Czechowicz et al.
143.4, 143.4, 141.0, 127.4, 126.7, 124.61, 124.57. 119.6, 77.2, 66.6,
54.5, 46.9, 46.8, 44.1, 42.7, 42.5, 37.5, 35.0, 30.9, 29.3, 25.6, 25.5,
24.1, 23.8, 22.2 ppm; HRMS (ESI) m/z calcd for C49H58N7O8 [M
+ H]+ 872.4341, found 872.4317.
in a hydrophobic pocket (the “Z-site”) on the enzyme that is
not utilized in the binding of the natural substrate.9c,23 The more
potent inhibition observed for our Fmoc derivative 4 suggests
that this putative Z-site may favorably accommodate aromatic
groups that are significantly larger than the Cbz moiety. On
the other hand, in addition to the Z-site, the enzyme also
possesses a hydrophobic patch comprising the side chains of
Leu18, Trp22, Tyr111, and Met114, where trypanothione’s
spermidine moiety binds.6 The aromatic acridine ring of a
quinacrine-based inhibitor was recently shown to bind in this
hydrophobic site,24 and this pocket is also the putative binding
site of a number of related hydrophobic tricyclic TR inhibitors.25
It seems unlikely, but it is possible that 3 and 4 are turned around
in the active site, binding in an orientation that places their
aromatic moieties in this hydrophobic site that normally binds
the substrate’s spermidine moiety.
In conclusion, we have developed syntheses of dethiotrypan-
othione and related trypanothione analogues featuring RCM
macrocyclizations. Dethiotrypanothione proved to be a very poor
inhibitor of TR despite its close structural similarity to the
natural substrate. However, the more hydrophobic analogues 3
and 4 are more effective inhibitors, underscoring the enzyme’s
general affinity for hydrophobic ligands.
N1,N8-Bis(benzyloxycarbonylglycyl)spermidine (5): mp 148-
1
149 °C (lit.13 149-150 °C); H NMR (400 MHz, DMSO-d6) δ
7.90 (t, 1H, J ) 5.4 Hz), 7.86 (t, 1H, J ) 5.7 Hz), 7.45-7.30 (m,
11.6H), 7.09-7.02 (m, 0.4H CONH, minor rotamer), 5.03 (s, 4H),
3.57 (d, 4H, J ) 6.0 Hz), 3.09 (dt, 2H, J ) 6.4, 6.4 Hz), 3.05 (dt,
2H, J ) 6.2, 6.2 Hz), 2.48-2.43 (m, 4H), 1.52 (quint, 2H, J ) 6.7
Hz), 1.44-1.33 (m, 4H); 13C NMR (100 MHz, DMSO-d6) δ 168.8,
168.7, 156.4, 137.0, 136.9, 128.2, 127.7, 127.6, 65.4, 48.8, 46.7,
43.5, 38.4, 36.9, 29.2, 26.9, 26.7 ppm; LRMS (ESI) m/z 528.4 [M
+ H]+, 550.3 [M + Na]+.
N1,N8-Bis(benzyloxycarbonylglycyl)-N4-tert-butoxycarbonyl-
spermidine (6): 1H NMR (400 MHz, CDCl3) δ 7.36-7.25 (m,
10.7H), 6.93-6.70 (m, 1.3H), 6.14-5.90 (m, 2H), 5.09 (s, 2H),
5.08 (s, 2H), 3.84-3.81 (m, 4H), 3.26-3.05 (m, 8H), 1.71-1.56
(m, 2H), 1.54-1.39 (m, 4H), 1.42 (s, 9H) ppm; 13C NMR (100
MHz, CDCl3) δ 169.4(C), 156.8(C), 156.1(C), 136.3(C), 136.2-
(C), 128.51(CH), 128.47(CH), 128.2(CH), 128.1(CH), 128.0(CH),
79.7(C), 67.0(CH2), 66.8(CH2), 46.8(CH2), 45.1(CH2), 44.5(CH2),
43.9(CH2), 38.8(CH2), 36.2(CH2), 29.3(CH3, tert-butyl, minor
rotamer), 28.4(CH3), 27.8(CH2), 26.7(CH2), 25.8(CH2) ppm; LRMS
(ESI) m/z 650.3 (M + Na)+.
1
Diene 8: H NMR (400 MHz, CDCl3,) δ 7.73-6.87 (m, 14H),
6.06-5.62 (m, 4H), 5.19-4.98 (m, 8H), 4.40-4.16 (m, 2H), 4.10-
3.66 (m, 4H), 3.36-3.00 (m, 8H), 2.69-2.34 (m, 4H), 1.77-1.60
(m, 2H), 1.59-1.33 (m, 13H, within this multiplet lies the tert-
butyl group CH3 singlet at 1.42 ppm) ppm; 13C NMR (100 MHz,
CDCl3) δ 172.1(C), 169.0(C), 156.5(C), 156.1(C), 136.1(C), 136.0-
(C), 133.0(CH), 132.7(CH), 128.6(CH), 128.5(CH), 128.3(CH),
128.2(CH), 128.0(CH), 119.2(CH2), 119.0(CH2), 79.7(C), 67.13-
(CH2), 67.10(CH2), 54.7(CH), 47.0(CH2), 45.3(CH2), 44.1(CH2),
43.2(CH2), 38.7(CH2), 36.6(CH2), 28.4(CH3), 27.8(CH2), 26.6(CH2),
25.9(CH2); HRMS (ESI) m/z calcd for C42H59N7NaO10 [M + Na]+
844.4227, found 844.4217.
Experimental Section
Dethiotrypanothione (2): 1H NMR (400 MHz, CD3OD) δ 4.26
(dd, 1H, J ) 6.8, 6.8 Hz), 4.25 (dd, 1H, J ) 5.9, 8.5 Hz), 4.04 (dd,
1H, J ) 6.5, 6.5 Hz), 4.03 (dd, 1H, J ) 6.3, 6.3 Hz), 3.88 (d, 1H,
J ) 16.1 Hz), 3.86 (d, 1H, J ) 16.1 Hz), 3.77 (d, 1H, J ) 16.1
Hz), 3.73 (d, 1H, J ) 16.1 Hz), 3.40-3.33 (m, 2H), 3.32-3.24
(m, 2H), 3.03 (t, 2H, J ) 6.8 Hz), 3.00 (t, 2H, J ) 7.2 Hz), 2.60-
2.53 (m, 4H), 2.29-2.10 (m, 4H), 1.93-1.84 (m, 2H), 1.84-1.66
(m, 6H), 1.65-1.57 (m, 2H), 1.51-1.35 (m, 2H) ppm; 13C NMR
(100 MHz, CD3OD) δ 175.2, 175.0, 174.6, 174.5, 172.8, 171.9,
171.6, 55.0, 53.5, 45.7, 44.0, 43.8, 39.0, 36.5, 32.1, 27.3, 27.1, 26.9,
26.1, 25.7, 23.9 ppm; HRMS (ESI) m/z calcd for C29H52N9O10 [M
+ H]+ 686.3832, found 686.3831.
Diene 9: 1H NMR (400 MHz, acetone-d6) δ 7.91-7.83 (m, 2H
CONH), 7.86 (d, 4H, J ) 7.6 Hz), 7.69 (dd, 4H, J ) 3.8, 7.5 Hz),
7.41 (t, 4H, J ) 7.5 Hz), 7.41-7.24 (m, 2H, NH), 7.32 (t, 4H, J )
7.6 Hz), 6.95 (d, 2H, J ) 7.0 Hz), 5.85 (dddd, 2H, J ) 7.0, 7.0,
10.2, 17.1 Hz), 5.15 (dddd, 2H, J ) 1.4, 1.4, 3.2, 17.1 Hz), 5.06
(dddd, 2H, J ) 1.0, 2.1, 3.2, 10.2 Hz), 4.43-4.35 (m, 2H), 4.35-
4.28 (m, 2H), 4.26-4.17 (m, 4H), 3.92 (dd, 2H, J ) 6.0, 16.7 Hz),
3.82-3.73 (m, 2H), 3.24-3.07 (m, 8H), 2.69-2.58 (m, 2H), 2.53-
2.43 (m, 2H), 1.74-1.59 (m, 2H), 1.52-1.40 (m, 4H), 1.39 (s,
9H) ppm; 13C NMR (100 MHz, acetone-d6) δ 172.7(C), 169.7(C),
157.4(C), 156.2(C), 144.9(C), 144.8(C), 142.0(C), 135.0(CH),
134.9(CH), 128.5(CH), 127.9(CH), 126.1(CH), 120.7(CH), 118.3-
(CH2), 79.3(C), 67.4(CH2), 56.0(CH), 55.5(CH2), 47.9(CH), 47.4-
(CH2), 45.0(CH2), 43.6(CH2), 43.5(CH2), 39.3(CH2), 36.9(CH2),
32.0(CH3), 28.6(CH3), 26.54(CH2) ppm; HRMS (ESI) m/z calcd
for C56H67N7NaO10 [M + Na]+ 1020.4847, found 1020.4851.
1
Inhibitor 3: H NMR (400 MHz, 15% CH3OD/CDCl3 with a
trace of TFA-d added) δ 7.39-7.28 (m, 10H), 5.15-5.05 (m, 2H),
4.17-4.09 (m, 2H), 3.90-3.72 (m, 4H), 3.52-3.11 (m, 4H), 3.00-
2.84 (m, 4H), 1.94-1.83 (m, 2H), 1.83-1.62 (m, 6H), 1.62-1.51
(m, 2H), 1.48-1.31 (m, 4H) ppm; 13C NMR (100 MHz, 15% CD3-
OD/CDCl3 with a trace of TFA added) δ 174.7(C), 174.6(C), 172.0-
(C), 170.9(C), 157.62(C), 157.56(C), 136.9(C), 129.0(CH), 128.6-
(CH), 128.2(CH), 67.4(CH2), 55.4(CH), 55.3(CH), 47.7(CH2),
45.0(CH2), 43.5(CH2), 43.3(CH2), 38.3(CH2), 35.8(CH2), 31.7(CH2),
30.1(CH2), 26.5(CH2), 26.2(CH2), 25.0(CH2), 24.6(CH2), 23.1(CH2)
ppm; HRMS (ESI) m/z calcd for C35H50N7O8 [M + H]+ 696.3721,
found 696.3738; m/z calcd for C35H49N7NaO8 [M + Na]+ 718.3541,
found 718.3557.
1
Diene 10: H NMR (400 MHz, CDCl3) δ 7.98-7.61 (m, 2H),
7.49-7.06 (m, 4H), 6.12-5.86 (m, 0.4H, NH minor rotamer), 5.77
(dddd, 2H, J ) 7.1, 7.1, 10.0, 17.1 Hz), 5.57-5.46 (m, 1.6H, NH
major rotamer), 5.15 (d, 2H, J ) 17.0 Hz), 5.11 (d, 2H, J ) 10.1
Hz), 4.59-4.36 (m, 2H), 4.18-4.07 (m, 2H), 4.04-3.81 (m, 4H),
3.35-3.09 (m, 8H), 2.67-2.56 (m, 2H), 2.56-2.43 (m, 2H), 2.41-
2.29 (m, 4H), 2.19-2.05 (m, 2H), 1.96-1.78 (m, 2H), 1.61-1.47
(m, 2H), 1.59-1.48 (m, 4H), [1.46 (s), 1.43 (s), 1.43 (s), 45H total]
ppm; 13C NMR (100 MHz, CDCl3) δ 172.9(C), 172.0(C), 171.6-
(C), 169.2(C), 155.9(C), 133.4(CH), 133.3(CH), 118.7(CH2), 118.6-
(CH2), 82.1(C), 79.8(C), 79.5(C), 69.6(C), 54.2(CH2), 53.6(CH),
53.4(CH), 53.2(CH), 47.0(CH2), 43.2(CH2), 38.8(CH2), 36.2(CH2),
32.1(CH2), 31.8(CH3), 30.9(CH3), 29.3(CH3), 28.7(CH2), 28.5(CH3),
28.4(CH3), 28.0(CH3), 26.5(CH2), 25.8(CH2) ppm; HRMS (ESI)
m/z calcd for C54H93N9NaO16 [M + Na]+ 1146.6633, found
1146.6594.
Inhibitor 4: 1H NMR (400 MHz, 10% CD3OD/CDCl3 with ca.
3% TFA) δ 7.76 (d, 4H, J ) 7.3 Hz), 7.62 (d, 4H, J ) 6.6 Hz),
7.40 (t, 4H, J ) 7.05 Hz), 7.31 (t, 4H, J ) 6.8 Hz), 4.52-4.33 (m,
4H), 4.25-4.17 (m, 2H), 4.15-4.08 (m, 2H), 3.90-3.77 (m, 4H),
3.47-3.13 (m, 4H), 3.01-2.83 (m, 4H), 1.93-1.49 (m, 8H), 1.45-
1.16 (m, 6H) ppm; 13C NMR (100 MHz, 10% CD3OD/CDCl3 with
ca. 1-2% TFA) δ 173.9, 173.8, 171.3, 170.2, 161.5, 156.7, 156.7,
(23) Khan, M. O. F.; Austin, S. E.; Chan, C.; Yin, H.; Marks, D.;
Vaghjiani, S. N.; Kendrick, H.; Yardley, V.; Croft, S. L.; Douglas, K. T. J.
Med. Chem. 2000, 43, 3148-3156.
(24) Saravanamuthu, A.; Vickers, T. J.; Bond, C. S.; Peterson, M. R.;
Hunter, W. H.; Fairlamb, A. H. J. Biol. Chem. 2004, 279, 29493-29500.
(25) Garforth, J.; Yin, H.; McKie, J. H.; Douglas, K. T.; Fairlamb, A.
H. J. Enzyme Inhib. 1997, 12, 161-173.
3692 J. Org. Chem., Vol. 72, No. 10, 2007