A switch in enantiomer preference between mitochondrial F1F 0-ATPase chemotypes

Article abstract of DOI:10.1016/j.bmcl.2005.03.115

The preferred absolute configuration of two series of F₁F ₀-ATP synthase inhibitors was determined. Although the configuration of the active enantiomer in each series is different, each series presents the same 'triaryl' pharmacophor

Full text of DOI:10.1016/j.bmcl.2005.03.115

Bioorganic & Medicinal Chemistry Letters 15 (2005) 2749–2751
A switch in enantiomer preference between mitochondrial
F1F0-ATPase chemotypes
Sharon N. Bisaha,a Mary F. Malley,b Andrew Pudzianowski,c Hossain Monshizadegan,d
Paulina Wang,d Cort S. Madsen,d Jack Z. Gougoutasb and Philip D. Steina,*
aDepartment of Discovery Chemistry, Bristol-Myers Squibb, Pharmaceutical Research Institute, PO Box 4000,
Princeton, NJ 08543-4000, USA
bDepartment of Solid State Structure, Bristol-Myers Squibb, Pharmaceutical Research Institute, PO Box 4000,
Princeton, NJ 08543-4000, USA
cDepartment of Computer-aided Drug Design Bristol-Myers Squibb, Pharmaceutical Research Institute, PO Box 4000,
Princeton, NJ 08543-4000, USA
dDepartment of Cardiovascular Biology, Bristol-Myers Squibb, Pharmaceutical Research Institute, PO Box 4000,
Princeton, NJ 08543-4000, USA
Received 22 February 2005; accepted 30 March 2005
Available online 4 May 2005
Abstract—The preferred absolute configuration of two series of F1F0-ATP synthase inhibitors was determined. Although the con-
figuration of the active enantiomer in each series is different, each series presents the same ÔtriarylÕ pharmacophore to the enzyme
binding site.
Ó 2005 Elsevier Ltd. All rights reserved.
Respiratory chain (or oxidative) phosphorylation is the
process by which cells obtain energy under aerobic con-
ditions, and ATP synthase (F1F0-ATPase) is the central
enzyme in this process. Under normal conditions in
mitochondria, this membrane-bound, multisubunit pro-
tein synthesizes ATP from ADP and Pi using a proton
gradient established and maintained by the electron
transport chain.1 Under hypoxic conditions, the proton
gradient disappears and F1F0-ATPase switches modes
to act as an ATP hydrolase.2 This hydrolase activity
wastes ATP, making it unavailable to drive cellular pro-
cesses, including those which maintain cellular function
and viability.
paper, we report the synthesis, absolute configuration,
and ATPase inhibitory activity for several analogs in
two closely related chemotypes (thiocarbamates and
acylguanidines) and the surprising finding that the two
series exhibit opposite stereospecificity for enzyme
inhibition.
The synthesis of the enantiomeric thiocarbamate inhibi-
tors 4 and 5 is outlined in Figure 1. The absolute config-
uration of 1a was assigned as R-based on both the mode
of synthesis5 and by correlation to econazole, an anti-
fungal agent of known absolute configuration.6 This
synthesis constitutes the first correlation of optical rota-
tion data7 to the absolute configuration of compounds
1–3. The synthesis and separation of acylguanidines 6a
and 6b were carried out as previously described.4a
Salvage of ATP through selective ATPase hydrolase
inhibition may provide a novel mechanism for the treat-
ment of unstable angina, MI, and other vascular dis-
eases.3 In pursuit of this objective, we and others have
disclosed selective inhibitors of mitochondrial ATP
hydrolase as potential antiischemic agents.4 In this
Crystallization of 6b from acetonitrile produced crystal-
line material suitable for the determination of a solid
state structure and assignment of the absolute configura-
tion by anomalous scattering.8 On this basis, the abso-
lute configuration of 6b is confirmed as S- (Fig. 2).
Keywords: Mitochondrial ATPase; Acylguanidine; Chiral synthesis.
*
Compounds 4–6 were evaluated as inhibitors of bovine
F1F0-ATPase using the assay systems previously
Corresponding author. Tel.: +1 609 252 6157; fax: +1 609 252
6804; e-mail: philip.d.stein@bms.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.03.115

2750
S. N. Bisaha et al. / Bioorg. Med. Chem. Lett. 15 (2005) 2749–2751
gest preference (5600-fold) is for 6b versus 6a, however,
the (S)-enantiomer is the more potent inhibitor.
Although initially surprising in light of the structural
similarity of the two chemotypes, the SAR is consistent
with the distinct conformational preferences of the two
chemotypes. The H-bonded conformation in the solid-
state structure of 6b represents the low energy confor-
mation about the acylguanidine.9 To overlay the three
aromatic rings of enantio-matching 5b onto the corre-
sponding groups in 6b, 5b would have to adopt a confor-
mation that is 1.7 kcal/mol higher than the preferred
thiocarbamate conformer, a situation that is energeti-
cally unfavorable. However, one of the low energy
conformers of enantio-opposite 5a can be superimposed
onto 6b if the overlay matches, respectively, the imidaz-
ole, the C-linked dichlorophenyl and the N-linked
dichlorophenyl of 5a with the imidazole, the C-linked
dichlorophenyl ring and the cyanobenzoyl groups of
6b. Qualitatively, this alignment of the aromatic rings
and their substituents appears better than all of the
possible alignments of 5b and 6b (Fig. 3).
Figure 1. (a) (ꢀ)-DIP-Cl, THF ꢀ25 °C, 16 h, 79%; (b) 5 N aq. NaOH,
THF; (c) (+)-DIP-Cl, THF ꢀ25 °C, 16 h, 98%; (d) imidazole, NaH,
DMF; ꢀ5 °C to rt, 16 h, 47%; (e) 4-Cl-C6H4-NCS or 2,4-Cl2-C6H3-
NCS, tert-butylimino-tri(pyrrolidino)-phosphorane, CH3CN, rt, 6 h,
74%.
Additionally, the role of the cyanobenzoyl moiety as a
pharmacophore is consistent with our previous SAR.10
So, although the absolute configurations of the two
chemotypes are different, the potent enantiomers (5a
and 6b) each present the hydrolase with an appropriate
ÔtriarylÕ pharmacophore array.
described3 and the data are collected in Table 1. The
pairwise comparisons of enantiomers in both chemo-
types (4a/b, 5a/b, and 6a/b) indicate enantiomer prefer-
ences. The preference for (R)- is moderate (28-fold) for
4a and 4b, but is stronger (116-fold) for 5a/b. The stron-
Figure 2. Solid state structure of 6b.

S. N. Bisaha et al. / Bioorg. Med. Chem. Lett. 15 (2005) 2749–2751
2751
Table 1. Data for inhibition of mitochondrial F1F0-ATP hydrolase and synthase by compounds 4-6
Compound
Absolute configuration
Inhibition of mATP hydrolase (nM)
Inhibition of mATP synthase (nM)
4a
4b
5a
5b
6a
6b
R
S
R
S
R
S
140 41
3600 1100
19 2.7
2200
>100,000 (racemate)
>100,000 (racemate)
>100,000
>100,000 (racemate)
>100,000
>100,000
>100,000
18 16
3. Atwal, K. S.; Wang, P.; Rogers, W. L.; Sleph, P.;
Monshizadegan, H.; Ferrara, F. N.; Traeger, S.; Green,
D. W.; Grover, G. J. J. Med. Chem. 2004, 47, 1081–1084.
4. (a) Atwal, K. S.; Ahmad, S.; Ding, C. Z.; Stein, P. D.;
Lloyd, J.; Hamann, L. G.; Green, D. W.; Ferrara, F. N.;
Wang, P.; Rogers, W. L.; Doweyko, L. M.; Miller, A. V.;
Bisaha, S. N.; Schmidt, J. B.; Li, L.; Yost, K. J.; Lan,
H.-J.; Madsen, C. S. Bioorg. Med. Chem. Lett. 2004, 14,
1027–1030; (b) Hamann, L. G.; Ding, C. Z.; Miller, A. V.;
Madsen, C. S.; Wang, P.; Stein, P. D.; Pudzianowski, A.
T.; Green, D. W.; Monshizadegan, H.; Atwal, K. S.
Bioorg. Med. Chem. Lett. 2004, 14, 1031–1034.
5. Brown, H. C.; Ramachandran, P. V. Acc. Chem. Res.
1992, 25, 16–24.
6. (a) La¨mmerhofer, M.; Lindner, W. Chirality 1994, 6, 261–
269; (b) Aasen, A. J.; Groth, P. Acta Chem. Scand. 1987,
A41(9), 496–500.
7. 1a [a]D ꢀ43 (c = 7.6 mg/mL); 1b [a]D +40 (c = 7.6 mg/mL);
rt
2a (½aꢁD ꢀ55 (c = 8.6 mg/mL); 2b ([a]D +56 (c = 6.3 mg/
mL); 3a ([a]D ꢀ79 (5.1 mg/mL); 3b ([a]D +83 (5.0 mg/mL);
4a ([a]D +85 (c = 5.6 mg/mL), 95% ee); 4b ([a]D ꢀ90
(c = 5.1 mg/mL), 97% ee); 5a ([a]D +68 (c = 5.2 mg/mL),
98% ee); 5b ([a]D ꢀ73 (c = 5.1 mg/mL), >99% ee). Rota-
tions were measured at rt in CHCl3. Enantiomeric excess
(% ee) was determined by chiral chromatography.
Figure 3. Overlay of thiocarbamate 5a (colored by atom type) and
acylguanidine 6b (salmon).
8. Crystallographic data (excluding structure factors) for 6b
have been deposited with the Cambridge Crystallographic
Data Centre as supplementary publication numbers
CCDC 251489 and 251490. Copies of the data can be
obtained, free of charge, on application to CCDC, 12
Union Road, Cambridge, CB2 1EZ, UK [fax: +44 (0)
1223 336033 or e-mail: deposit@ccdc.cam.ac.uk].
In summary, we have determined the absolute configur-
ations for two related series of potent mATPase inhibi-
tors. We found that the stereospecificity for hydrolase
inhibition in the two series is opposite to one another
and postulate that the SAR is consistent with present-
ation of a Ôtriaryl pharmacophoreÕ by both chemotypes.
9. Semi-empirical gas-phase conformations and heats of
formation were calculated using AMPACÕs SAM-1
parameter set. For SAM-1, see Dewar, M. J. S.; Jie, C.;
Yu, J. Tetrahedron 1993, 49, 5003–5038, AMPAC 6.55,
Semichem Inc., Box 1649, Shawnee KS 66222.
10. The conformationally similar cyanoguanidine-based
inhibitors lack this group and are generally inferior
inhibitors (Ref. 4a). In addition, the overlay aligns the
m-cyano substituent in 6b and the o-chloro group in 5a.
The presence and location of both of these substituents are
important for inhibitor potency (Ref. 4a and Table 1).
References and notes
1. For a general discussion see Lodish, H.; Baltimore, D.;
Berk, A.; Zipursky, S. L.; Matsudaira, P.; Darnell, J.
Molecular Cell Biology, 3rd ed.; W. H. Freeman and Co.:
New York, 1995, pp. 745–765.
2. (a) Rosulin, W.; Erickson, J. L.; Solaro, R. J. Am. J. Physiol.
1986, 250, H503–H508; (b) Jennings, R. B.; Reimer, K. A.;
Steenbergen, C. J. Mol. Cell. Cardiol. 1991, 23, 1383–1395.