ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, May 2009, p. 2052–2058
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Vol. 53, No. 5
Novel S-Adenosylmethionine Decarboxylase Inhibitors for the
Treatment of Human African Trypanosomiasis?†
Robert H. Barker, Jr.,1* Hanlan Liu,1Bradford Hirth,1Cassandra A. Celatka,1Richard Fitzpatrick,1
Yibin Xiang,1Erin K. Willert,2Margaret A. Phillips,2Marcel Kaiser,3Cyrus J. Bacchi,4,5
Aixa Rodriguez,4Nigel Yarlett,4,6Jeffrey D. Klinger,1and Edmund Sybertz1
Genzyme Corporation, 153 Second Avenue, Waltham, Massachusetts 024511; Department of Pharmacology, University of
Texas Southwestern Medical Center, 6001 Forest Park Road, Dallas, Texas 75390-90412; Swiss Tropical Institute,
Parasite Chemotherapy, Socinstrasse 57, P.O. Box CH-4002, Basel, Switzerland3; and Haskins Laboratory,4
Department of Biological and Health Sciences,5and Department of Chemistry and
Physical Sciences,6Pace University, 41 Park Row, New York, New York 10038
Received 19 December 2008/Returned for modification 28 January 2009/Accepted 4 March 2009
Trypanosomiasis remains a significant disease across the sub-Saharan African continent, with 50,000 to
70,000 individuals infected. The utility of current therapies is limited by issues of toxicity and the need to
administer compounds intravenously. We have begun a program to pursue lead optimization around MDL
73811, an irreversible inhibitor of S-adenosylmethionine decarboxylase (AdoMetDC). This compound is potent
but in previous studies cleared rapidly from the blood of rats (T. L. Byers, T. L. Bush, P. P. McCann, and A. J.
Bitonti, Biochem. J. 274:527-533). One of the analogs synthesized (Genz-644131) was shown to be highly active
against Trypanosoma brucei rhodesiense in vitro (50% inhibitory concentration, 400 pg/ml). Enzyme kinetic
studies showed Genz-644131 to be approximately fivefold more potent than MDL 73811 against the T. brucei
brucei AdoMetDC-prozyme complex. This compound was stable in vitro in rat and human liver microsomal and
hepatocyte assays, was stable in rat whole-blood assays, did not significantly inhibit human cytochrome P450
enzymes, had no measurable efflux in CaCo-2 cells, and was only 41% bound by serum proteins. Pharmaco-
kinetic studies of mice following intraperitoneal dosing showed that the half-life of Genz-644131 was threefold
greater than that of MDL 73811 (7.4 h versus 2.5 h). Furthermore, brain penetration of Genz-644131 was
4.3-fold higher than that of MDL 73811. Finally, in vivo efficacy studies of T. b. brucei strain STIB 795-infected
mice showed that Genz-644131 significantly extended survival (from 6.75 days for controls to >30 days for
treated animals) and cured animals infected with T. b. brucei strain LAB 110 EATRO. Taken together, the data
strengthen validation of AdoMetDC as an important parasite target, and these studies have shown that analogs
of MDL 73811 can be synthesized with improved potency and brain penetration.
Sleeping sickness, or human African trypanosomiasis (HAT),
afflicts 50,000 to 70,000 people across sub-Saharan Africa, with
17,000 new cases reported in the year 2004 (9) and 10,769 re-
ported in 2007 (31). Untreated, the disease is inevitably fatal.
Current treatments include drugs first developed over 50 years
ago, and while not without efficacy, some have high toxicity and
generally need to be administered by intravenous (i.v.) infusion—
hardly a practical solution in locations where this disease is prev-
alent (17). Despite the obvious need for new, easily administered
therapies, the rate of development of new drugs for HAT by the
Drugs for Neglected Diseases Initiative (DNDi) was formed in
2003 to facilitate the formation of partnerships among industry,
academia, and public-sector organizations to develop affordable
solutions for this urgent unmet medical need (www.dndi.org).
Studies presented here were conducted under one such partner-
Polyamines are small-molecule cationic structures that are
critical to the survival of eukaryotic cells, including trypano-
somes (2, 4, 19). Difluoromethyl ornithine (DFMO) is an in-
hibitor of ornithine decarboxylase, a key enzyme in the poly-
amine biosynthetic pathway. DFMO is an effective and
relatively well tolerated agent for the treatment of the central
nervous system (CNS) second stage of HAT caused by
Trypanosoma brucei gambiense. However, DFMO must be
given as an i.v. infusion of up to 30 g per day four times per day
for 14 days. The amount required and frequency of dosing
demonstrate the limitations of this drug in terms of potency,
cost, and logistics of administration in the field (10, 16). Nev-
ertheless, the extraordinary efficacy of DFMO serves to vali-
date the polyamine pathway as an attractive target for new
drug discovery for trypanosomiasis. A potent, safe, orally bio-
available inhibitor of polyamine biosynthesis would represent a
major breakthrough for the treatment of this disease.
A second key enzyme in the polyamine biosynthetic pathway is
S-adenosylmethionine decarboxylase (AdoMetDC). AdoMetDC
somatid parasites; the functional form of the enzyme is a het-
erodimer between the active subunit and a paralog (termed
prozyme) that arose through gene duplication (32). Gene knock-
out and small interfering RNA experiments have indicated that
suppression of this enzyme by knockdown of either AdoMetDC
or its regulatory subunit prozyme is lethal to the parasite (33).
* Corresponding author. Mailing address: Genzyme Corporation,
153 Second Avenue, Waltham, MA 02451. Phone: (781) 434-3425. Fax:
(781) 434-3400. E-mail: Robert.firstname.lastname@example.org.
† Supplemental material for this article may be found at http://aac
?Published ahead of print on 16 March 2009.
Inhibitors of this enzyme have been shown to be highly efficacious
in killing trypanosomes in vitro (3, 14) and in curing T. brucei-
infected mice (6, 8). However, these agents lack both the potency
and, especially, the pharmacokinetic (PK) and tissue (CNS) dis-
tribution characteristics (11) that are essential to meet the im-
proved target product profile for a new antitrypanosomal drug.
MDL 73811 is an irreversible inhibitor of AdoMetDC and is
believed to form a Schiff base with a pyruvate group within the
active site of the enzyme (13). The compound displays a high
level of selectivity (?100-fold) for killing parasites compared
with its toxicity for mammalian cells (24). Reasons for this
selectivity are not clear, but it has been postulated that the
compound is more readily taken up by the parasites (12).
Alternatively, selectivity may result from differences in enzyme
turnover (33). The mechanism of killing is believed to be as-
sociated with the buildup of S-adenosylmethionine (11). Re-
cent studies utilizing RNA interference silencing, however,
suggest that a major method by which AdoMetDC inhibition
kills trypanosomes is by depleting reserves of trypanothione
(33). Finally, MDL 73811 induces increased expression of the
prozyme in T. brucei bloodstream form parasites, providing
strong evidence that the primary target of MDL 73811 respon-
sible for parasite death is AdoMetDC inhibition (33). MDL
73811 has been demonstrated to reduce parasitemia within 5 h
and to effect cures of acute infections in T. brucei brucei- and T.
brucei rhodesiense-infected mice when administered at a dose
of 20 mg/kg of body weight twice a day (BID) for 4 days (11,
24). This compound was originally developed in the 1980s by
Merrell-Dow as an anticancer therapy.
Despite its trypanocidal activity, MDL 73811 is not itself an
attractive drug candidate for a number of reasons. This com-
pound is not effective as monotherapy against the CNS stage of
infection, although it is curative when given in combination
with DFMO (6). It has been hypothesized that DFMO may in
some way temporarily alter the blood-brain barrier (BBB) to
effect this; however, data to support this contention are cur-
rently lacking. Recent studies (28) showed that DFMO by itself
penetrated the BBB poorly and that addition of 250 ?M
DFMO did not improve the uptake of other solutes, although
a statistically significant increase in BBB penetration was seen
at day 28 and later after the trypanosome infection. Finally, the
drug has poor oral bioavailability and was thought to have
limited metabolic stability (P. Casara, personal communica-
tion). We therefore initiated a program to synthesize analogs
of MDL 73811 and determine whether they could overcome
these issues. Studies reported here describe initial progress.
MATERIALS AND METHODS
Chemicals. MDL 73811 and analogs were synthesized as described in the
Trypanosomes in vitro. In vitro trypanosome killing assays were performed as
previously described using Trypanosoma brucei rhodesiense strain STIB 900, a
clone which is known to be susceptible to all currently used drugs. Briefly, serial
dilutions of drugs (90 to 0.123 ?g/ml) in supplemented minimal essential medium
(GIBCO-BRL catalog no. 072-1100) were inoculated with 104bloodstream trypo-
mastigotes and incubated 72 h, and then viability was determined using Alamar
Trypanosomes in vivo. Collaborators utilized two different protocols. In vivo
studies at the Swiss Tropical Institute (STI) were performed as previously de-
scribed (30) under a protocol reviewed and approved by the local veterinary
authorities of the Canton Basel-Stadt. Mice were infected with 1 ? 104trypano-
somes (T. b. brucei strain STIB 795) on day 0 and then treated once/day (QD)
intraperitoneally (i.p.) with 50 mg/kg test compound for 4 days starting on day 3.
Animals were assessed by microscopic examination of blood smears twice/week
through day 30. Untreated animals generally were moribund and were eutha-
nized by days 7 to 9. Studies conducted at Pace University (under a protocol
approved by the university’s Institutional Animal Care and Use Committee)
utilized the LAB 110 EATRO strain of T. b. brucei as previously described (8).
Briefly, groups of five animals were infected i.p. on day 0 with 2.5 ? 105parasites,
and dosing was initiated on day 1. Genz-644131 and MDL 73811 were dosed at
50 mg/kg/day i.p. either QD for 7 days or split and administered BID for 4 days.
Treatment controls received 2 mg/kg pentamidine QD i.p. for 4 days. Animals
were assessed by microscopic examination of at least 20 fields of wet blood
smears twice/week through day 40 before being considered cured. In this model,
untreated animals generally were moribund and were euthanized by days 3 to 4.
Enzyme kinetics. T. brucei AdoMetDC-prozyme complex and human
AdoMetDC were expressed in Escherichia coli and copurified by Ni2?-agarose
and anion-exchange chromatography as previously described (7, 22, 32). Previous
studies indicated that the activity of the enzyme complex consisting of recombi-
nant His-tagged AdoMetDC enzyme and Flag-tagged recombinant prozyme was
equivalent to that of the native complex in lysates (32). AdoMetDC activity was
determined by trapping of labeled14CO2, also as previously described (7, 22, 32).
Kinetic analysis of the described inhibitors was conducted by the methods of Kitz
and Wilson, which describes analysis of time-dependent irreversible inhibition
(23). Enzyme (0.1 ?M) was preincubated with inhibitor at various concentrations
(0.1, 0.3 0.6, and 1.0 ?M) in buffer (100 mM HEPES [pH 8.0], 50 mM NaCl,1
mM dithiothreitol) at 37°C over a time range of 1 to 22 min. Aliquots were
removed at various time points and diluted 10-fold into assay mix containing 1
mM AdoMetDC, and the activity remaining (vi) relative to that of the no-
inhibitor control at the same time point (vo) was determined in a 10-min assay.
Data were fitted to equation 1 to determine the observed rate constant (kobs).
The first-order rate constant of inactivation (kinact) and the apparent Ki(Kiapp)
were determined by fitting kobsto equation 2. Data are reported as kinact/Kiapp
ratios, which represent the apparent second-order rate constants defining the
efficiency of enzyme inactivation by the inhibitor.
kobs? kinact?I?/?Kiapp ? ?I??
where e??kobst?is the negative logarithm of the apparent first-order rate constant
(kobs) at time t and [I] is the inhibitor concentration.
In vitro pharmaceutical properties. Solubility was determined using a kinetic
method. Briefly, the compound was dissolved in dimethyl sulfoxide, was diluted
to 0.5% in phosphate-buffered saline, pH 7.4 (PBS), was allowed to equilibrate
for 16 to 24 h, and then was filtered using a 1.2-?m filter, and the concentration
was measured using a 96-well plate UV spectrophotometer.
Passive permeability was measured using the parallel artificial membrane
permeation assay (PAMPA). Compound was added to the donor well (pH 6.5 in
PBS), which was separated from the acceptor well (pH 7.4 in PBS) by a phos-
pholipid membrane in dodecane. Samples were incubated 4 h, and then the
concentrations were measured in donor and acceptor wells to calculate perme-
ability (1, 21).
Metabolic stability was determined using both rat and human liver micro-
somes. Compound was incubated with rat or human liver microsomes at 0.5
mg/ml protein and NADPH cofactors. Postincubation at 0, 10, 20, 30, and 45
min, samples were withdrawn for liquid chromatography-tandem mass spectrom-
etry (LC–MS-MS) analysis. Half-life (t1/2) was determined by plotting ln(peak
area ratio) versus time (min). The intrinsic clearance was calculated based on the
well-stirred model (26).
The logarithm of the concentration ratio was calculated by measuring the
partitioning of compound between octanol and PBS (pH 7.4) after shaking and
Plasma protein binding was calculated by diluting a 25 mM stock solution in
DMSO to 5 ?M in plasma, dialyzing for 4 h against PBS (pH 7.4) at 37°C, and
determining the concentration remaining in each compartment.
Cytochrome P450 (CYP) inhibition was determined for human CYP isozymes
1A2, 2D6, 3A4, 2C9, and 2C19 by measuring the inhibition of each isozyme’s
ability to process its specific fluorescein-labeled substrate. This assay tested
inhibition at a single concentration (5 ?M) (15, 29).
In vivo PKs. Groups of six animals (split into two subgroups to minimize the
number/volume of blood collections) were administered doses either i.v. or by
mouth (p.o.) ranging from 10 mg/kg to 50 mg/kg. Blood samples were drawn at
15 min, 30 min, 2 h, 4 h, 8 h, and 24 h and were subjected to LC–MS-MS to
determine drug levels.
Brain penetration was determined by harvesting brains at selected time points
VOL. 53, 2009AdoMetDC INHIBITORS FOR TREATING HAT2053
and by weighing and extracting total tissue homogenate for LC–MS-MS analysis.
The amount of test compound associated with the brain was calculated by
subtracting an amount assumed to be attributable to blood vessels (3%, times the
blood exposure levels).
MDL 73811 and Genz-644131 are highly active against T. b.
rhodesiense and purified T. b. brucei AdoMetDC-prozyme en-
zyme complex in vitro. MDL 73811 and its analogs were tested
for their ability to kill parasites in vitro and for activity against
the purified AdoMetDC prozyme-enzyme complex. Results
(Table 1) show that MDL 73811 is highly active in vitro against
the parasite (50% inhibitory concentration [IC50], 0.004 ?g/ml;
0.0014 ?M) and also against the T. brucei AdoMetDC het-
erodimer (kinact/Kiapp ? 1.5 ?M?1min?1). Some analogs
(Genz-644390, Genz-644043, Genz-644053) showed a ?100-
fold loss of activity against the parasite. However, one analog
(Genz-644131) showed 10-fold-greater potency against the
parasite (IC50? 400 pg/ml; 0.0001 ?M) and ?5-fold activity
against the purified T. brucei AdoMetDC heterodimer
AdoMetDC (kinact/Kiapp ? 7.8 M?1min?1). Similar kinetic
constants of inactivation were obtained for the human enzyme
(Table 1). The efficiency of inactivation measured for Genz-
644043 is likely to be an underestimate, because the enzyme
activity was nearly completely depleted by the first measurable
time point (2.5 min) for all concentrations of inhibitor above
0.1 ?M. Additionally, the results were affected by stoichiomet-
ric inhibition, because the enzyme concentration ([E]) re-
quired to obtain reliable data was similar to the inhibitor con-
centration ([E] ? 0.1 ?M). It was therefore not possible to test
lower inhibitor concentrations.
ADME profiles of MDL 73811 and its analogs are generally
favorable. Results from in vitro absorption, distribution, me-
tabolism, and excretion (ADME) profiling studies are shown in
Table 2. The aqueous solubility of all the analogs was ?47
?g/ml. Compound stability was examined in a number of assays
because previous work suggested rapid clearance of MDL
73811 from the blood of rodents (11). Rates of clearance by
microsomes and hepatocytes were low, and CYP inhibition was
minimal. Plasma protein binding (41%) by the most active
analog, Genz-644131, was approximately half that of MDL
73811 in either human (72.9%) or mouse (69.8%) plasma. The
TABLE 1. Activities against parasites and purified AdoMetDC
IC50for T. b.
kinact/Kiapp (?M?1? min?1) againsta:
MDL 738110.004; 0.0014 1.501.59
0.489; 0.172 0.19ND
aND, not determined.
TABLE 2. In vitro ADME profiling
Stability clearance (ml/min/kg)
IC50(?M) for human CYP isozyme:
% of protein
bound in plasma
RatHumanRatHuman3A42D62C9 2C19 1A2 HumanMouse
2054BARKER ET AL.ANTIMICROB. AGENTS CHEMOTHER.
only poor in vitro ADME characteristic of these compounds
was their membrane permeability, which was low.
Additional in vitro studies were done to characterize MDL
73811 and to create a benchmark against which to compare
future analogs. MDL 73811 was highly stable in rat and human
plasma, with no degradation over a 4-h period (100% re-
mained). Similarly, MDL 73811 was highly stable in rat whole
blood (100% remaining) and showed low permeability, no ev-
idence of efflux in CaCo-2 cells ([3.25 ? 0.2] ? 10?6and [2.7 ?
0.2] ? 10?6cm/s for apical (Ap) to basolateral surface (BL)
and for BL to Ap, respectively), and modest partitioning into
red blood cells (partitioning coefficient, 0.28). Because these
results strongly suggested that neither blood stability nor par-
titioning into red blood cells was responsible for clearance
from plasma, Genz-644131 was not tested for these parame-
PK parameters of Genz-644131 in rats. Two studies were
performed assessing PK parameters in rats (Table 3). Oral
bioavailability was 2%; the t1/2ranged from 3.2 to 7.0 h be-
tween the two studies. Only 4% was cleared in urine by 8 h
postadministration; by 24 h, 21% was cleared in urine and
0.2% in bile.
Genz-644131 shows a longer t1/2and brain penetration in
vivo in mice than MDL 73811. Selected PK parameters were
examined following single intraperitoneal administration at 50
mg/kg in mice. Results (Table 4) show that the t1/2of Genz-
644131 is approximately threefold longer than that of MDL
73811 (7.43 h versus 2.48 h). Similarly, brain penetration is also
?4.3-fold higher for Genz-644131. This is noteworthy because
of the importance of brain penetration for treating stage 2
Genz-644131 shows a longer t1/2and higher IC50levels in
the blood of mice over time than does MDL 73811. PK studies
were conducted with mice to establish the benchmark values
for MDL 73811. Animals were injected with either 50 mg/kg or
20 mg/kg MDL 73811, and levels in blood were measured over
8 h. Results show that by 8 h, remaining drug following a
50-mg/kg dose was still ?10-fold above the IC50after correc-
tion for protein binding (Fig. 1A). At 8 h, following a single
dose of 20-mg/kg MDL 73811, levels had fallen to approxi-
mately the IC50. Based upon these studies, two situations were
modeled. The first follows the hypothetical clearance of the
single original 50-mg/kg dose from 8 h through 24 h and pre-
dicts that levels of MDL 73811 will fall below the IC50by 12 h
(Fig. 1A). The second situation assumes that animals are dosed
two more times (at 8 and 16 h) with 20-mg/kg doses. Under this
scenario, blood levels are expected to remain ?10-fold above
the IC50throughout the 24-h period.
A similar, single-dose study was done with Genz-644131
administered at 50 mg/kg (Fig. 1B). Because of the increased
potency, lower protein binding and longer t1/2, plasma levels of
Genz-644131 were expected to remain ?3 orders higher than
the IC50corrected for plasma protein binding of the com-
pound. Based on these observations, a model showing the
elimination profile of a 5-mg/kg dose was developed (Fig. 1B).
Even at this dose, blood levels of Genz-644131 were expected
to remain ?10-fold above the IC50for 24 h.
Both MDL 73811 and Genz-644131 significantly increase
the survival of T. b. brucei-infected mice. Mice were infected on
day 0 with T. b. brucei strain STIB 795, and treatment was
administered to groups of four animals at 50 mg/kg once per
day for 4 days starting on day 3. Animals were then monitored
for survival (daily) and for parasitemia twice/week through day
30. Results (Table 5) show that the mean survival time for
untreated control animals was 6.75 days but that treated ani-
mals survived though day 30 (when the study was terminated
and animals were euthanized). By day 26, one animal in the
MDL 73811-treated group was deemed cured, and no parasites
were found upon examination of blood smears. This animal
remained free from parasitemia through day 30. In the Genz-
644131-treated group, there was no detectible parasitemia in
any of the animals through day 20, though by day 30, parasites
were again found in three out of four animals.
Additionally, we studied the effect of increased dose fre-
quency and duration. Results (Table 6) show that treatment
with 50 mg/kg/day either BID for 4 days or QD for 7 days
resulted in sterile cure (no parasites through day 40) of the T.
b. brucei LAB 110 EATRO strain.
Despite the obvious unmet medical need, progress in devel-
oping new drugs for HAT has been very slow. Studies initiated
in the late 1970s and early 1980s demonstrated that the poly-
TABLE 4. Selected systemic PK parameters and brain penetration
following single-dose i.p administration in mice
(?g ? h/ml)
(?g ? h/ml)
TABLE 3. PK parameters of Genz-644131 in rats
Mean value (SD)a
% of dose
(collection interval, h)
(ng ? h/ml)
3681 (290) 3.17 (0.02)
853 (632) 81 (35)Urine
0.20 (0–24)100.25 (0) 39 (5)1.53 (1.11)2.1 (0.60)
2i.v.31,550 (696) 7.03 (1.01) 2,830 (2,120)38 (19)Urine21.1 (0–24)
aTmax, time to maximum concentration of drug in serum; Vss, volume of distribution at steady state; CL, clearance; F, percent oral bioavailability.
VOL. 53, 2009AdoMetDC INHIBITORS FOR TREATING HAT2055
amine biosynthetic pathway was essential in trypanosomes and
that inhibitors could be potential drug candidates (2, 5, 14).
DFMO, which inhibits ornithine decarboxylase, has become
widely used in treating HAT yet has issues of administration
requiring large amounts of drug to be infused i.v. over a 2-week
period (10, 20).
Irreversible inhibitors of AdoMetDC, another enzyme in the
polyamine biosynthetic pathway, have also shown considerable
promise (6, 8, 11). However, drug characteristics were not
optimal; MDL 73811 is not curative against the CNS stage of
infection when used as monotherapy (6), and studies examin-
ing levels in the blood of rats appeared to indicate that the
compound is cleared with a t1/2of 10 to 20 min (11). We
therefore set out to characterize the ADME properties of
MDL 73811 to provide a benchmark against which to compare
subsequent analogs that hopefully will begin to address some
of these issues.
MDL 73811 (32) and four analogs were synthesized. Com-
pounds were tested for efficacy against trypanosomes in vitro,
and two were tested against the purified T. brucei AdoMetDC-
prozyme heterodimer (Table 1). All halogenated compounds
were ?100-fold less active than MDL 73811 against trypano-
somes. The activities of two of these analogs against the puri-
fied enzyme were also lower than that of MDL 73811, though
it was not reduced by the same proportion. While the reasons
for this are uncertain, the uptake of MDL 73811 by trypano-
somes is mediated by purine transporters (12, 18), and the
halogenated compounds may disrupt the electron distribution
required by the transporter. In contrast, the activity of Genz-
644131 against both T. brucei blood form parasites in whole-
cell assays and the purified AdoMetDC prozyme-enzyme het-
erodimer wasincreased (10-
compared with that of MDL 73811.
In vitro, these compounds were found to be stable in rat and
human liver microsomes and hepatocyte stability tests, did not
inhibit human CYP enzymes, and had relatively low plasma
protein binding (Table 2). Further studies examined com-
pound stability in rat and human plasma as well as whole blood
from rats (see above). Over a 4-h period, there was no appar-
ent change in compound structure, nor was there significant
uptake into red blood cells. Additional studies examined
whether lung endothelial cells (human microvascular lung en-
dothelial cells) or kidney cells (renal proximal tubule epithelial
cells) altered the stability of these AdoMetDC inhibitors.
Again, there was no apparent change in the chemical structure
of the compound after a 4-h exposure (data not shown). In vivo
PK studies showed that by 24 h, ?21% of Genz-644131 was
cleared in urine while only 0.2% was cleared in bile (Table 3).
Because MDL 73811 and its analogs appear stable in these in
vitro assays and the proportion excreted is low, the clearance
mechanism(s) remains unknown.
PK studies of mice examined the clearance of compound
from plasma and penetration into the brain (Table 4). While
and 5-fold, respectively)
TABLE 6. Antiparasitic activity in T. b. brucei strain LAB 110
Treatment groupDose (days)
2 mg/kg QD (1–4)
50 mg/kg/day BID (1–4)
50 mg/kg/day BID (1–7)
50 mg/kg/day QD (1–7)
50 mg/kg/day QD (1–7)
FIG. 1. Measured and predicted levels of MDL 73811 and Genz-644131 in the blood of mice. (A) Mice were injected i.p. with MDL 73811
either at 50 mg/kg (open circles) or 20 mg/kg (open triangles); samples were collected and analyzed over an 8-h period. Based upon those values,
the predicted exposure level was modeled. Filled circles indicated predicted levels following a single injection of 50 mg/kg, plotted from 8 to 24 h.
Filled triangles indicated predicted blood levels modeled on thrice daily injections over a 24-h period. The dashed line indicates the calculated IC50
level, corrected for protein binding. (B) Mice were injected once with 50 mg/kg Genz-644131, and blood levels were sampled over 8 h (open circles).
Predicted values extending clearance of the 50-mg/kg dose through 24 h are modeled (open triangles). Filled triangles indicate predicted values
for a hypothetical dose of 5 mg/kg, based upon results with 50 mg/kg. The dashed line indicates the calculated IC50level, corrected for protein
TABLE 5. Antiparasitic activity in T. b. brucei strain STIB
No. cured/no. infected on dayb:
aThe dose of both MDL 73811 and Genz-644131 was 50 mg/kg i.p. on days 3
bND, not done.
2056 BARKER ET AL.ANTIMICROB. AGENTS CHEMOTHER.