Synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxamides with broad-spectrum human herpesvirus polymerase inhibition

Article abstract of DOI:10.1016/j.bmcl.2008.06.060

A versatile synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxylate esters has been developed which has lead to the identification of a new series of non-nucleoside inhibitors of human herpesvirus polymerases HCMV, HSV-1, EBV, and VZV with high specificity compared to human DNA polymerases.

Full text of DOI:10.1016/j.bmcl.2008.06.060

Bioorganic & Medicinal Chemistry Letters 18 (2008) 3856–3859
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxamides
with broad-spectrum human herpesvirus polymerase inhibition
a
a
a
a
Mark E. Schnute a, , Roger J. Brideau , Sarah A. Collier , Michele M. Cudahy , Todd A. Hopkins ,
Mary L. Knechtel a, Nancee L. Oien a, Robert S. Sackett b, Allen Scott b, Mari L. Stephan b,
Michael W. Wathen a, Janet L. Wieber a
*
a Global Research and Development, Pfizer Inc., 700 Chesterfield Parkway West, St. Louis, MO 63017, USA
b Research API, Pfizer Inc., 7000 Portage Road, Kalamazoo, MI 49007, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A versatile synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxylate esters has been devel-
oped which has lead to the identification of a new series of non-nucleoside inhibitors of human
herpesvirus polymerases HCMV, HSV-1, EBV, and VZV with high specificity compared to human
DNA polymerases.
Received 2 March 2008
Revised 14 June 2008
Accepted 17 June 2008
Available online 20 June 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Cytomegalovirus
HCMV
Varicella-zoster
VZV
HSV
EBV
Infections by viruses of the Herpesvirus family lead to highly
debilitating or even life-threatening disease and pose a signifi-
cant threat to the general and especially the immunocompro-
mised population. Herpes simplex virus (HSV), human
cytomegalovirus (HCMV), Epstein–Barr virus (EBV), and vari-
cella-zoster virus (VZV) are the most widespread pathogens of
this family. Human cytomegalovirus is associated with clinical
symptoms such as pneumonia, retinitis, and graft rejection in
the immunocompromised, as well as congenital birth defects in
neonates.1 The existing therapies to treat HCMV infection are
hampered by the development of drug resistance and significant
drug-associated toxicities which limit their duration of use.2
Childhood vaccination has reduced the incidence of chickenpox
resulting from primary infection by varicella-zoster virus; how-
ever, reactivation of latent infection in the form of herpes zoster
(shingles) still represents a significant risk to the elderly and
those who are immunocompromised.3 Acyclovir, Famciclovir,
and Valacyclovir are prescribed for VZV infection though at high
doses due to their modest potency against the virus.3d Initial
infection by Epstein–Barr virus resulting in infectious mononu-
cleosis can have a profound personal impact, and there is a
strong association for the role of the virus in the development
of post-transplant lymphoproliferative disease and lympho-
mas.1a,4 A well tolerated, broad-spectrum agent against herpesvi-
rus infection is unfortunately not currently available and would
constitute a significant break-through in this field.
Recently, we have described two novel series of 4-oxo-4,7-
dihydrothieno[2,3-b]pyridine-5-carboxamides (1a and 1b) which
were broad-spectrum inhibitors of herpesvirus polymerases, Fig-
ure 1.5 One focus of our SAR program was to explore alternative
bicyclic ring systems, and of specific interest was furo[2,3-b]pyr-
idine based on the bioisosteric replacement of sulfur for oxygen.
Initial efforts based on compound 1a (e.g., 7a, vide infra) were
disappointing due to poor observed antiviral activity as well as
the limited scope and scalability of existing syntheses to the
O
1a
N
O
N
O
N
R
N
N
OH
H
S
Cl
1b
* Corresponding author. Tel.: +1 636 247 3662; fax: +1 636 247 5400.
Figure 1. Representative 4-oxo-4,7-dihydrothieno[2,3-b]pyridine-based herpesvi-
E-mail address: mark.e.schnute@pfizer.com (M.E. Schnute).
rus DNA polymerase inhibitors.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.06.060

M. E. Schnute et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3856–3859
3857
furo[2,3-b]pyridine ring system. The improved potency observed
with analogs such as 1b prompted us to reconsider this series
though overcoming the synthetic challenges were paramount.
Herein, we report the development of a versatile synthesis of
4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxylate esters and
the realization of antiherpetic activity.
Compared to the analogous quinoline and thieno[2,3-b]pyri-
dine ring systems, there are relatively few syntheses of the 4-
oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxylate scaffold in the
literature. Reported syntheses have utilized the Gould–Jacob
cyclization consisting of condensation of a 2-aminofuran with
ethoxyethylene malonate followed by thermal ring closure.6 In
fact, 2-aminofuran is not a viable precursor for this chemistry
due to its intrinsic instability,7 and syntheses have only been
successful starting from 2-aminofurans bearing strongly electron
withdrawing substituents at the 5-position (e.g., CO2CH3). For
our purposes we needed a route to access the Gould–Jacob prod-
uct derived from 2-aminofuran and preferably avoiding the high
temperature (240 °C) cyclization required for this chemistry. A
general and versatile synthesis of quinolone antibacterials has
been described, which relies on an intramolecular SNAryl substi-
tution of an electron deficient fluorobenzene by an enamine to
construct the pyridinone ring.8 A similar approach has been re-
ported to thieno[2,3-b]pyridin-4-ones utilizing 2-chlorothiophene
derivatives.9 To our knowledge this approach has not been ap-
plied to the synthesis of furo[2,3-b]pyridin-4-ones, and we envi-
sioned that such an approach would address the limitations of
the existing Gould–Jacob route. Chlorination of the lithium dian-
ion derived from 3-furoic acid (2) with hexachloroethane affor-
ded 2-chloro-3-furoic acid (3), Scheme 1.10 The magnesium
enolate of ethyl malonate was then acylated with the imidazo-
lide of 3 to provide b-ketoester 4.11 The condensation of 4 with
triethylorthoformate and acetic anhydride provided the 3-eth-
oxypropenoate which underwent facile addition of amines
(RNH2, Table 1) to afford the corresponding enamine with elim-
ination of ethanol.12 Pyridinone ring closure was then affected
by the addition of potassium tert-butoxide to yield furo[2,3-
b]pyridin-4-one esters 5a–h in 30–67% yield from compound 3.
The above sequence allows for the introduction of a wide range
of functionality to the N7-position in an effective one-pot, three-
step procedure.
O
O
O
b
OH
O
O
O
X
Cl
2 X = H
3 X = Cl
4
a
O
O
c,d
O
4
O
N
R
5a-h
Scheme 1. Synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine ester 5a–h. Reagents
and conditions: (a) LDA, THF, ꢀ70 °C; C2Cl6, 82%; (b) CDI, THF; potassium ethyl
malonate, CH3CN, MgCl2, Et3N, 25 °C; (c) (EtO)3CH, Ac2O, 135 °C; (d) RNH2, THF,
25 °C; t-BuOK, 30–40 °C.
Following the precedent of Mannich elaboration of the C2-posi-
tion established in the thienopyridine series5 and further sup-
ported by the increased relative rate of electrophilic aromatic
substitution for furan compared to thiophene,13 a similar strategy
in the furopyridine series was explored. Indeed furopyridines 5a–
h reacted with 4-methylenemorpholin-4-ium chloride14 in aceto-
nitrile at 60 °C to provide 6a–h in good yields, Scheme 2. The reg-
iochemistry of addition was confirmed by single crystal X-ray.15
Subsequent direct transformation to the corresponding 4-chloro-
benzylamides 7a–h followed by treatment with ethylchlorofor-
mate provided the alkylchlorides 8a–h which were reacted with
(R)-1-(2-furyl)-2-(methylamino)ethanol16 to afford compounds
9a–h.
Table 1
HCMV polymerase and antiviral inhibition for compounds 9
Compound
R
HCMV polymerase
IC50 (nM)
HCMV antiviral
IC50 (nM)
a
a
9a
9b
9c
9d
9e
9f
CH3
CH2CH3
C3H5
CH2CH2CH3
Phenyl
Pyridin-2-yl
CH2CH2Ph
CH2CH2N(C2H5)2
76 ( 5)
510
420
1120
830
140
26 ( 10)
700
1200
50 ( 10)
9g
9h
690
3920
a
Compounds 9a–h were assayed for their ability to inhibit the
Values are means of at least two experiments where standard deviation is given
in parentheses, see Table 4 for reference compounds.
incorporation of 3H-labeled nucleotide into template-primer by
O
O
O
O
O
O
a
b
N
N
N
5a-h
O
H
O
O
N
R
Cl
N
R
6a-h
7a-h
O
O
O
O
Cl
N
c
d
N
N
H
7a-h
H
O
O
O
OH
N
R
Cl
N
R
Cl
8a-h
9a-h
Scheme 2. Synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine amides 9a–h. Reagents and conditions: (a) 4-methylenemorpholin-4-ium chloride, CH3CN, 60 °C; (b) 4-
chlorobenzylamine, ethylene glycol, 130 °C; (c) ethylchloroformate, CHCl3; (d) (R)-1-(2-furyl)-2-(methylamino)ethanol, DMF, i-Pr2EtN, 90 °C.

3858
M. E. Schnute et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3856–3859
improvement over the corresponding thienopyridine analog
(0.4
M).5b
l
Further refinements to the benzylamide and aminoalcohol
substituent were next explored. Docking of compound 9a into
the previously reported crystal structure of HSV-1 DNA poly-
merase18 revealed that the 4-chlorobenzylamide substituent fits
into a well-defined pocket partially border by valine 823, Figure
2. We had previously established that mutation of this residue
conveys resistance to related quinolone and thienopyridine
antivirals suggesting that this pocket plays a critical role in
compound binding.5a,19 A small bulge at the end of the pocket
nicely accommodates the chlorine substituent found in 9a. To
determine if chlorine was the optimal substituent, ester 10
was condensed with several 4- and 3,4-substituted benzylam-
ines to provide compounds 11a–f, Scheme 3. Indeed, the chlo-
rine substituent contributes substantially to the polymerase
inhibition of 9a as compared to benzyl alone 11f while alterna-
tive substituents at the 4-position (Br, F, CH3, and CF3) re-
mained inferior to chloride itself, Table 2. Compound 8a was
alkylated with a variety of N-methylaminoalcohols, Scheme 4.
Most promising were compounds 12a–c each substituted by a
six-membered nitrogen heterocycle, which showed good HCMV
polymerase inhibition and antiviral activity, Table 3. Further
profiling of compounds 9a and 12a indicated that both com-
pounds were broad-spectrum inhibitors of herpesvirus poly-
merases including HCMV, HSV, and VZV, while not being
Figure 2. Representation of benzylamide binding pocket based on docking of
compound 9a into crystal structure of HSV-1 DNA polymerase (Ref. 18).
purified HCMV DNA polymerase using a scintillation proximity
assay and for antiviral activity in an HCMV plaque reduction as-
say, Table 1.17 Polymerase inhibition was noticeably sensitive to
the nature of the N7-substituent with methyl (9a) and pyrimi-
din-2-yl (9f) being the best suggesting the potential for a steric
clash above or below the ring plane in this region. This would be
consistent with a binding model in which the furopyridine ring
system stacks between the polymerase and the DNA duplex.18
Overall, methyl substitution (9a) imparted the best combination
active against human DNA polymerases (a, c, and d), Table 4.
Broad-spectrum herpes antiviral activity was also demonstrated
with potencies comparable to thienopyridine 1b. Compound 9a
demonstrated good oral bioavailability in rat (67%) and in dog
(76%).20
By applying SAR learnings from previous thieno[2,3-b]pyri-
dine herpesvirus polymerase inhibitors, furo[2,3-b]pyridine
analogs have been identified demonstrating comparable po-
tency to their progenitors with improved aqueous solubility.
Compound 9a exhibited broad-spectrum herpes antiviral activ-
ity superior to that of available therapies with respect to
HCMV, VZV, and EBV. The full optimization of this chemical
series was made possible by the development of a practical
synthesis of 4-oxo-4,7-dihydrofuro[2,3-b]pyridine-5-carboxyl-
ate esters.
of potency and aqueous solubility (60 lM) which was a marked
O
N
O
a,b
N
O
6a
O
O
OH
10
O
O
Y
X
O
O
N
c
N
10
H
N
a
N
O
O
OH
8a
N
R'
H
O
OH
N
Cl
11a-f
12a-c
Scheme 3. Synthesis of amides 11a–f. Reagents and conditions: (a) ethylchloro-
formate, CHCl3, 38%; (b) (R)-1-(2-furyl)-2-(methylamino)ethanol, DMF, i-Pr2EtN,
90 °C, 72%; (c) benzylamine, ethylene glycol, 130 °C.
Scheme 4. Synthesis of amides 12a–c. Reagents and conditions: (a) i-Pr2EtN, DMF,
R0CH(OH)CH2NH(CH3), 90 °C.
Table 2
HCMV polymerase inhibition for compounds 11
a
Table 3
Compound
X
Y
HCMV polymerase IC50 (nM)
HCMV polymerase and antiviral inhibition for compounds 12
9a
Cl
Br
F
H
H
H
F
H
H
H
76 ( 5)
340 ( 80)
610
1020
3140
Compound
R0
HCMV polymerase
IC50 (nM)a
HCMV antiviral
11a
11b
11c
11d
11e
11f
a
IC50 (nM)
F
9a
2-Furyl
76 ( 5)
220 ( 80)
170
26 ( 10)
170 ( 20)
360 ( 5)
28 ( 15)
CH3
CF3
H
12a
12b
12c
Pyridin-2-yl
Pyrimidin-2-yl
Pyrazin-2-yl
6450
11,400
220
a
a
Values are means of at least two experiments where standard deviation is given
in parentheses, see Table 4 for reference compounds.
Values are means of at least two experiments where standard deviation is given
in parentheses, see Table 4 for reference compounds.

M. E. Schnute et al. / Bioorg. Med. Chem. Lett. 18 (2008) 3856–3859
3859
Table 4
Broad-spectrum activity of 9a and 12a compared to that of 4-oxo-4,7-dihydrothieno[2,3-b]pyridine 1b, and established therapies
a
a,b
c
Compound
Polymerase IC50 (nM)
Antiviral IC50 (nM)
CC50
HCMV
HSV-1
VZV
a
/
c/d
HCMV
HSV-1
VZV
EBV
(lM)
1b
9a
12a
Ganciclovir
Acyclovir
Foscarnet
Aphidicolin
AZT-TPd
61 ( 1)
76 ( 5)
220 ( 80)
76
220
350
21
78
100
>20,000
>20,000
>20,000
100
3000
4300
5400
nd
2
20
90
nd
8100
200
400
nd
nd
6900
>100
>100
>100
>100
>100
26 ( 10)
170 ( 20)
1300
>20,000
2100
2,500
487 ( 74)
22,100
nd
nd
473 ( 90)
5800
<280 (
2600 (a
c
)
)
438 ( 136)
3300
2300 (d)
a
Values are means of at least two experiments where standard deviation is given in parentheses (nd = not determined).
Determined by plaque reduction assay (HCMV, Davis strain; HSV-1, KOS strain; VZV, Webster strain).
CC50, 50% cellular cytotoxicity in HFF cells derived from single determination, Ref. 21.
AZT-TP = zidovudine triphosphate.
b
c
d
became a solution as the mixture was stirred over 2 h. In a separate reaction
vessel, slurry of potassium ethyl malonate (530.5 g, 3.12 mol) and dry
Acknowledgments
a
acetonitrile (4560 mL) was cooled to 10–15 °C. Triethylamine (435 mL,
3.12 mol) was then added. Solid anhydrous MgCl2 (370 g, 3.89 mol) was
added portion-wise maintaining the reaction temperature below 25 °C. The
resulting slurry was stirred at room temperature for 2.5 h, and then the
previously prepared imidazolide solution was added via cannula maintaining
the temperature below 25 °C. The slurry was stirred overnight at room
temperature. The solvent was removed in vacuo and the residue was treated
with 0.5 N HCl (9.3 L) and toluene (4.6 L). The aqueous layer was extracted
with toluene (1.9 L) and the combined organic layers were washed with
saturated aqueous sodium bicarbonate (1.9 L). The organic layer was
concentrated to provide 317.9 g of 4 as a brown oil which could be used as
is in subsequent steps. 1H NMR (400 MHz, DMSO-d6) d ppm (major isomer)
7.80 (d, J = 2.3 Hz, 1H), 7.01 (d, J = 2.3 Hz, 1H), 4.09 (q, J = 7.1 Hz, 2H), 3.95 (s,
2H), 1.15 (t, J = 7.1 Hz, 3H); (minor isomer) 12.35 (s, 1H), 7.79 (d, J = 2.3 Hz,
1H), 7.00 (d, J = 2.3 Hz, 1H), 5.69 (s, 1H), 4.18 (q, J = 7.1 Hz, 2H), 1.22 (t,
J = 7.0 Hz, 3H).
The authors thank Garold Bryant for X-ray structural determi-
nations, Francis Schwende, Karen Wilkinson, and Bob Rush for
pharmacokinetic analysis, Kevin Stafanski for solubility measure-
ments, and Paul Herrington, William Perrault, William McGhee,
and Thomas Beauchamp for providing chiral aminoalcohols.
References and notes
1. (a) Fishman, J. A. N. Engl. J. Med. 2007, 357, 2601; (b) Steininger, C.;
Puchhammer-Stöckl, E.; Popow-Kraupp, T. J. Clin. Virol. 2006, 37, 1; (c)
Gaytant, M. A.; Steegers, E. A. P.; Semmekrot, B. A.; Merkus, H. M. M. W.;
Galama, J. M. D. Obstet. Gynecol. Surv. 2002, 57, 245.
2. (a) Gilbert, C.; Boivin, G. Antimicrob. Agents Chemother. 2005, 49, 873; (b)
Chakrabarty, A.; Pang, K. R.; Wu, J. J.; Narvaez, J.; Rauser, M.; Huang, D. B.;
Beutner, K. R.; Tyring, S. K. Expert Opin. Emerg. Drugs 2004, 9, 237; For
prospective therapies see: (c) Biron, K. K. In New Concepts of Antiviral Therapy;
Bogner, E., Holzenburg, A., Eds.; Springer: Dordrecht, The Netherlands, 2006; pp
309–336.
3. (a) Heininger, U.; Seward, J. F. Lancet 2006, 368, 1365; (b) Guenther, L. C. Exper.
Rev. Dermatol. 2006, 1, 607–618; (c) Kimberlin, D. W.; Whitley, R. J. N. Engl. J.
Med. 2007, 356, 1338; (d) Gnann, J. W., Jr.; Whitley, R. J. N. Engl. J. Med. 2002,
347, 340.
4. Thorley-Lawson, D. A.; Gross, A. N. Engl. J. Med. 2004, 350, 1328.
5. (a) Schnute, M. E.; Cudahy, M. M.; Brideau, R. J.; Homa, F. L.; Hopkins, T. A.;
Knechtel, M. L.; Oien, N. L.; Pitts, T. W.; Poorman, R. A.; Wathen, M. W.; Wieber,
J. L. J. Med. Chem. 2005, 48, 5794; (b) Schnute, M. E.; Anderson, D. J.; Brideau, R.
J.; Ciske, F. L.; Collier, S. A.; Cudahy, M. M.; Eggen, M.; Genin, M. J.; Hopkins, T.
A.; Judge, T. M.; Kim, E. J.; Knechtel, M. L.; Nair, S. K.; Nieman, J. A.; Oien, N. L.;
Scott, A.; Tanis, S. P.; Vaillancourt, V. A.; Wathen, M. W.; Wieber, J. L. Bioorg.
Med. Chem. Lett. 2007, 17, 3349.
12. General procedure for preparation of esters 5a–h.
A mixture of 4 (10.0 g,
46.2 mmol), triethylorthoformate (15.4 mL, 92.4 mmol), and acetic anhydride
(15.3 mL, 161.7 mmol) was heated to 135 °C with removal of ethyl acetate
distillate with
40 Torr (100 °C) and then at 0.2 Torr (65 °C, 1 h) to afford a black oil. The
resulting oil was dissolved in THF (50 mL) and corresponding amine
a Dean–Stark trap. After 3 h, volatiles were removed at
a
(50.8 mmol) was added while cooling in an ice bath. The mixture was
allowed to stir at room temperature for 20 h. A solution of potassium tert-
butoxide (1.0 M in THF, 50.8 mL, 50.8 mmol) was then added maintaining
the internal temperature below 0 °C. The mixture was allowed to warm to
room temperature and was then held at 30–40 °C for 1 h. The mixture was
diluted with ethyl acetate (400 mL) and satd aq ammonium chloride
(200 mL). The organic layer was washed with brine (50 mL), dried
(Na2SO4), and concentrated. The product was purified by column
chromatography. Yield from 3: 5a, 67%; 5b, 50%; 5c, 38%; 5d, 30%; 5e,
34%; 5f, 37%; 5g, 52%; 5h, 44%.
13. (a) Heaney, H.; Papageorgiou, G.; Wilkins, R. F. Tetrahedron Lett. 1988, 29, 2377;
(b) Heaney, H.; Papageorgiou, G.; Wilkins, R. F. Tetrahedron 1997, 53, 2941.
14. Dimmock, J. R.; Erciyas, E.; Bigam, G. E.; Kirkpatrick, D. L.; Duke, M. M. Eur. J.
Med. Chem. 1989, 24, 379.
15. X-ray determination employed the 4-bromobenzoate ester of 9a.
Crystallographic data have been deposited with the Cambridge
Crystallographic Data Centre as supplementary publication number CCDC
679175.
16. Tanis, S. P.; Evans, B. R.; Nieman, J. A.; Parker, T. T.; Taylor, W. D.; Heasley, S. E.;
Herrinton, P. M.; Perrault, W. R.; Hohler, R. A.; Dolak, L. A.; Hester, M. R.; Seest,
E. P. Tetrahedron: Asymmetry 2006, 17, 2154.
17. Assay methods have been previously decribed in Ref. 5 to assess inhibition of
compounds against viral DNA polymerases (HCMV, HSV-1, and VZV) and
6. (a) Snyder, H. R., Jr.; Ebetino, F. F. J. Heterocycl. Chem. 1966, 3, 202; (b) Bhupathy,
M.; Conlon, D. A.; Wells, K. M.; Nelson, J. R.; Reider, P. J.; Rossen, K.; Sager, J. W.;
Volante, R. P.; Dorsey, B. D.; Hoffman, J. M., Jr.; Joseph, S. A.; McDaniel, S. L. J.
Heterocycl. Chem. 1995, 32, 1283.
7. Ramsden, C. A.; Milata, V. Adv. Heterocycl. Chem. 2006, 92, 1.
8. Clay, R. J.; Collom, T. A.; Karrick, G. L.; Wemple, J. Synthesis 1993, 290.
9. El-Abadelah, M. M.; Sabri, S. S.; Al-Ashqar, H. A. Heterocycles 1997, 45, 255.
10. 2-Chloro-3-furoic acid (3). A solution of diisopropylamine (713 mL, 5.08 mol) in
dry THF (5.1 L) was cooled to less than ꢀ50 °C. A solution of 2.5 M n-butyl
lithium in hexanes (2033 mL, 5.08 mol) was added over 40 min maintaining
the temperature below ꢀ50 °C. A solution of 3-furoic acid (285 g, 2.54 mol) in
dry THF (1742 mL) was added maintaining the temperature below ꢀ70 °C. The
solution was stirred at ꢀ70 °C for 40 min and then
a
solution of
human DNA polymerases (a, c, and d). Antiviral activities against herpesviruses
were determined using plaque reduction assays (HCMV, VZV, and HSV-1) or
lytic replication (EBV) as described in Ref. 5.
hexachloroethane (662 g, 2.8 mol) in dry THF (1017 mL) was added
maintaining the temperature below ꢀ70 °C. The solution was stirred for 3 h
at ꢀ70 °C and then was allowed to warm to room temperature overnight. The
reaction mixture was quenched with water (10.9 L). The aqueous layer was
extracted with MTBE (5.8 L) and then acidified with 2 N HCl to a pH 1–2. The
product was extracted into ethyl acetate (2 ꢁ 3.8 L), and then the solvent was
removed in vacuo to provide a solid. Recrystallization from water (7.5 L)
provided 233 g (82%) of 3 as pale tan crystals on cooling to 0–5 °C overnight.
Mp 135.3–137.5 °C; 1H NMR (400 MHz, DMSO-d6) d ppm 13.06 (br s, 1H), 7.75
(d, J = 2.3 Hz, 1H), 6.80 (d, J = 2.3 Hz, 1H); 13C NMR (100 MHz, DMSO-d6) d ppm
162.42, 143.38, 140.12, 113.78, 112.65; MS (ESI+) m/z 147 (M+H). Anal. Calcd
for C5H3ClO3: C, 40.98; H, 2.06. Found: C, 40.83; H, 1.90.
18. Liu, S.; Knafels, J. D.; Chang, J. S.; Waszak, G. A.; Baldwin, E. T.; Deibel, M. R., Jr.;
Thomsen, D. R.; Homa, F. L.; Wells, P. A.; Tory, M. C.; Poorman, R. A.; Gao, H.;
Qiu, X.; Seddon, A. P. J. Biol. Chem. 2006, 281, 18193.
19. Thomsen, D. R.; Oien, N. L.; Hopkins, T. A.; Knechtel, M. L.; Brideau, R. J.;
Wathen, M. W.; Homa, F. L. J. Virol. 2003, 77, 1868.
20. Compounds were dosed IV at 5 mg/kg and PO at 15 mg/kg using a vehicle of
80/20 PEG400/0.01 M methanesulfonic acid.
21. Toxicity of compounds to non-infected mammalian cells was determined using
replicating HFF cells seeded as subconfluent monolayers and treated with
compound for 3 days. Cell viability determinations were performed using both
microscopic evaluation and a quantitative neutral red dye uptake assay as
previously described. Lowik, C. W. G. M.; Alblas, M. J.; van de Ruit, M.;
Papapoulos, S. E.; van der Pliijm, G. Anal. Biochem. 1993, 213, 426.
11. Ethyl 3-(2-chloro-3-furyl)-3-oxopropanoate (4).
A solution of 3 (228.0 g,
1.56 mol) in dry THF (373 mL) was added over 30 min to a slurry of N,N0-
carbonyldiimidazole (278.3 g, 1.72 mol) in dry THF (2280 mL). The slurry