Chemistry Reference
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Table 3.3
In vitro GK enzyme kinetics data for compounds 2a and 9a.
Parameter
2a
9a
V
max
(mmol min
1
mg protein
1
)
Control
18.0
19.1
1 mM (fold change versus control)
24.7 (1.4)
23.7 (1.2)
20 mM (fold change versus control)
31.2 (1.7)
26.1 (1.4)
S
0.5
(mmol L
1
)
Control
8.0
8.7
1 mM (fold change versus control)
5.2 (0.66)
3.3 (0.38)
20 mM (fold change versus control)
1.8 (0.22)
0.84 (0.10)
half-maximal velocity) for glucose (Figure 3.10C). The magnitude of these
effects at 1 mM and 20 mM activator concentrations was determined by fitting
the data to the Hill equation (Table 3.3). Compound 2a exerted a relatively
larger increase in V
max
(a 1.7-fold change for 2a versus a 1.4-fold change for 9a
at 20 mM concentration of each), whereas 9a showed a greater effect on redu-
cing the enzyme's S
0.5
for glucose (1.8mmol L
1
for 2a versus 0.84mmol L
1
for 9a at 20 mM concentration of each). Consistent with the role of pancreatic b-
cell GK, treatment of rodent islets with either 2a or 9a shifted the glucose-
stimulated insulin release curve to the left, as demonstrated in perifusion studies
using isolated pancreatic islets.
16
In order to better discriminate between the two GK activators, head-to-head
in vivo ecacy studies were performed. Acute oral administration of 2a and 9a
(50mg kg
1
) to male normal C57Bl/6J mice caused a statistically significant
reduction in fasting glucose levels and improvement in glucose tolerance rela-
tive to the vehicle-treated animals (Figure 3.11A). Compound 9a showed a
statistically significant superior effect during an oral glucose tolerance test
(OGTT) relative to 2a (Figure 3.11B). Furthermore, comparison of rat phar-
macokinetic (PK) parameters indicated that 9a displayed lower clearance and
higher oral bioavailability compared to 2a (Table 3.4).
Based on these and other safety related properties, 9a was advanced to a
single ascending dose (25, 100, 200, and 400mg) study in healthy volunteers.
Following an oral dose, 9a reduced fasting and postprandial glucose levels
(minimal effective dose was between 25 and 100mg) following an OGTT, was
well tolerated, and displayed no adverse effects related to drug administration
other than hypoglycemia at a single dose of 400mg, which defined the max-
imum tolerated dose.
15
Thus, 9a was the first GK activator that provided the
proof-of-concept for this mechanism in humans.
3.3 Lead Optimization and Discovery of Piragliatin
16
Compound 9a, however, caused reversible hepatic lipidosis in rats and dogs in
sub-chronic toxicology studies and thus was unsuitable for full clinical devel-
opment. With this observation, we were faced with a key question: is this
finding related to the compound structure or due to drug-induced GK
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