inhibitors and explore the three-dimensional space of the
inhibitor binding site. While several modifications led to
inactive compounds, replacement of the quinolinone portion
of the inhibitor with a tetramic acid was investigated due to
its conservation of what was considered to be an essential
hydrogen bonding network along the interior of the mol-
ecule.7
The above synthetic sequence was employed utilizing a
variety of R-amino acids in order to rapidly investigate the
SAR of this novel inhibitor template (Table 1). Certain trends
Table 1. SAR of Benzothiadiazine-Substituted Tetramic Acids
R1
R2
% yielda
IC50 (nM)
[R]D
b
compound
6b
6c
6d
6e
6f
6g
6h
6i
H
Me
Me
H
H
Me
71
69
85
83
77
65
69
65
58
87
51
2059
1436
5068
3468
314
208
6394
247
546
19
+23.8°
Figure 1. Benzothiadiazinylquinolinone screening lead.
-CH2CH2-
Ph
H
H
i-Pr
Me
H
+127.8°
-47.5°
+45.0°
-10.5°
-43.8°
-16.6°
-142.1°
i-Pr
H
i-Pr
i-Bu
c-Hex
t-Bu
One of the most prominently employed methodologies in
the synthesis of tetramic acids is the Dieckmann cyclization.8
Unfortunately, under standard conditions (NaOMe, MeOH,
reflux),9 racemic products are often obtained. However, investi-
gations by Ley and co-workers demonstrated that such cycli-
zations can be performed with potassium t-butoxide in tert-
butyl alcohol at ambient temperature without racemization.10
Initial investigations into benzothiadiazine-substituted tet-
ramic acids began with the reductive amination11 of (S)-
phenylalanine methyl ester followed by acylation with
benzothiadiazine acid 412 (Scheme 1). Cyclization under the
6j
6k
6l
H
H
19
a Overall yield for two steps. b c )1.0, CH2Cl2.
became readily apparent, leading to highly potent inhibitors
of NS5B. First, the S configuration was greatly preferred
over the R configuration (e.g., 6g vs 6h). Second, inhibitor
potency improved with an increase in steric bulk proximal
to the stereocenter (R1 ) t-Bu ∼ c-Hex > i-Pr > Ph > Me
> H). Third, spirocyclic derivatives offered no obvious
potency advantage (e.g., 6e). Importantly, addition of a small
substituent to the R2 position of inhibitor 6g provided a
compound of roughly equal potency (6i, R2 ) Me), obviating
the risk of racemization.
Scheme 1. First Generation Approach to
Benzothiadiazine-Substituted Tetramic Acids
Given the success of the above solution-phase synthesis
of benzothiadiazine-substituted tetramic acids, an analogous
solid-phase approach was investigated employing Wang resin
(7) Tedesco, R.; Shaw, A. N.; Bambal, R.; Chai, D.; Concha, N. O.;
Darcy, M. G.; Dhanak, D.; Fitch, D. M.; Gates, A.; Gerhardt, W. G.;
Halegoua, D. L.; Han, C.; Hofmann, G. A.; Johnston, V. K.; Kaura, A.;
Liu, N.; Keenan, R. M.; Lin-Goerke, J.; Sarisky, R. T.; Wiggall, K. J.;
Zimmerman, M. N.; Duffy, K. J. J. Med. Chem. submitted.
(8) For a review of tetramic acids, see: Royles, B. J. L. Chem. ReV.
1995, 95, 1981-2001.
(9) Lacey, R. N. J. Chem. Soc. 1954, 850-854.
(10) Ley, S. V.; Smith, S. C.; Woodward, P. R. Tetrahedron 1992, 48,
1145-1174.
(11) Ramanjulu, J. M.; Joullie´, M. M. Synth. Commun. 1996, 26, 1379-
Ley conditions provided tetramic acid 6a in good overall
yield and high optical purity.13 Analogue 6a demonstrated
modest but encouraging activity against NS5B (∼3.5 µM),
prompting further investigation.14
1384.
(12) Acid 4 was prepared by hydrolysis (see Supporting Information for
details) of the known ethyl ester: Kovalenko, S. N.; Chernykh, V. P.;
Shkarlat, A. E.; Ukrainets, I. V.; Gridasov, V. I.; Rudnev, S. A. Chem.
Heterocycl. Compd. (Engl. Transl.) 1998, 34, 791-795.
(13) Optical purity was determined by chiral HPLC analysis (Chiralpak
AD column, 0.1% trifluoroacetic acid in ethanol, 1 mL/min, Tr (S) ) 13.8
min, Tr (R) ) 25.9 min). Analysis of a sample of 6a stored for >1 month
at ambient temperature showed no sign of racemization.
(14) A scintillation-proximity assay (SPA) using N-terminal-truncated
∆21-NS5B was employed for the determination of IC50 values. See refs 6
and 7 for details.
(6) Dhanak, D.; Duffy, K. J.; Johnston, V. K.; Lin-Goerke, J.; Darcy,
M. G.; Shaw, A. N.; Gu, B.; Silverman, C.; Gates, A. T.; Nonnemacher,
M. R.; Earnshaw, D. L.; Casper, D. J.; Kaura, A.; Baker, A.; Greenwood,
C.; Gutshall, L. L.; Maley, D.; DelVecchio, A.; Macarron, R.; Hofmann,
G. A.; Alnoah, Z.; Cheng, H. Y.; Chan, G.; Khandekar, S.; Keenan, R. M.;
Sarisky, R. T. J. Biol. Chem. 2002, 277, 38322-38327.
5522
Org. Lett., Vol. 7, No. 24, 2005