9, 173.3, 172.8, 172.6, 155.8, 153.8, 140.4, 139.5, 139.1, 138.9, 135.6, 135.6, 135.4, 129.7, 129.5, 129.3, 129.2, 129.0, 128.9, 128.5, 128.5, 127.6, 127.5, 127.4, 127.2, 126.9, 126.9, 126.57, 78.5, 78.4, 63.9, 62.2, 55.5, 54.7, 50.3, 50.1, 49.8, 48.1, 42.8, 38.9, 37.4, 32.9, 32.7, 32.2, 28.5, 27.8, 21.1. MS (ES+) m/z: 675.54 (M + 1)⁺. HPLC retention time = 9.35 min (99% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₂H₅₀N₄O₄ (C, H, N): C, 74.75; H, 7.47; N, 8.30. Found: C, 74.91; H, 7.21; N, 8.63.

Methyl(S)-4-(benzyl(4-benzylcarbamoyl-1-phenethylpiperidin-4-yl)amino)-3-benzyloxycarbonylamino-4-oxobutanoate (53): A solution of 1-phenethylpiperidin-4-one (0.22 mmol, 0.016 mL), benzylamine (0.44 mmol, 0.071 mL), Cbz-Asp(OMe)-OH (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) was reacted. After purification of the final residue, compound 53 (122 mg, 80%) was obtained as a white foam. ¹H NMR (400 MHz, CDCl₃) δ 7.20 (m, 20 H), 7.08 (d, J = 7.4 Hz, 1H), 6.71 (bt, J = 4.8 Hz, 1H), 5.02 (m. 1H), 4.9 (m, 3H), 4.76 (m, 3H), 4.32 (m, 2H), 3.47 (s, 3H), 2.55 (m, 8H), 1.97 (s, 2H), 2.37 (m, 2H). ¹³C NMR (101 MHz, CDCl₃) δ 172.9, 172.5, 171.3, 171.2, 155.3, 138.7, 137.9, 135.9, 129.1, 128.7, 128.6, 128.5, 128.4, 128.3, 128.1, 127.7, 127.6, 127.2, 126.2, 126.1, 67.3, 64.9, 60.441, 59.9, 51.9, 50.3, 49.9, 49.6, 48.1, 43.6, 37.5. MS (ES+) m/z: 691.63 (M + 1)⁺. HPLC retention time = 8.40 min (99% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₁H₄₆N₄O₆ (C, H, N): C, 71.28; H, 6.71; N, 8.11. Found: C, 71.30; H, 6.55; N, 8.34.

Methyl(S)-4-((1-benzyl-4-benzylcarbamoylpiperidin-4-yl)(2-(phenylamino)ethyl)amino)-3-benzyloxycarbonylamino-4-oxobutanoate (57): 1-benzylpiperidin-4-one (0.22 mmol, 0.016 mL), N¹-phenylethane-1,2-diamine (0.44 mmol, 0.071 mL), Cbz-Asp(OMe)-OH (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) reacted to give 57 (64 mg, 41%) as a white foam. ¹H NMR (400 MHz, CDCl₃) δ 7.32 (m, 18), 6.65 (t, J = 7.5 Hz, 1H), 6.52 (d, J = 7.5 Hz, 2H), 5.26 (m, J = 9.1 Hz, 1H), 4.97 (m, 4H), 4.33 – 4.11 (m, 3H), 3.76 (m, 2H), 3.46 (s, 3H), 3.40 (m, 4H), 2.52 (m, 9H). ¹³C NMR (101 MHz, CDCl₃) δ 173.3, 172.8, 171.3, 155.6, 147.4, 138.7, 135.8, 129.7, 129.4, 128.8, 128.6, 128.5, 128.4, 128.4, 128.3, 128.1, 127.7, 127.6, 127.5, 127.2, 127.0, 118.2, 113.3, 67.4, 63.8, 62.4, 60.4, 52.0, 51.8, 50.2, 49.8, 48.9, 44.2, 43.8, 43.5, 37.4, 32.2. MS (ES+) m/z 706.72 (M + 1)⁺. HPLC retention time = 8.61 min (95% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₁H₄₇N₅O₆ (C, H, N): C, 69.77; H, 6.71; N, 9.92. Found: C, 69.81; H, 6.54; N, 9.73.

Methyl(S)-4-((4-benzylcarbamoyl-1-phenethylpiperidin-4-yl)(2-(phenylamino)ethyl)amino)-3-benzyloxycarbonylamino-4-oxobutanoate (58): 1-phenethylpiperidin-4-one (0.22 mmol, 0.016 mL), N¹-phenylethane-1,2-diamine (0.44 mmol, 0.071 mL), Cbz-Asp(OMe)-OH (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) reacted according to the general procedure to give, after purification, a white foam that was identified as compound 58 (32 mg, 20%). ¹H NMR (400 MHz, CDCl₃) δ 7.13 (m, 18H), 6.66 (t, J = 7.5 Hz, 1H), 6.52 (d, J = 7.5 Hz, 2H), 5.29 (m, 1H), 5.23 (s, 1H), 5.02 (d, J = 12.0 Hz, 1H), 4.93 (d, J = 12.0 Hz, 1H), 4.92 (m, 1H), 4.14 (m, 3H), 3.77 (m, 2H), 3.47 (s, 3H), 3.40 (m, 2H), 2.73 (m, 9H), 2.49 (m, 4H). ¹³C NMR (101 MHz, CDCl₃) δ 206.9, 173.2, 172.9, 171.3, 155.6, 147.4, 139.9, 138.7, 135.8, 129.4, 128.7, 128.6, 128.5, 128.5, 128.4, 128.1, 127.8, 127.2, 126.2, 118.3, 113.3, 77.4, 77.2, 77.0, 76.7, 67.4, 63.9, 59.9, 52.0, 50.2, 49.9, 48.9, 44.2, 43.8, 43.5, 37.7, 33.7, 32.7, 30.9. MS (ES+) m/z: 720.80 (M + 1)⁺. HPLC retention time = 8.68 min (97% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flow rate of 1 mL/min). Anal. calculated for C₄₂H₄₉N₅O₆ (C, H, N): C, 70.08; H, 6.86; N, 9.73. Found: C, 70.29; H, 6.51; N, 9.93.

Methyl(S)-4-((4-benzylcarbamoyl-1-phenethylpiperidin-4-yl)(4-methylbenzyl)amino)-3-benzyloxycarbonylamino-4-oxobutanoate (59): Reaction of 1-phenethylpiperidin-4-one (0.22 mmol, 0.016 mL), 4-methylbenzylamine (0.44 mmol, 0.071 mL), Cbz-Asp(OMe)-OH (0.44 mmol, 122 mg) and benzyl isocyanide (0.44 mmol, 0.066 mL) in MeOH (2 mL) gave 59 (112 mg, 72%) as a white foam. ¹H RMN (400 MHz, DMSO-d6) δ 7.22 (m, 19H), 4.99 (dd, J = 16.0, 12.0 Hz, 2H,), 4.83 (d, J = 16.0 Hz, 1H), 4.80 (t, J = 8.1 Hz, 1H), 4.71 (d, J = 16.0 Hz, 1H), 4.28 (d, J = 8.1 Hz, 2H,), 3.58 (s, 3H), 3,09 (s, 2H), 2.85 (dd, 2H), 2.53 (m, 4H), 2.30 (s, 3H), 1.87 (2H, td, J = 12.2, 4.4 Hz, 4H), 1.72 (td, 2H, J = 12.2, 4.4 Hz, 4H). ¹³C RMN (100 MHz, CDCl₃) δ 172.8, 172.5, 171.2, 155.3, 140.2, 138.7, 137.2, 136.0, 134.8, 129.7, 128.7, 128.6, 128.5, 128.4, 128.3, 128.1, 127.7, 127.2, 126.1, 126.0, 67.2, 65.0, 60.0, 51.9, 50.3, 49.9, 49.6, 47.8, 43.6, 37.5, 33.6, 32.8, 32.1, 21.1. MS (ES+) m/z: 705.72 (M + 1)⁺. HPLC retention time = 8.76 min (99% analytical purity) (H₂O/CH₃CN from 15/85 to 0/100 in 5 min flowrate of 1 mL/min). Anal. calculated for C₄₂H₄₈N₄O₆ (C, H, N): C, 71.57; H, 6.86; N, 7.95. Found: C, 71.42; H, 6.99; N, 7.74.