Effect of low-dose ritonavir on the pharmacokinetics of the CXCR4 antagonist AMD070 in healthy volunteers.
ABSTRACT AMD070, a CXCR4 antagonist, has demonstrated antiretroviral activity in human immunodeficiency virus-infected patients. Since AMD070 is a substrate of cytochrome P450 3A4 and P-glycoprotein, both of which may be affected by ritonavir, we tested for a ritonavir effect on AMD070 pharmacokinetics. Subjects were given a single 200-mg dose of AMD070 on days 1, 3, and 17. Ritonavir (100 mg every 12 h) was dosed from day 3 to day 18. Blood samples to test for AMD070 concentrations were collected over 48 h after each administration of AMD070. Twenty-three male subjects were recruited. Among them, 21 completed the study, and 2 were discontinued for reasons other than safety. All adverse events were grade 2 or lower. AMD070 alone had the following pharmacokinetic features, given as medians (ranges): 3 h (0.5 to 4 h) for the time to peak blood concentration, 256 ng/ml (41 to 845 ng/ml) for the peak concentration (C(max)), 934 h x ng/ml (313 to 2,127 h x ng/ml) for the area under the concentration-time curve from 0 h to infinity (AUC(0-infinity)), 214 liters/h (94 to 639 liters/h) for apparent body clearance, and 4,201 liters (1,996 to 9,991 liters) for the apparent volume of distribution based on the terminal phase. The initial doses of ritonavir increased the C(max) of AMD070 [geometric mean (90% confidence interval)] by 39% (3 to 89%) and the AUC(0-infinity) by 60% (29 to 100%). After 14 days of ritonavir dosing, the pharmacokinetic changes in AMD070 persisted. The plasma pharmacokinetics of ritonavir were consistent with previous reports. It is concluded that AMD070 concentrations were increased with concomitant ritonavir dosing for 14 days in healthy volunteers.
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ABSTRACT: The chemokine CXCR4 receptor is a G-protein-coupled receptor that has multiple functions in normal physiologies involving the hematopoietic and immune systems. Although the CXCR4 receptor was initially discovered as one of the co-receptors involved in human immune deficiency virus (HIV) cell entry, it has also been linked to many central immune system functions through the direct regulation of cell trafficking and adhesion, and is present in many cell types within the body. The receptor ligand combination of CXCL12 (SDF-1) and CXCR4 underlies pathologies such as HIV infection, cancer metastasis and drug resistance, leukemia progression, rheumatoid arthritis, and lupus. This alternative and widespread functionality has allowed researchers to discover new potential uses for substances that are CXCR4 antagonists. This has resulted in approval of the first CXCR4 antagonist, plerixafor injection (AMD3100), for hematopoietic stem cell mobilization. Newer agents are under clinical evaluation for HIV, cancer, and stem cell mobilization. As these agents are further investigated, new uses and therapeutic interventions for CXCR4 antagonists will be discovered. Drug Dev Res 72:598–602, 2011. © 2011 Wiley Periodicals, Inc.Drug Development Research 11/2011; 72(7). · 0.87 Impact Factor
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ABSTRACT: 1. Recently, the Food and Drug Administration (FDA) and European Medicines Agency have shown decision trees to determine whether a drug candidate is an inhibitor of P-glycoprotein (P-gp). However, there has been no clear information on whether P-gp inhibition can be significant in clinical drug–drug interactions (DDIs). The purpose of this study was to confirm the effect of P-gp inhibition through comprehensive analysis of the clinical DDI studies. 2. Clinical information on P-gp inhibition was collected using the University of Washington Metabolism and Transport Drug Interaction Database™. The risks of P-gp inhibition-related DDI were qualitatively evaluated in terms of the contribution of CYP3A inhibition. The degrees of DDI risk were categorized using the area under the plasma concentration–time curve increase ratio (AUCR), according to the FDA DDI criteria. 3. When both P-gp and CYP3A were inhibited, the DDI risks were potent in 25% of the studies. When CYP3A inhibition did not contribute to the DDI, no study was categorized as potent DDI risk, and the detailed analysis revealed that AUCRs were basically <3.0. The DDI risk caused by P-gp inhibition solely would be limited, although the use of P-gp substrates with narrow therapeutic range should be carefully controlled.Xenobiotica 06/2014; · 2.10 Impact Factor
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ABSTRACT: A de novo hit-to-lead effort involving the redesign of benzimidazole-containing antagonists of the CXCR4 receptor resulted in the discovery of a novel series of 1,2,3,4-tetrahydroisoquinoline (TIQ) analogues. In general, this series of compounds show good potencies (3-650 nM) in assays involving CXCR4 function, including both inhibition of attachment of X4 HIV-1IIIB virus in MAGI-CCR5/CXCR4 cells and inhibition of calcium release in Chem-1 cells. Series profiling permitted the identification of TIQ-(R)-stereoisomer 15 as a potent and selective CXCR4 antagonist lead candidate with a promising in vitro profile. The drug-like properties of 15 were determined in ADME in vitro studies, revealing low metabolic liability potential. Further in vivo evaluations included pharmacokinetic experiments in rats and mice, where 15 was shown to have oral bioavailability (F = 63%) and resulted in the mobilization of white blood cells (WBCs) in a dose-dependent manner.ACS Medicinal Chemistry Letters 11/2013; 4(11):1025-30. · 3.07 Impact Factor
ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, May 2008, p. 1630–1634
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
Vol. 52, No. 5
Effect of Low-Dose Ritonavir on the Pharmacokinetics of the CXCR4
Antagonist AMD070 in Healthy Volunteers?†
Ying Jun Cao,1* Charles W. Flexner,1Shelia Dunaway,2Jeong-Gun Park,3Karin Klingman,4
Ilene Wiggins,1Jeanne Conley,2Christine Radebaugh,1Angela D. Kashuba,5
Ron MacFarland,6Stephen Becker,6and Craig W. Hendrix1
Johns Hopkins University School of Medicine, Baltimore, Maryland1; University of Washington and Harborview Medical Center,
Seattle, Washington2; Frontier Science and Technology Research Foundation/Harvard School of Public Health, Boston,
Massachusetts3; Division of AIDS, NIAID/NIH, Bethesda, Maryland4; University of North Carolina at Chapel Hill,
Chapel Hill, North Carolina5; and AnorMED, Inc., Langley, British Columbia, Canada6
Received 11 November 2007/Returned for modification 14 January 2008/Accepted 11 February 2008
AMD070, a CXCR4 antagonist, has demonstrated antiretroviral activity in human immunodeficiency virus-
infected patients. Since AMD070 is a substrate of cytochrome P450 3A4 and P-glycoprotein, both of which may
be affected by ritonavir, we tested for a ritonavir effect on AMD070 pharmacokinetics. Subjects were given a
single 200-mg dose of AMD070 on days 1, 3, and 17. Ritonavir (100 mg every 12 h) was dosed from day 3 to
day 18. Blood samples to test for AMD070 concentrations were collected over 48 h after each administration
of AMD070. Twenty-three male subjects were recruited. Among them, 21 completed the study, and 2 were
discontinued for reasons other than safety. All adverse events were grade 2 or lower. AMD070 alone had the
following pharmacokinetic features, given as medians (ranges): 3 h (0.5 to 4 h) for the time to peak blood
concentration, 256 ng/ml (41 to 845 ng/ml) for the peak concentration (Cmax), 934 h ? ng/ml (313 to 2,127
h ? ng/ml) for the area under the concentration-time curve from 0 h to infinity (AUC0–?), 214 liters/h (94 to 639
liters/h) for apparent body clearance, and 4,201 liters (1,996 to 9,991 liters) for the apparent volume of
distribution based on the terminal phase. The initial doses of ritonavir increased the Cmaxof AMD070
[geometric mean (90% confidence interval)] by 39% (3 to 89%) and the AUC0–?by 60% (29 to 100%). After 14
days of ritonavir dosing, the pharmacokinetic changes in AMD070 persisted. The plasma pharmacokinetics of
ritonavir were consistent with previous reports. It is concluded that AMD070 concentrations were increased
with concomitant ritonavir dosing for 14 days in healthy volunteers.
Despite the availability of many effective antiretroviral
drugs, treatment options are still challenged by drug toxicities
and the emergence of drug-resistant human immunodeficiency
virus (HIV). The identification of a new class of drugs with a
novel mechanism of action, durable efficacy, and lack of cross-
resistance with existing antiretroviral drug classes remains a
continuing therapeutic need. AMD070 belongs to a novel class
of antiretroviral entry inhibitors (AnorMED, Inc., data on file).
It is a specific and reversible antagonist of the CXCR4 che-
mokine receptor, the coreceptor used by T-cell tropic (X4),
syncytium-inducing HIVs, and has a potent and selective abil-
ity in vitro to inhibit X4 viral replication by blocking fusion and
viral entry into the cell. The investigational CXCR4 antagonist
AMD3100 given intravenously has been shown to reduce the
X4 viral load in HIV-infected subjects; however, its develop-
ment for HIV infection was terminated due to poor oral bio-
availability and adverse effects (7, 8). AMD3100 is currently in
phase III clinical trials as an intravenous adjunct for stem cell
mobilization in malignant diseases. In contrast, orally admin-
istered AMD070 is well absorbed and well tolerated and dis-
plays a terminal elimination half-life of 11 to 16 h (19). A
majority of the subjects dosed with 200 mg twice daily attained
plasma concentrations at or near the in vitro 90% effective
concentration 24 h after dosing. The combined results of two
related 10-day phase II studies of HIV-infected subjects re-
ceiving 200 mg AMD070 twice daily demonstrated an antiviral
effect, defined as a 1-log reduction in X4 relative luminescence
units in the profile assay, in 7 of 15 subjects (13, 17). Further-
more, like its predecessor, AMD3100, AMD070 showed a dose
response for leukocytosis which may serve as a surrogate
marker for CXCR4 inhibition and a second potential indica-
tion for use of the drug.
AMD070 is cleared primarily by metabolism, with ?1% of
the oral dose appearing unchanged in the urine (AnorMED,
Inc., data on file). In vitro studies using human liver micro-
somes have shown that AMD070 is metabolized by cytochrome
P450 3A4 (CYP3A4) and, to a lesser extent, CYP2D6 en-
zymes; it is a substrate of P-glycoprotein (P-gp) (AnorMED,
Inc., data on file). CYP3A4 is a monooxygenase encoded by
the CYP3A gene. Many CYP3A enzyme substrates can also be
metabolized by other CYP enzymes. Ritonavir is a substrate
for multiple CYP enzymes, including CYP3A4 and CYP2D6.
It is also an inhibitor and inducer of several CYP enzymes,
such as CYP3A4 and CYP2C8. Ritonavir is commonly given
with other protease inhibitors to “boost” their plasma concen-
trations through hepatic or enterocytic CYP enzyme inhibition
(4). Additionally, ritonavir inhibits P-gp, which enhances the
absorption and alters the distribution of substrate drugs (3, 21).
In rat studies, the coadministration of ritonavir increased the
area under the concentration-time curve (AUC), the peak
* Corresponding author. Mailing address: Osler 501, 600 N. Wolfe
Street, Baltimore, MD 21287. Phone: (410) 502-3252. Fax: (410) 614-
9978. E-mail: firstname.lastname@example.org.
† AIDS Clinical Trial Group study A5191.
?Published ahead of print on 19 February 2008.
concentration in blood plasma (Cmax), and the oral bioavail-
ability (F) of AMD070 by 58%, 158%, and 66%, respectively.
In dog studies, there was no significant increase in the AUC for
AMD070 when it was coadministered with either a single dose
of ritonavir or at ritonavir steady state. In vitro and clinical
studies have also shown that AMD070 is a weak inhibitor of
the CYP2D6 and CYP3A4 enzymes (AnorMED, Inc., data on
Because of the potential for an AMD070-ritonavir interaction
based on in vitro studies, we performed this pharmacokinetic
drug interaction study to assess the interactions of AMD070 with
ritonavir given both as a single dose and twice a day for 2 weeks.
We hypothesized that the coadministration of a low dose of
ritonavir would increase the oral bioavailability of AMD070.
(The results of this study were presented previously at the
Conference on Retroviruses and Opportunistic Infections, Los
Angeles, CA, 2007.)
MATERIALS AND METHODS
Study design. This was a phase I, open-label, multiple-dose, sequential-design
drug interaction study that consisted of one cohort of healthy male volunteers.
Volunteers were dosed initially with 200 mg AMD070 alone on day 1. On days
1 to 3, blood samples were taken for analysis of the AMD070 concentrations.
Volunteers did not receive any drug on day 2 to allow for the clearance of nearly
all (estimated to be 93%) of the first AMD070 dose. On day 3, volunteers began
taking ritonavir (100 mg twice daily) and continued through day 18 (for a total of
16 consecutive days and 32 total doses). A single 200-mg dose of AMD070 was
also given simultaneously with the morning ritonavir dose on day 3. On days 3 to
5, frequent blood samples were taken for pharmacokinetic analysis of the
AMD070 concentrations. Blood was collected to measure ritonavir concentra-
tions within 6 h after dosing on day 10 or 11 to indirectly assess the adherence to
the ritonavir dosing regimen. On day 17, volunteers received another 200-mg
dose of AMD070 simultaneously with their morning ritonavir dose. The evening
ritonavir dose on day 18 was the last dose. On days 17 to 19, blood samples were
again taken for analysis of the AMD070 concentrations. The safety of taking
AMD070 alone and during coadministration with ritonavir was assessed through-
out the study period.
The study was conducted at two research sites, Johns Hopkins University and
the University of Washington. Subjects were healthy men with no active medical
illness, as determined by history, physical, or laboratory evaluation. Uninfected
women were not enrolled due to the lack of reproductive toxicity data from
preclinical studies. The inclusion criteria, by which each subject was evaluated
within 28 days prior to commencing the study drug, included the following. (i)
The subjects were healthy males age 18 to 55 years (inclusive). (ii) If participating
in sexual activity that could lead to pregnancy, the subjects agreed that two
reliable methods of contraception would be used simultaneously while partici-
pating in the study and for 2 weeks after stopping the medication. (iii) The
subjects had acceptable laboratory values, including an HIV-seronegative result
determined by any licensed enzyme-linked immunosorbent assay; a white blood
cell count within normal limits; a hemoglobin level of ?12.5 g/dl; a platelet count
of ?150,000 cells/mm3; pertussis toxin level, partial thromboplastin time, and
total bilirubin level within normal limits; aspartate transaminase (serum glutamic
oxaloacetic transaminase) and alanine aminotransferase (serum glutamic pyruvic
transaminase) levels of ?1.3? the upper limit of normal; and calcium and
magnesium levels within normal limits. (iv) The subjects had electrocardiograms
deemed normal or with abnormalities not considered clinically significant by the
site investigators. (v) The subjects had body weights within 33% of the ideal
weight for their height based on the Metropolitan Life table (20). (vi) The
subjects had a willingness to abstain from exercise for at least 24 h prior to study
entry. Subjects were excluded if they (i) had participated in a previous AMD070
study with any adverse event of grade 3 or higher; (ii) had used prescription
medications, herbal supplements, or aspirin within 7 days prior to study entry or
nonsteroidal anti-inflammatory drugs, over-the-counter medications, or other
supplements, including multivitamins, within 1 day prior to study entry; (iii) had
received any immunizations within 30 days prior to study entry or been treated
with radiation therapy or cytotoxic chemotherapeutic agents or immunomodu-
lating agents within 30 days prior to study entry; (iv) had experienced an active
infection or acute illness of any kind within 14 days prior to study entry; (v) had
experienced active drug or alcohol abuse or dependence; (vi) had a history of
gastrointestinal bleeding or ulcer or had chronic diarrhea (defined as ?3 stools/
day) for more than 4 weeks prior to study entry; (vii) had a history of cardiac
conduction abnormalities, cardiac arrhythmias, cardiomyopathy, repolarization
delay (rate-corrected QT interval, ?500 ms), or additional risk factors for tor-
sade de pointes (e.g., heart failure, hypokalemia); and (viii) had any medical or
psychological condition that, in the opinion of the site investigator, might inter-
fere with participation in the study or put the volunteer at undue risk. The
protocol was approved by the institutional review board at both research sites,
and all study participants provided written informed consent prior to enrollment.
After screening and enrollment, the subjects were admitted to the General
Clinical Research Center for 5 nights to determine the pharmacokinetics of the
first two doses of AMD070 (days 1 to 5) and 3 nights to determine the pharma-
cokinetics of the last AMD070 dose (days 17 to 19). In the evening (on day 0 and
day 16), subjects were admitted to the General Clinical Research Center and
concomitant medications and vital signs were obtained. A targeted physical
exam, including signs, symptoms, diagnosis, and body weight, was performed,
and an indwelling intravenous catheter was inserted before the standardized
breakfast on the second day. A single, 200-mg dose of AMD070 was given 30 min
after breakfast. Ritonavir was given on days 3 to 18 twice a day, 30 min after
breakfast and 30 min after dinner. Urine for a urine dipstick test with micro-
scopic exam was collected for 24 h following AMD070 dosing. Blood was col-
lected into heparinized tubes predose and at 0.5, 1, 1.5, 2, 3, 4, 8, 12, 24, 34, and
48 h following AMD070 dosing. Food and drink were given ad libitum 4 h after
AMD070 dosing. Breakfast, lunch, and dinner were served at approximately 8:00
a.m., 12:00 p.m., and 5:00 p.m. during hospitalization. A follow-up safety visit was
conducted approximately 2 weeks after discharge (on day 19) or upon premature
discontinuation of the study. A review of the subject’s medical history and a
targeted physical exam were performed daily during hospitalization and on either
day 10 or 11. Similarly to the screening visit, the final follow-up visit included a
history review, a targeted physical exam, and laboratory evaluations.
Blood samples were kept on ice and sent to the clinical laboratory for pro-
cessing within 1 h of collection. During processing, the sample was centrifuged at
2 to 8°C at 900 ? g for 10 min. The blood plasma was aliquoted and frozen at
?20°C. After the sample from the last volunteer had been collected, all samples
were shipped on dry ice to the BAS Northwest Laboratory (McMinnville, OR)
for the AMD070 assay.
AMD070 assay. Concentrations of AMD070 in plasma and urine were deter-
mined by high-performance liquid chromatography–mass spectrometry (HPLC-
MS) using validated methods. Validations included assessments of linearity;
within-run and between-run precision and accuracy; selectivity; long-term (fro-
zen), short-term (room temperature), autosampler, freeze-thaw, and processed
sample stability; extraction efficiency; carryover; the effect of dilution; and po-
tential over-the-counter and HIV drug interferences. The validated calibration
range was 0.5 to 500 ng/ml for both plasma and urine and was selected based on
linearity (R2? 0.95), accuracy, and precision criteria for the validation runs. The
acceptance criteria were a 15% (20% at the lower limit of quantification) devi-
ation for accuracy and a ?15% (?20% at the lower limit of quantification)
coefficient of variation (CV) for precision. Actual within-run precision and ac-
curacy for the validation runs ranged from 3.7 to 14% CV and ?11 to 15%
deviation. Between-run precision and accuracy ranged from 6.6 to 11% CV and
?5.2 to 12% deviation. The maximum validated dilution factor was 100-fold.
Sample runs were accepted or rejected based on the results obtained for quality
control samples included in each run: at least 67% (e.g., six out of nine) of the
quality control samples were required to be within ?15% of their respective
Standard calibrators, quality control samples, and study samples were pre-
pared and analyzed in identical manners. Briefly, samples (100 ?l) were heated
for 30 min at 57°C, followed by the addition of 50 ?l of a 0.5-mg/ml internal
standard solution (AMD025, a structural analog of AMD070) and 50 ?l of a 1 N
sodium hydroxide (NaOH) solution. After the samples were mixed briefly, 1.0 ml
of methyl tert-butyl ether (MTBE) was added, and the samples were vortexed (10
min) and then centrifuged (3 min, ?10,000 rpm). Samples were then frozen at
??60°C for approximately 60 min. The MTBE layer was decanted into a second
tube and evaporated to dryness in a water bath at 30°C under nitrogen. Following
its reconstitution in 200 ?l of 5% acetonitrile-95% water-0.1% trifluoroacetic
acid, the samples were analyzed by reversed-phase HPLC with tandem MS
detection (mobile phase, 7% acetonitrile-93% water-0.1% trifluoroacetic acid;
flow rate, 0.5 ml/min; run time, 4.5 min). Detection by tandem MS incorporated
an electrospray interface in the positive ion mode.
Ritonavir assay. The concentration of ritonavir was determined by the Phar-
macology Support Laboratory at the University of North Carolina at Chapel Hill
using an HPLC/UV method. Briefly, 200 ?l of plasma was combined with 50 ?l
VOL. 52, 2008RITONAVIR-AMD070 DRUG INTERACTION1631
of a 5-?g/ml midazolam internal standard working solution. Samples were sub-
jected to liquid-liquid extraction using MTBE, and the organic layer was evap-
orated to dryness under a gentle stream of nitrogen. Samples were reconstituted
in the mobile phase before injection onto an Agilent (Wilmington, DE) 1100
HPLC system. Samples were separated on a Zorbax C18analytical column (3.5
?m, 150 by 4.6 mm; Agilent, Wilmington, DE) with a Zorbax C18guard column
(3.5 ?m, 12.5 by 4.6 mm; Agilent) and facilitated via a gradient elution. Cali-
bration standard curves ranged from 25 to 10,000 ng/ml. Intraday and interday
CVs were less than 6% (16).
Pharmacokinetic analysis of AMD070. Noncompartmental analysis was per-
formed using WinNonlin professional software (version 5.0.1; Pharsight Corp.,
Cary, NC). The AUC, Cmax, time to maximum concentration (Tmax), apparent
total clearance (CL/F), and apparent volume of distribution based on the ter-
minal phase (Vz/F) were estimated. The AUC was calculated with the linear/log
trapezoidal method. CL/F was calculated as the quotient of the dose to the
AUC0–?, where the AUC0–?was the AUC0–48plus the quantity of C48divided
by ?z(AUC0–48is the AUC from 0 to 48 h post-AMD070 dosing, C48is the
AMD070 concentration at 48 h postdosing, and ?zis the slope of the linear
regression of the log concentration versus time for the terminal portion of the
log concentration-time curve). Vz/F was calculated by dividing the dose by
AUC0–?? ?z. The AMD070 concentration during the acute ritonavir phase was
corrected for any residual concentration from the AMD070-alone phase, assum-
ing exponential decay with time based on the C48during the AMD070-alone
phase. Only the AUC0–?was reported since the percentage of extrapolation from
the time of last measurable concentration to infinity is relatively small [the
medians (ranges) were 6% (2 to 10%) for the first AMD070 dose, 8% (4 to 17%)
for the second AMD070 dose, and 6% (3 to 24%) for the last AMD070 dose].
Pharmacokinetic analysis of ritonavir. Similar methods were used for ritona-
vir pharmacokinetic analysis as were used for AMD070 except that the summary
parameters were estimated only for 0 to 12 h postdosing since ritonavir was given
twice daily. CL/F for the initial ritonavir dose and Vz/F were not estimated due
to the relatively large Tmax, the limited number of concentration points during
the decay phase, and the still-high concentration at 12 h (C12) postdosing. For
the first dose of ritonavir, the concentrations of the samples collected within the
individual lag time were set to zero for the calculation of the AUC. CL/F for
steady-state ritonavir was estimated by dividing the dose by the AUC at steady
Statistical consideration. Based on the data from our previous study (19), we
determined that with a paired study design, we would need 19 subjects to detect
a 35% change in the AUC of AMD070 (80% power; 5% two-sided significance
level; logarithmically transformed data). Assuming that 25% of the volunteers
would be lost or not evaluable, we planned to enroll up to 26 volunteers. In order
to be evaluable, volunteers would have to have taken at least 80% of their
scheduled ritonavir doses, including the last three scheduled doses prior to the
last AMD070 dose.
Data are presented as either medians and ranges or geometric means and 95%
confidence intervals (CI), except for the ratios of pharmacokinetic parameters,
which were presented as geometric means and 90% CI computed with the Proc
Mixed program of SAS v9.1.3 (SAS Institute, Inc., Cary, NC).
Subjects. A total of 23 subjects were enrolled: 17 subjects at
Johns Hopkins University and 6 subjects at the University of
Washington. Twenty-one subjects completed the study and
were included in the final pharmacokinetic analysis; one sub-
ject discontinued in the first week due to consent withdrawal,
and one discontinued in the second week due to unwillingness
to come to the clinic. The ages and weights of the 21 subjects
included in the final pharmacokinetic analysis, given as medi-
ans (ranges), were 42 years (18 to 53 years) and 77.8 kg (55.2
to 101.2 kg), respectively. Ten subjects were white (non-His-
panic), and 13 subjects were black (non-Hispanic).
Safety. All subjects tolerated the study drugs well. Revers-
ible, grade 1 to grade 2 adverse events were seen in 7 subjects
(30%) on AMD070 alone (a 2-day period) and in 16 subjects
(70%) on ritonavir, sometimes in association with an AMD070
dose during a longer 16-day period. Among nine of the grade
2 adverse events, five were assigned this grade solely because
either ibuprofen or acetaminophen was prescribed. All adverse
events were resolved. There was no grade 3- or grade 4-related
adverse event or finding of hepatotoxicity. Since the study was
powered for potential ritonavir-induced changes in AMD070
pharmacokinetics, not for safety endpoints, further statistical
analysis of the frequency of adverse events was not performed.
Pharmacokinetics. The blood plasma pharmacokinetic pa-
rameters of a single 200-mg oral dose of AMD070 are listed in
Table 1. Ritonavir increased the concentration of AMD070 in
blood plasma (Fig. 1). The Cmaxand AUC0–?of AMD070
coadministered with the first dose of ritonavir were increased
[90% geometric mean (90% CI)] by 39% (3 to 89%) and 60%
(29 to 100%), respectively, compared to those with the admin-
TABLE 1. AMD070 pharmacokinetic parametersa
AUC0–?(h ? ng/ml)
CL/F (liters/h)Vz/F (liters)
aThe number of subjects who completed the study was 21.
bAMD070 was given on day 1 (no ritonavir treatment), day 3 (first dose of ritonavir), and day 17 (29th dose of ritonavir).
FIG. 1. Concentration-time profile of AMD070 in blood plasma.
The inset shows the full time course of measured AMD070 concen-
trations. Data are geometric means with 95% CIs.
1632 CAO ET AL.ANTIMICROB. AGENTS CHEMOTHER.
istration of AMD070 alone (Table 2). These changes remained
after ritonavir had been administered twice daily for 14 days
There was a lag of 1 h (0 to 4 h) [median (range)] for
ritonavir to reach a detectable blood plasma concentration
after the first dose of ritonavir. The Cmaxand AUC0–12follow-
ing the first dose of ritonavir, given as geometric means (95%
CI), were 184 ng/ml (103 to 329 ng/ml) and 1,209 ng ? h/ml
(565 to 2,586 ng ? h/ml), respectively. The Cmax, Cmin, and
AUC at steady state following the morning dose of ritonavir on
day 17 were 741 ng/ml (354 to 1,551 ng/ml), 242 ng/ml (133 to
440 ng/ml), and 5,014 ng ? h/ml (2,048 to 12,277 ng ? h/ml),
respectively. The trough ritonavir concentration appeared sta-
ble starting at the second dosing interval. The blood samples
for the indirect assessment of ritonavir regimen adherence at
day 10 or 11, after the first week of ritonavir dosing, were taken
between 1.3 and 5.3 h postdosing. The ritonavir concentration
in these samples, given as the geometric mean (95% CI), was
642 ng/ml (424 to 973 ng/ml), which was not different from
those obtained 1 week later when the subjects were dosed in
the hospital [geometric mean (95% CI), 617 ng/ml (525 to 725
ng/ml) for the postdosing time between 1 and 4 h and 616
ng/ml (535 to 711 ng/ml) for the postdosing time between 1 and
We have shown that low-dose ritonavir caused weak in-
creases in the Cmaxand AUC of oral AMD070, during both the
acute and steady-state phases of ritonavir treatment. In addi-
tion, the combination of single-dose AMD070 with low-dose
ritonavir in our study appeared safe, as the subjects tolerated
the study regimen well and no adverse events of grade 3 or 4
were reported. The exclusion of female subjects is a limitation
of this study in that the study results may not be generalizable
The common mechanisms contributing to the increase of the
AUC of oral drugs include increasing oral bioavailability (by
increasing either absorption or decreasing presystemic clear-
ance or both) and decreasing elimination (by inhibiting metab-
olism and/or excretion). The apparent decrease of Tmaxand
Vz/F and the increase of Cmaxin our study were consistent with
an increase in oral bioavailability. Thus, ritonavir may inhibit
P-gp (thus increasing AMD070 absorption) and intestinal
CYP3A4 (thus decreasing presystemic clearance). The inhibi-
tion of the hepatic CYP3A4 enzyme may also be involved.
Alteration of renal clearance is unlikely since less than 1% of
AMD070 is excreted unchanged in the urine (AnorMED, Inc.,
data on file).
In our study, ritonavir was given for 14 days at a low dose. In
a study with HIV-infected subjects (11), ritonavir (given as a
single agent) was administered in dosing regimens of 200 mg to
500 mg twice daily; the trough ritonavir concentration de-
creased from day 8 to day 16, whereas the AUC and Cmaxwere
relatively time invariant. One of the proposed mechanisms was
the induction of CYP3A4. Based on the model of the time
course, the authors concluded that the half-life of the potential
ritonavir-mediated CYP3A4 induction was ?3.5 days. Their
data show that the trough concentrations of ritonavir tended to
stabilize by the end of a 2-week dosing period. Apparently,
ritonavir in our study reached steady state and the ritonavir
pharmacokinetic parameters were similar to those reported by
Aarnoutse et al. (1). Since ritonavir induces CYP3A4 enzyme
activity, we had expected slightly higher CYP3A4 activity after
2 weeks of ritonavir treatment. Although the geometric mean
ratio of AMD070 Cmaxand AUC0–?with both acute (day 3)
and chronic (day 17) ritonavir dosing to that with no ritonavir
dosing showed the lack of bioequivalence of AMD070, it is still
not clear whether the impacts of acute and chronic ritonavir
treatment on AMD070 are different. An in vitro study has shown
that AMD070 weakly inhibits CYP2D6, CYP3A4, and CYP1A2
(AnorMED, Inc., data on file), which can metabolize ritonavir
and add to the complexity of the AMD070-ritonavir drug inter-
Single-dose rather than steady-state pharmacokinetics of
AMD070 were studied because dosing once or twice daily as
anticipated in future efficacy studies would yield an accumula-
tion less than or equal to two times the single-dose concentra-
tions only during the terminal elimination phase, with little or
no detectable difference in peak concentration and AUC (19).
Furthermore, concentrations achieved with single 200-mg
doses are near the range expected to be clinically relevant (19).
The AMD070 dose chosen (200 mg) was based on demon-
strated safety in the study of up to 400 mg twice daily (AIDS
Clinical Trials Group study A5191) (19).
AMD070 has modest antiretroviral activity in some patients
with dosing of 200 mg every 12 h (17); the doses used in the
antiretroviral studies are also below the concentrations associ-
ated with maximal leukocyte mobilization (19). The pharma-
cokinetic interaction we report here may provide a way to
increase concentrations of AMD070, and perhaps the antiviral
activity, without increasing the AMD070 dose, as occurs for
numerous other antiretroviral drugs, primarily protease inhib-
itors. However, after the clinical phase of this study was com-
pleted, the FDA placed AMD070 on clinical hold due to liver
histology changes observed in longer-term preclinical toxicity
experiments. Currently, AMD070 is still on clinical hold. How-
ever, it has been reported that disrupting the CXCR4 receptor-
mediated trafficking of hematopoietic progenitor cells pro-
duces a stem cell mobilization effect (5, 12, 15). Therefore, the
favorable ritonavir interaction may also have relevance in an-
other clinical setting, as AMD070 has leukocyte-mobilizing
activity similar to that of AMD3100, another CXCR4 inhibitor
which is in advanced clinical development as a stem-cell-mo-
bilizing drug (2, 18). AMD070, however, has the significant
advantage of being orally bioavailable (19).
In conclusion, the coadministration of ritonavir with AMD070
weakly increased the Cmaxand AUC of AMD070 in blood
plasma. The increased Cmaxand AUC of AMD070 due to con-
comitant ritonavir dosing may result in clinically beneficial effects.
TABLE 2. Comparison of AMD070 pharmacokinetic parameters
Geometric mean ratio (90% CI)
Day 3/day 1 Day 17/day 1Day 17/day 3
VOL. 52, 2008RITONAVIR-AMD070 DRUG INTERACTION1633
This work was supported in part by NIH General Clinical Research
Center grants M01RR000052 (Johns Hopkins University) and
M01RR00037 (University of Washington) and AIDS Clinical Trials
Unit grants U01AI027668 (Johns Hopkins University), U01AI27664
(University of Washington), and AI038855.
We recognize the sustained excellence of all members of the ACTG
A5191 study team and the essential contributions of our healthy vol-
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