Design and synthesis of a potent macrocyclic 1,6-napthyridine anti-human cytomegalovirus (HCMV) inhibitors

Article abstract of DOI:10.1016/j.bmcl.2005.01.050

A novel class of macrocyclic 1,6-napthyridines designed to adopt the presumed bioactive conformation of anti-HCMV acyclic 1,6-napthyridines are described. Both 14- and 15-membered macrocycles were shown to be highly potent against HCMV HSV-1 and HSV-2.

Full text of DOI:10.1016/j.bmcl.2005.01.050

Bioorganic & Medicinal Chemistry Letters 15 (2005) 1693–1695
Design and synthesis of a potent macrocyclic 1,6-napthyridine
anti-human cytomegalovirus (HCMV) inhibitors
Guy Falardeau,a,* Hugo Lachance,a Annie St-Pierre,a Constantin G. Yannopoulos,a
b
Marc Drouin, Jean Bedard and Laval Chan
a
a
´
a
´
ViroChem Pharma Inc., 275 Armand-Frappier Blvd., Laval, Quebec, Canada H7V 4A7
b
´ ´
Universite de Sherbrooke, Sherbrooke, Quebec, Canada J1K 2R1
Received 5 August 2003; revised 17 January 2005; accepted 19 January 2005
Abstract—A novel class of macrocyclic 1,6-napthyridines designed to adopt the presumed bioactive conformation of anti-HCMV
acyclic 1,6-napthyridines are described. Both 14- and 15-membered macrocycles were shown to be highly potent against HCMV
HSV-1 and HSV-2.
Ó 2005 Elsevier Ltd. All rights reserved.
Of the DNA viruses, the herpes group is the source of
the most common viral illnesses in man. The group con-
sists of herpes simplex virus (HSV) type I and II, vari-
cella zoster (VZV), Epstein-Barr virus (EBV), and
human cytomegalovirus (HCMV).1 As with other her-
pes viruses, infection with HCMV leads to a lifelong
association of virus and host. Following a primary infec-
tion, virus may be shed for a number of years. Infection
in healthy individuals is frequently asymptomatic and
up to 80% of the adult population harbors the virus in
latent form. In immunocompromised individuals, such
as chemotherapy patients, organ transplant patients
and particularly AIDS sufferers, latent HCMV can be
re-activated resulting in microcephaly, hepatospleno-
megaly, jaundice, convulsive seizures, mononucleosis,
retinitis, and even death. With the introduction of
HAART and improved patientsꢀ compliance, the inci-
dence of CMV infection has greatly diminished in the
past few years in the AIDS population.1d
infections, as have the nucleoside analogs, cytosine-ara-
binoside, adenine-arabinoside, iodoxy-uridine, and acy-
clovir (ACV), which is presently the treatment of choice
for herpes simplex type I infection. Unfortunately, drugs
such as acyclovir that have proven effective to treat cer-
tain herpes viruses infections are not sufficiently effective
to treat HCMV. And, drugs currently used to treat
HCMV infection, such as ganciclovir (GCV), cidofovir
(CDV), foscarnet (PFA), an valganciclovir, an orally
bioavailable valine ester prodrug of ganciclovir, lack
the acceptable side effect and safety profiles of the drugs
approved for treatment of other herpes viruses.2
As part of our ongoing search for new anti-HCMV com-
pounds, we have discovered the [1,6]-naphthyridines,3 a
novel class of inhibitors in which compound 1 (Fig. 1) is
25 times more potent than GCV. The resulting SAR
study showed that the optimal positions for the nitro-
gens on the naphthyridine ring were 1 and 6.3b In addi-
tion, substitution at C8 and a bulky alkoxy group at the
20-position were beneficial for activity.3a We have also
demonstrated that an internal hydrogen bond (IHB)
likely maintains the [1,6]-naphthyridine-2-carboxylic
acid benzylamide in an active conformation.4,5 Exami-
nation of the X-ray crystal structure of 1 reveals the
presence of two conformations. Conformer 1a with
two IHB, is believed to be the active conformation5
whereas, conformer 1b with only one IHB is about
0.4 kcal higher in energy than 1b thus explaining the
presence of the two conformers in the crystal structure.
A variety of drugs have been developed to treat herpes
virus infection, including naturally occurring proteins
and synthetic nucleoside analogs. For example, inter-
feron, has been used in the treatment of herpes virus
Keywords: Anti-proliferative agents; Anti-virals; Polycyclic hetero-
cyclic compounds; HCMV; HSV; Naphthyridine; Macrocycle.
*
Corresponding author. Tel.: +1 45 0978 8431; fax: +1 45 0978
7777; e-mail: gfalardeau@virochempharma.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.01.050

1694
G. Falardeau et al. / Bioorg. Med. Chem. Lett. 15 (2005) 1693–1695
N
N
a
N
b
OH
O
OH
N
N
N
Br
O
Br
N
O
5
6
4
HO
N
N
O
O
N
N
c
d
Br
N
N
O
O
n
(Bu)3Sn
n
7 n=1, 2
2 n=1
3 n=2
Figure 1. Crystal structure of 1.
Scheme 1. Reagents and conditions: (a) AcOH, Br2, 80%; (b) HOBT,
EDCI, 2-hydroxy-benzylamine, Et3N, DMF, 93%; (c) (n = 1) DEAD,
4-tributylstannanyl-but-3-en-1-ol, Et3N, DMF (53%); (n = 2) DEAD,
4-tributylstannanyl-pent-3-en-1-ol,
Et3N,
DMF
(50%);
(d)
Pd(dba)3ÆCHCl3, DMF, 110 °C, 3 h. n = 1, 35%; n = 2, 28%.
tion between phenol 6 and 4-tributylstannanyl-but-3-en-
1-ol giving E-vinylstannane 2 in 53% yield. The Stille
coupling7 membered macrocycle 3 was constructed from
the corresponding pent-4-en-1-ol.
Other approaches to form the macrocycle by an intra-
molecular amidation, olefin metathesis or phenolic
alkylation failed to give the desired product in useful
yields.
Examination of conformer 1a reveals that the C8 methyl
is in close proximity to the alkoxy moiety. We therefore
decided to evaluate the effect of introducing a covalent
link between C8 methyl and the alkoxy group on anti-
viral activity. A low energy computer generated model
of a 14-membered macrocycle 2 (Fig. 2) gave two con-
formers with an energy difference of 0.1 kcal, the differ-
ence in the 2 conformers resided mainly in the
orientation of the phenyl group. Interestingly, both con-
formers 2a and 2b could be superimposed on the open
conformer 1a. Encouraged by these results, we therefore
embarked on the synthesis of 14-membered macrocycle
2 as well as 15-membered macrocycle 3.
Both macrocycles were almost equipotent (Table 1) and
macrocycle 2 showed an impressive gain in activity com-
pared to the acyclic analog 1. The 14-membered macro-
cycle 2 with an IC50 of 0.6 ng/mL and a selectivity index
of 3000 is probably one of the most potent anti-HCMV
compound reported to date. In addition, as with the pre-
viously reported napthyridines, activity against a clinical
isolate (P8 low passage) and GCV resistant strains (UL
97 and UL 54 mutants) was retained thus, indicating
that the mode of action of 2 does not involve phospho-
transferase or viral DNA polymerase enzymes as tar-
gets. Macrocycles 2 and 3 were also tested against
herpes simplex virus (HSV) in order to determine the
specificity toward the herpesviredae family (Table 2).
The results are reported in Table 2. The 15-membered
macrocycle 3 is more potent than the 14-membered mac-
rocycle 2 against HSV-1 but interestingly, both are far
more potent against HSV-2 than both GCV and cidofo-
vir (CDV). Both macrocycles show IC50ꢀs of 4–9 ng/mL
against this virus and good selectivity indices.
Both macrocycles were constructed by an intramolecu-
lar Stille coupling between an appropriate vinyl stann-
ane and 8-bromo-napthyridine 5 (Scheme 1). Thus,
bromination of the carboxylic acid 43a with bromine in
acetic acid yielded the 8-bromo-[1,6]-naphthyridine-2-
carboxylic acid 5 in 80% yield. Reaction of the carbox-
ylic acid 5 with 2-hydroxy-benzylamine in the presence
of EDCI/HOBt gave amide 6 in 93% yield. The stannane
intermediate 7 was assembled using a Mitsunobu6 reac-
Furthermore, macrocycle 2 was successfully crystallized
and the X-ray structure (Fig. 3) indicated the presence of
two conformations similar to what was predicted by
modeling.
In summary, we have designed novel macrocyclic naph-
thyridines which are believed to adopt the bioactive con-
formation of the previously described acyclic
napthyridines. Subsequent X-ray crystallographic stud-
ies and the 2–3 log enhancement in potency provide evi-
dence that our hypothesis is correct. Since both the
Figure 2. Molecular model of macrocycle 2.

G. Falardeau et al. / Bioorg. Med. Chem. Lett. 15 (2005) 1693–1695
1695
Table 1. Anti-cytomegaloviral activity of acyclic and macrocyclic naphthyridine analogs against HCMV laboratory strains (Davis, Ad169), clinical
isolate (P8), and GCV resistant clinical isolates (C8704, D16)
a
Compound
Anti-viral activity: IC50 (lg/ml)
Cytotoxicity (lg/ml)
Davisb
Ad169c
P8c
C8704 (UL97 mutation)c
D16 (UL54 mutation)c
1
0.09
0.0006
ND
0.13
0.0052
ND
0.11
0.1
2.4d
1.8d
2
<0.0015
0.003
2.7
0.00024
ND
8.7
0.0056
ND
7.8
3
GCV
0.2e
2.2
1.2
>1000d
a IC50, represents the compound concentration required to reduce virus plaque formation by 50% when compared to control samples without
compound.
b Numbers represent mean of duplicate values (SD < 15%), all experiments were performed at least twice; HEL cell line was used for the anti-viral
assay.
c Numbers represent mean of duplicate values (SD < 15%); MRC-5 cell line was used for the anti-viral assay.
d Number represents CC50, which represents the compound concentration required to reduce neutral red uptake by 50% when compared to control
samples without compound, mean of duplicate values (SD < 15%), MRC-5 cell line was used for the cytotoxicity assay.
e Number represents MTC value, which is defined as the minimum concentration of compound required to induce microscopically detectable
alteration of cell morphology, mean of duplicate values (SD < 15%), HEL cell line was used for the cytotoxicity assay.
Table 2. Anti-herpes simplex activity (IC50) and cytotoxicity (MTC) of
macrocycles 2 and 3
Acknowledgements
We wish to thank Dr. Robert Sidwell, Institute for Anti-
viral Research, Utah State University, Logan, UT, USA
as well as Johan Neyts and E. De Clercq, Rega Institute,
Leuven, Belgium for providing the biological results;
Compound
Anti-viral
activity: IC50
(lg/ml)
Anti-viral
activity: IC50
(lg/ml)
Cytotoxicityb
a
a
(lg/ml)
HSV-1 (KOS)
0.391
0.098
HSV-2 (G)
0.009
0.004
MTC
6
25
´ `
Ms. Therese Godbout and Ms. Andree Brisebois for
their assistance in the preparation of this manuscript.
´
2
3
CDV
GCV
3.937
1.274
1.276
1.220
>100
>100
a IC50, represents the compound concentration required to reduce virus
cytopathic effect by 50% when compared to control samples without
compound, number represents mean of duplicate values (SD < 15%),
all experiments were performed at least twice. Vero cell line was used
for the anti-viral assay.
b MTC is defined as the minimum concentration of compound required
to induce microscopically detectable alteration of cell morphology,
mean of triplicate values (SD < 15%). Vero cell line was used for the
cytotoxicity assay.
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