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kinase inhibitory potency, regardless of the kind of
substituents at C20 of the pyridinyl ring moiety of the
C7 pyridopyrimidino core (Fig. 1). The implication of
this observation is that the C20 part of the molecule does
not bind to a specific site of the enzyme. In an effort to
find out the docking mode of this subtype of analogues
within adenosine kinase, we have studied a known
crystal structure of adenosine kinase8 and did molecular
modeling with our analogues. These studies suggested
that the terminal of the C7 substituent is likely projected
outside the binding pocket of the enzyme, which may
explain why the terminals of the C7 substituent did not
significantly alter AK inhibitory activity of these analo-
gues (Fig. 2). Furthermore, it has been noticed that the
majority of the metabolic reactions occur at C20 sub-
stituent. These studies and observations suggest that if
we find a suitable group at C20 to avoid rapid metabolism
occurring at that site, we could improve the PK profiles
of these analogues without adversely affecting their AKi
(adenosine kinase inhibitor) potency. Therefore, a wide
range of substituents at the C20 of the C7 pyridino moiety
could be screened to improve half-life and oral bioavail-
ability of analogue 9e in light our rational (Fig. 2).
Table 1 is a summary of typical examples of nitrogen
linked tertiary amino analogues. SAR results indicated
that analogues with cyclic tertiary amines exhibited
better oral activities than those with acyclic tertiary
amines. Compound 9a is a potent AKi in vitro (AK
(enzyme): IC50=4.1 (ꢁ0.020) nM; AK (intact cell):
IC50=42.3 (ꢁ5.6) nM). This compound also potently
reduced carrageenan-induced thermal hyperalgesia in
the rat (ED50=1.0 mmol/kg, ip). Its in vivo potency,
however, was diminished when the compound was
administrated orally (inflammatory hyperalgesia (po):
ED50>10 mmol/kg). The lack of oral activity of this
analogue (9a) was easily explained by its PK profile,
which revealed several unidentified metabolites and
poor plasma concentration of the parent compound
following oral administration in the rat.10 Cyclic tertiary
amine substituents on the C20 position also yielded
potent AK inhibitors. Five-membered ring pyrrolidino
analogues 9b and 9c had in vivo activity when adminis-
tered ip. However, these compounds had very low oral
bioavailability. Better oral activity and bioavailability
were achieved by expanding the five-membered ring to a
six-membered ring system at the C20 position of these
analogues (entries 5–8, Table 1). The 4-morpholinyl
analogue 9e (ABT-702) was one of the best analogues
and showed improved, broad spectrum oral efficacy.6
Although its plasma half-life still needed to be improved
[t1/2: 0.9 h (iv)], its oral bioavailability reached a rea-
sonable level (23% in rat and 59% in dog). Moving the
oxygen heteroatom of the morpholinyl substituent out-
side the piperidino ring system gave 4-hydroxy-piper-
idinyl derivative 9f, which had improved plasma half-
life of 2.1 h, oral bioavailability of 18.6%, and good in
vivo activity (inflammatory hyperalgesia in rat (po):
ED50=3 mmol/kg). Incorporating two oxygen hetero-
atoms in the spiral ketal derivative 9g further enhanced
the oral bioavailability (F=44.7%), and retained good
half-life (t1=2=2.4 h) following iv administration.
Finally, modification of ketal functionality to a 4-
OTHP oxime derivative 9h, improved plasma half-life to
t1=2=3.6 h, retained good oral bioavailability
(F=46.2%, po) in the rat and improved PK profiles in
the dog. Following oral administration, 9h exhibited an
ED50 of 1 mmol/kg in reducing carrageenan-induced
thermal hyperalgesia in the rat (Table 1).
After comparing analogues with different X linkages at
C20, the nitrogen atom was found to be the best among
carbon, oxygen, and sulfur9 in terms of in vitro and in
vivo potencies (Fig. 1). Less metabolic problems were
observed when X is a nitrogen atom. Analogues with
tertiary amines at C20 demonstrated good correlation
between intact cell inhibition values and in vivo efficacy.
The synthesis of the nitrogen-linked analogues was
simple and straightforward, which allowed access to a
variety of analogues in order to compare their physical
chemical properties, such as solubility, PK (pharmaco-
kinetic) profiles, as well as in vivo activities.
Figure 1.
Chemistry
The C20-derivatives are easily accessible and prepared as
shown in Schemes 1, 2, and 3. In the presence of a cat-
alytic amount of glycine, condensation of malono-nitrile
with 3-bromobenzaldehyde gave a quantitative yield of
2-(30-bromophenyl)-1,1-dicyanyl-ethene (3) (Scheme 1).
Figure 2. Molecular modeling shown here demonstrates that the oxy-
gen of the morpholino moiety of 9e (red) is outside the binding pocket
of the adenosine kinase.
Scheme 1.