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IN VITRO ANTI-HIV EFFICACY OF THE CHEMOKINE RECEPTOR 5 (CCR5) ANTAGONIST TBR-652 IN COMBINATION WITH FOUR OTHER CLASSES OF ANTIRETROVIRAL AGENTS
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Reported by Jules Levin
ICAAC Sept 2009 SF, CA
S. Palleja1, R. Ogden1, F. Hamy2, V. Vidal2, T. Klimkait2, D. Martin1, R. Driz1, J Sapirstein1 1Tobira Therapeutics, Inc.; 2InPheno AG
AUTHOR SUMMARY & DISCUSSION
The results of the combination profiling of TBR-652 with clinically used anti-HIV drugs are summarized in Table 3. Under the conditions of the deCIPhR assay, the results of the additive effects of TBR-652/TBR-652 were compared to the effects of TBR-652/Other agent.
AUTHOR CONCLUSION
TBR-652 demonstrated additivity or weak synergy in all the combinations tested (TBR-652 plus lopinavir, atazanavir, darunavir, tenofovir, etravirine, or raltegravir).
Importantly, no antagonism was noted with any combination.
There was no indication of cytotoxicity with any combination at the concentrations tested.
The results suggest that combination regimens containing TBR-652 and
one or more of these agents are possible and should be studied in HIV-infected
patients.
ABSTRACT
Background: TBR-652 is a promising once daily CCR5 antagonist in phase 2 clinical trials. Because effective treatment of HIV infection requires combinations of antiretrovirals, it is important to assess the potential for virologic antagonism and synergy between the various components of a regimen. To assess the potential for future effective combination therapy, the interaction of TBR-652 and other classes of anti-HIV agents were investigated in vitro.
Methods: TBR-652 was tested in combination with lopinavir (LPV), darunavir (DRV), atazanavir (ATV), tenofovir (TDF), etravirine (ETV), and raltegravir (RAL) using the infectious replicative assay deCIPhR. CCR5-tropic virus was produced from proviral DNA and exposed to 12 concentrations of TBR-652 (0-100 nM) and 8 concentrations of each of the other inhibitors for 4 days. The degree of HIV-1 inhibition was read optically and compared to TBR-652/TBR-652 pairings to determine cut-off values for synergy and antagonism. Each pairing was run in triplicate. Cytotoxicity (cell number, morphology) was also assessed.
Results: The average 50% inhibitory concentration (IC50) for TBR-652 was 0.99±0.19 (range 0.72-1.43) nM. Weak synergy was seen in combinations of TBR-652 with LPV (IC50 26.90-40.27 nM), ATV (IC50 57.66-78.89 nM), DRV (IC50 2.12-2.78 nM) and ETV (IC50 71.77-83.35 nM), mostly at sub-IC50 concentrations for both drugs in a pair. Additive effects were seen with TBR-652 plus TDF (IC50 10.66-11.13 nM) or RAL (IC50 29.66-103.72 nM). No antagonism and no cytotoxicity were observed with any 2-drug combination at any concentration tested.
Conclusions: TBR-652 demonstrated additivity or synergy with LPV, DRV, ATV, TDF, ETV, and RAL. Importantly, no antagonism was noted with any combination. These results suggest that combination regimens containing TBR-652 and one or more of these agents are possible and should be studied in HIV-infected patients.
BACKGROUND
Because effective treatment of HIV infection requires combinations of antiretroviral agents, it is important to assess the potential for pharmacokinetic interactions between the components of multidrug regimens. Synergistic effects may be desirable, but they may cause an increase in number or severity of adverse events. Antagonistic effects may reduce antiviral activity and require dosage adjustments. Additive antiviral effects are desired. In general, drugs with the least potential interaction are recommended for regimen construction.1 TBR-652 is a promising once-daily CCR5 antagonist in phase 2 clinical trials. In this study, the potential for drug-drug interactions with 6 drugs from 4 different classes, all in clinical use today, was tested.
OBJECTIVE
To assess in vitro the potential for future effective combination therapy with
TBR-652 plus other widely used anti-HIV agents from four other drug classes.
METHODS
TBR-652 was tested in two-drug combinations with lopinavir (Abbott Laboratories), atazanavir (Bristol-Myers Squibb), darunavir (Tibotec Therapeutics), tenofovir (Gilead Sciences), etravirine (Tibotec Therapeutics), or
raltegravir (Merck & Co., Inc.). TBR-652 was prepared by InPheno as a 20 mM
solution in 100% DMSO and 0.5% acetic acid. One-third log dilutions were
prepared by serial dilutions in phosphate buffer to a final dilution of 10 uL,
which was applied directly to 200 uL cell cultures. Other agents were obtained
from their respective pharmaceutical companies. CCR5-tropic provirus was
derived from provirus pNL4-3 in which Env region was replaced with that of
the CCR5-tropic clone AD-8.
Because of the wide range of IC50 values of the test agents, a 3-dimensional
combination methodology (Syn3RD) was used that crossed the two
concentration ranges of tested pair: 12 concentrations of TBR-652 (100, 46,
22, 10, 4.6, 2.2, 1, 0.46, 0.22, 0.1, 0.05, and 0 nM) and 8 concentrations of
each of the other inhibitors (Table 1). The concentration ranges were made
using third-log (2.154-fold) dilution, with the expected IC50 concentration
placed in the first one third of the range. Cultures were incubated for 4 days
(3 to 5 cycles of viral replication). The virus was assayed for fitness using the
proprietary system deCIPhR™ (InPheno AG).
Each pairing was run in triplicate, and the degree of HIV-1 inhibition was
read optically and compared to TBR-652/TBR-652 pairings to determine cutoff
values for synergy and antagonism. In the TBR-652/TBR-652 pairings, the
average combination factor was -0.64 with a calculated standard deviation
(SD) of 2.9 (Figure 1).
The average values ±3 times the SD was defined as the cut-off for an
additive effect. A combination factor above 8.1 was considered synergistic
and below -9.3 was considered antagonistic. Cell number and morphology
were assessed microscopically for all experiments.
Figure 1. 3-Dimensional Plot
of Combination Factor Values
of TBR-652/TBR-652
Averaged From 3
Independent Experiments
Note: yellow = additive effect; green = synergistic effect; red = antagonistic effect
RESULTS
The average 50% and 90% inhibitory concentrations (IC50 and IC90) for all agents are presented in Table 2. Weak synergy was seen in combinations of
TBR-652 with LPV, ATV, DRV, and ETV, mostly at sub-IC50 concentrations for
both drugs in a pair. Additive effect was seen with TBR-652 plus TDF or RAL.
Graphic summaries of the results are presented in Figure 2a-2f.
No antagonism and no cytotoxicity were observed with any 2-drug combination
at any concentration tested.
Figure 2. 3-Dimensional Plots of Combination Factor Values of TBR-652 Combined With Other Antiretroviral Agents, Each Averaged From 3 Independent Experiments
Note: yellow = additive effect; green = synergistic effect; red = antagonistic effect
a) TBR-652/Lopinavir
c) TBR-652/Darunavir
e) TBR-652/Etravirine
b) TBR-652/Atazanavir
d) TBR-652/Tenofovir
f) TBR-652/Raltegravir
REFERENCE
1. Back DJ. Perspective: Drug-Drug Interactions that Matter. Topics in HIV Medicine 14(2):2006.
http://www.iasusa.org/pub/topics/2006/issue2/88.pdf.
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