The Discovery of GS-9131, an Amidate Prodrug of a Novel Nucleoside Phosphonate HIV Reverse Transcriptase Inhibitor - Accounts in Drug Discovery Case Studies in Medicinal Chemistry - page 233

30h was cleared at a rate similar to 11, whilst ethylalanine 30g had a lower
clearance rate, consistent with this prodrug demonstrating high hepatic stabi-
lity. Although both of these prodrugs had lower Cat A cleavage rates than 11,
their loading eciencies were comparable to 11 as a result of the more favor-
able plasma exposures. Initial oral studies on 30g as a diastereomeric mixture
demonstrated modest oral bioavailability for the prodrug (12% F) and excel-
lent loading of the PBMCs with the diphosphate 22-DP. Both the ethyl ester
30g and cyclobutyl ester 30h prodrugs were separated using chiral chromato-
graphy into their individual diastereoisomers for further oral evaluation.
Although 30h had a higher Cat A cleavage rate and corresponding loading
eciency, the lower hepatic and intestinal stability resulted in a lower oral
bioavailability of the seperated prodrugs. Combined with potential toxicity
concerns due to the release of cyclobutanol, this prodrug was abandoned in
favor of the ethyl ester isomers 31 and 32. Oral administration of the diaster-
eomerically pure isomers 31 and 32 was undertaken at 3mg kg 1 in beagle dogs,
and established that the second eluting isomer 32 was the most promising
prodrug isomer, resulting in PBMC levels of the active diphosphate 22-DP that
exceeded 9 mM from a single dose. 29 The full pharmacokinetic profiles of 32 and
its parent are shown in Figure 10.8 and established that the prodrug is cleared
from circulation within the first 1-2 h and has a half-life of less than 20min.
Levels of diacid 22 in the plasma stay below 1 mM and peak in the first 2 h,
consistent with the elimination of the prodrug. This suggests the cleavage of the
prodrug in the liver and other sites is generating the diacid in the plasma. It is
important to note that iv dosing of diacid 22 does not lead to significant levels
of 22-DP inside PBMCs. Intracellular PBMC levels of 22 peaked early at
B

10 mM, more than 10-fold higher than the plasma levels. This early C max is
consistent with the prodrug clearance from plasma and supports the prodrug
being the dominant species for loading the PBMCs. Intracellular metabolism of

Figure 10.8 Plasma and PBMC pharmacokinetics of 32 delivered orally (3mg kg 1 )
in beagle dogs (n ¼ 3).