Z. K. Sweeney et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4348–4351
4351
Acknowledgment
F
OH
NC
O
N
a
CO2Et
We thank Grace Lam, Witold Woroniecki, and Jessica Brilliant
for their analytical expertise.
O2
NC
Cl
Cl
References and notes
b, c, d
1. Spence, R. A.; Kati, W. M.; Anderson, K. S.; Johnson, K. A. Science 1995,
267, 988.
2. Balzarini, J. Curr. Top. Med. Chem. 2004, 4, 921.
3. Sorbera, L. A.; Serradell, N.; Bolos, E.; Rosa, J. Drugs Future 2007, 32, 1046.
4. Sweeney, Z. K.; Dunn, J. P.; Li, Y.; Heilek, G.; Dunten, P.; Elworthy, T. R.; Han, X.;
Harris, S. F.; Hirschfeld, D. R.; Hogg, J. H.; Huber, W.; Kaiser, A.; Kertesz, D. J.;
Kim, W.; Roepel, T.; Mirzadegan, M. G.; Saito, Y. D.; Silva, T. M. P. C.; Swallow,
S.; Tracy, J. L.; Villasenor, A.; Vora, H.; Zhou, A. S.; Klumpp, K. Bioorg. Med. Chem.
Lett. 2008, 18, 4352.
F
e, f
NHNH2
NC
O
Br
7c
O
Cl
Scheme 1. Reagents: (a) KOtBu, ethyl (2, 3-difluoro-4-nitrophenyl)acetate, THF,
5. Enzymatic assay: IC50 values were obtained from a scintillation counting assay
using poly rA primer and tritiated dTTP.
t
90%; (b) Fe, NH4Cl, EtOH, H2O, 91%; (c) BuONO, CuBr2, LiBr, CH3CN, 50%; (d) N2H4,
EtOH; (e) OCNCH3, THF; (f) KOtBu, tBuOH, 74% for three steps.
6. Replication assay: EC50 values were obtained following incubation of
compound with infected MT4 cells in the presence of 10% fetal bovine
serum. The assay assesses the reduction in the cytopathic effect of HIV-1 on
MT4 cells in the presence of HIV-RT inhibitors. Each value represents the
average of at least two independent assays.
7. Surface plasmon resonance experiments were performed using HIV-RT protein
immobilized to the CM5 sensor chip via an amide-coupling reaction. The
binding experiments were performed using Hepes buffer (50 mM Hepes (pH
8.0), 150 mM NaCl, 10 mM MgCl2, 0.01% Tween 20, 5% DMSO) as the running
buffer. For characterization of other NNRTIs using SPR see: Geitmann, M.;
Danielson, U. H. Bioorg. Med. Chem. 2007, 15, 7344.
8. CYP3A4 inhibition was assessed using 7-benzyloxy-4-(trifluoromethyl)-
coumarin as a fluorogenic substrate.
9. Interestingly, the differences in solubility between these inhibitors cannot
simply be attributed to differences in intermolecular interactions in the solid
state. The melting temperature of crystals of 1 (181–182 °C) is nearly identical
to that of 7c (185–186 °C).
The synthesis of inhibitor 7c is outlined in Scheme 1.14 An aro-
matic substitution reaction of 3-cyano, 5-chlorophenol with a
difluoronitrophenyl acetate provided the diaryl ether in excellent
yield. Reduction of the nitro group with iron, followed by a Sand-
meyer reaction gave the chlorinated product. Sequential reaction
of the ester with hydrazine and methyl isocyanate provided a
semicarbazide that could be dehydrated with a catalytic amount
of KOtBu in BuOH to form the triazolinone NNRTI.
t
In summary, a number of non-nucleoside inhibitors of HIV re-
verse transcriptase were prepared. Evaluation of the potency and
physical properties of these compounds led to the identification
of a series of triazolinones that strongly inhibited wild-type and
NNRTI-resistant viruses in vitro. These compounds had signifi-
cantly improved physical properties relative to their pyridazinone
analogs, and inhibitor 7c was determined to have excellent bio-
availability in animal studies. A deconstruction exercise was used
to determine the suitability of fragment-based approaches for lead
identification for the NNRTI binding site.
10. Abad-Zapatero, C. Expert Opin. Drug Disc. 2007, 2, 469.
11. For an early example of such
a ‘‘inhibitor deconstruction” exercise see:
Babaoglu, K.; Shoichet, B. K. Nat. Chem. Biol. 2006, 2, 720.
12. Hopkins, A. L.; Groom, C. R.; Alex, A. Drug Discovery Today 2004, 9, 430.
13. Hajduk, P. J. J. Med. Chem. 2006, 49, 6972.
14. For complete details of the synthesis of inhibitor 7c and other NNRTIs
described in this communication see: Saito, Y. D.; Smith, M.; Sweeney, Z. K. US
2007/078128 and Dunn, J. P.; Elworthy, T. R.; Stefanidis, D.; Sweeney, Z. K. WO
2006/010545.