icon-folder.gif   Conference Reports for NATAP  
  12th Conference on Retroviruses and Opportunistic Infections (CROI)
Feb 22-25, 2005
Boston, MA
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Advancing towards the therapeutic use of chemokine receptor blockade (CCR5-Entry Inhibitors)
  Reported for NATAP by David Margolis, MD
University of Texas, Southwestern Medical Center Dallas; VA Dallas; ACTG Many of the major sessions from CROI are available for viewing as webcasts http://www.retroconference.org/2005/Pages/webcasts.htm. Don Mosier (abstr. 56) reviewed the preclinical development of drugs that block the attachment of HIV to cellular chemokine receptors. Chemokine receptors (CCRs) serve to regulate cellular responses by signaling in response to chemokines, the hormones of the immune system signals. However, in addition to binding a CD4 receptor to infect a cell, HIV also requires interaction with a CCR.
This link to the NATAP Dec 2004 "POSITIVE+INFORMATION" newsletter is a comprehensive review of new drug developments in HIV in 2004, including 12 pages on entry inhibitor drugs in development:
CCRs, particularly CCR5 as it is the CCR most-often used by HIV, are attractive drug targets for several reasons. There is a track record in the pharmaceutical industry of successful drug development of inhibitors of the G protein-coupled receptors, of which CCRs are a member. A proportion of the populations has a mutation of CCR5 (delta 32), which renders the receptor nonfunctional, is protective against HIV-1 transmission, but confers no physiologic abnormality.
Mosier made the point that even if CCR antagonists prove difficult to develop or use and therapeutic antivirals, they are likely to be useful to prevent HIV transmission. CCR5 antagonists have been shown to be effective microbicides, blocking sexual transmission of HIV, in model systems (Veazey, J. Exp. Med. 2003; Lederman, Science 2004). In the laboratory, CCR5 antagonist-resistant mutants are slow to develop. Another pathway to drug resistance might be switching of the virus from the use of one CCR to another, CCR5 to CXCR4, for example. This is unlikely as at least 6 mutations are required to accomplish this switch.
Dan Kuritzkes of Harvard Medical School (abstr. 57) followed Mosier with a report of the clinical information on such drugs to date. Inhibitors of CCR5 and of CXCR4 (an alternate CCR used by some viruses, most often those found in patients with late-stage HIV) have shown potent anti-HIV-1 activity in the laboratory. Kuritzkes discussed the issue of the need to therapeutically target both primary viral coreceptors (CCR5 and CXCR4). Although the predominant circulating HIV species in most patients through most of the course of HIV infection use CCR5, CXCR4-using viruses may exist as a minority population in many patients, or as a majority population in some. Use of a CCR5 inhibitor may select for amplification of the rare CXCR4 species present, unless these viruses are blocked by the simultaneous use of a CXCR4 inhibitor, or other effective HAART. Testing to identify how much of which kind of virus is circulating are available, but are costly and time-consuming. The pathway to drug development of CXCR4 inhibitors is particularly challenging, as these inhibitors have measurable effects on other cell populations, and the long-term effects of administration of these inhibitors is unknown.
Proof-of-concept studies in HIV-1-infected subjects have established the in vivo activity of three CCR5 inhibitors, and provide preliminary safety data for their long-term use. Schering D is an agent that can be given once a day when boosted by ritonavir. Short-term decline of up to 1.5 log in viral load were seen; expanded phase II studies are underway. UK 427,857 has been given bid in studies of up to 4 weeks, with declines of up to 1.5 log in viral load. It is metabolized by Cyp 3A4, and ritonavir boosting might be possible. CXCR4 virus was detected in 2 patients during UK 427,857 therapy. Expanded phase II studies are also underway. Finally,GW873140 is under development by Glaxo SmithKline. It too has been given bid in studies, with declines of up to 1.5 log in viral load observed. Dual tropic virus, able to use both CCR5 and CXCR4 was observed in one subject. Finally, AMD3100, an inhibitor of CXCR4 HIV, is also under study.
Craig Hendrix (abstr. 58) of Hopkins discussed the pharmacokinetics and pharmacodynamics for this class of drugs. How to correlate the blood level of a CCR inhibitor with its antiviral effect is not yet clear. At least for UK427,857 antiviral effect appears to correlate with total drug concentration AUC (area under the curve) rather than trough level as is typical for many other antivirals. For some CCR antagonists antiviral effect is seen well after the drug is cleared from the blood. Also, prolonged binding of drug to CCR is seen in some cases. How or whether these unique features of CCR blockers will affect their implementation is yet to be uncovered.
James Demarest (abstr. 77) presented data on prolonged occupancy of CCR5 by the Glaxo SmithKline inhibitor 873140. Flow cytometry analysis showed that this spirodiketopiperazine CCR5 antagonist occupied a median of 98% of CD4+ cell CCR5 receptors following 7-10 days of twice daily dosing in occupancy in 8 HIV- and 31 HIV+ subjects. Follow-up analysis up to day 13 suggested that the half-life of receptor occupancy was 127 hours post-last dose. Higher doses tended to result in longer duration of occupancy, up to a half life of 152 hours. However, occupancy cannot yet be linked to antiviral efficacy, and so these results must be interpreted cautiously.
Chris Petropoulos of ViroLogic (abstr. 59) then reported on resistance seen to this class of antivirals. He stated that HIV could develop resistance by: 1) switch to use another co-receptor (X4 ot R5, or R5 to X4), 2) develop the capacity to enter the cell by a non-receptor pathway, or 3) use the CCR despite the fact that it was occupied by the drug ("reaching around the inhibitor"). In laboratory experiments, this third, non-competitive mechanism of resistance was in fact observed. Laboratory passage experiments yielded HIV with 100% resistance to Schering C and UK 427,857, as well as the CD4 antibody TNX355 and enfuvirtide (T-20).
However, Mike Westby of Pfizer (abstr. 96) found differences in CCR5 binding of viruses that had evolved resistance to UK 427,857. Small molecule CCR5 antagonists bind to a similar region of the receptor, raising the possibility of generalized class-resistance. Westby tested UK-427,857 and structurally related inhibitor compounds against a panel of site-directed mutants and R5 viruses, including clinical isolates highly resistant to UK-427,857 (427res). They found that all CCR5 antagonists bind CCR5 in the same region, a pocket formed by the trans-membrane helices of the receptor and an extracellular loop (ECL2). However, while UK-427,856 congeners that contain a triazole moiety were inactive against 427res viruses, congeners that contained a different substitutent (imidazopiperidine) were active against 427res viruses. This raised the hope that resistance to one HIV co-receptor antagonist will not necessarily lead to drug-class resistance.
Preclinical Development of Chemokine Receptor Inhibitors
CROI abstract #56
Donald E Mosier
The Scripps Res Inst, La Jolla, CA, USA
Background: Chemokine receptors, particularly CCR5, are attractive targets for antiviral drug development because they are members of a large family of G protein-coupled receptors (GPCR) known to be sensitive to small molecule inhibitors, and because a null mutation of CCR5 (delta 32) is protective against HIV-1 transmission and confers no known disease susceptibility.
Methods: We and others have examined the activity of N-terminal modifications of RANTES and a number of small molecule inhibitors as antiviral agents and as modulators of CCR5 conformation or surface expression. Several of these agents have also been tested for in vivo activity in human xenograft models and in primate models of mucusal transmission. We have also analyzed envelope mutations associated with resistance to entry inhibitors and the process of coreceptor switching from CCR5 to CXCR4.
Results: One common finding has been that entry inhibitors show considerable variation in activity (IC50) with different HIV-1 isolates. This variation appears to reflect the considerable flexibility in the interaction between the viral envelope trimer and the exposed domains of the chemokine receptor. The plasticity of this binding event suggests that the efficacy of entry inhibitors may change over the course of infection, and that different virus isolates may be more or less prone to develop resistance. Generation of resistant mutants during selection by entry inhibitors has been rare, and resistance due to coreceptor switching even more rare. One explanation for these findings is that mutations involved in coreceptor switching decrease the entry fitness of the virus. Two entry inhibitors have shown activity in a SHIV mucosal transmission model, suggesting that they may be useful in preventing infection as well as treating established infection.
Conclusions: It will be important to understand the evolution of HIV envelope:coreceptor interactions to use entry inhibitors in the most productive manner.
Clinical Activity and Efficacy Trials of Chemokine Receptor Inhibitors
CROI abstract #57
Daniel Kuritzkes
Brigham and Women's Hosp, Harvard Med Sch, Boston, MA, USA
Chemokine receptor blockade provides an attractive target for inhibition of HIV-1 entry. Large- and small-molecule inhibitors of CCR5 and CXCR4 have shown potent anti-HIV-1 activity in vitro. Phase 1-2 studies in HIV-1-infected subjects have established proof-of-concept for the in vivo activity of these drugs, and provide preliminary safety data. Several of these candidates are now in phase 2/3 clinical trials. Although results to date have been encouraging, a number of important uncertainties remain to be clarified. A particular challenge is the occurrence of mixed infection with R5 and X4 virus in patients with advanced disease. Whether drugs that specifically inhibit only a portion of the virus population can contribute to an overall net reduction in plasma viremia is an important objective of ongoing studies. Another concern is whether emergence of X4 viruses will be accelerated by CCR5 inhibition, and if so, what the consequences will be on disease progression. In addition, the long-term safety of CCR5 or CXCR4 blockade remains to be established. Lastly, whether the chemokine receptor antagonists are best used as a component of initial treatment regimens or reserved for use in later regimens must be explored through treatment strategies trials.
Clinical Pharmacokinetics and Pharmacodynamics of Chemokine Inhibitors:
Implications for Rational Dosing
CROI abstract #58
Craig W Hendrix
Bloomberg Sch of Publ Hlth, John Hopkins Univ, Baltimore, MD, USA
Rational Dosing. Rational dosing of drugs is best informed by an understanding of both the pharmacokinetics of the drug (concentration-time relationship) as well as the pharmacodynamics of the drug (exposure-response relationship). Armed with an understanding of drug exposure levels which achieve desired efficacy and avoid undesired toxicity combined with the knowledge of how drug concentration changes over time, one can build a useful dose-exposure-response model. This model enhances the ability to rationally choose a dosing regimen that achieves the optimal balance of antiviral effect with minimized toxicity.
Traditional Antiretroviral Characteristics. During the development of currently marketed antiretroviral drugs, the pharmacodynamic relationship between antiretroviral effect and drug exposure (trough, peak, area under the concentration - time curve, inhibitory quotient) were usually discovered, if at all, well after the marketing of the drug. In some cases, especially for nucleoside analogs with complicated intracellular pharmacokinetics like zidovudine, this has led to the adoption of more rational dosing regimens with improved adherence and reduced toxicity without sacrificing efficacy. Additionally, where it has been determined, trough concentrations relative to viral sensitivity to the drug (inhibitory quotient, Cmin/IC50) are often the drug exposure variable that best correlates with antiviral effect.
Chemokine Inhibitor Differences. Chemokine inhibitors appear to differ from previously developed ARV classes in several important ways. In some cases their biologic effect is not best correlated with trough concentrations. For several, there exists a prolonged period of antiviral effect well after the drug is cleared from the blood. These findings suggest that dosing amount and frequency should not be based simply on drug half-life alone. While the mechanism for the apparent differences between chemokine inhibitors and other ARV classes remains unclear, our understanding of the relationship between drug exposure and biologic effect are emerging sufficiently early in clinical development that this PK-PD knowledge about chemokine inhibitors can inform useful dose-concentration-response models which enable a more rational choice of dosing regimens to be incorporated into definitive phase 3 clinical studies.
Prolonged Duration of CCR5 Occupancy by 873140 in HIV-negative and HIV-positive Subjects
CROI abstract #77
S Sparks, K Adkison, A Shachoy-Clark, S Piscitelli, and James Demarest*
GlaxoSmithKline, Research Triangle Park, NC, USA
Background: 873140 is a spirodiketopiperazine CCR5 antagonist that binds specifically to human CCR5 and demonstrates potent in vitro and in vivo anti-HIV activity. In vitro studies suggest prolonged CCR5 receptor occupancy by 873140 with an offset half-life of > 100 hours. Previous in vivo studies have shown that 873140 selectively inhibits monoclonal antibody binding to CCR5 and exhibits substantial (> 97%) CCR5 receptor occupancy in blood during repeated oral administration of 873140. Furthermore, 873140 shows sustained viral suppression for 24 to 48 hours after therapy discontinuation. The objective of the current study was to assess the duration of CCR5 receptor occupancy in HIV+ and HIV- subjects following repeat-dose administration of 873140.
Methods: Flow cytometric analysis was used to determine duration of in vivo blood CCR5 receptor occupancy in 8 HIV- and 31 HIV+ subjects administered various doses of 873140 for 7 or 10 days, respectively. In vivo CCR5 receptor occupancy was analyzed at baseline, last day of dosing, and over 7 to 14 days following multiple dose administration. The half-life (t1/2) for receptor occupancy decline, a measure of the duration of CCR5 receptor occupancy, was estimated by fitting a first-order exponential model to the resulting receptor occupancy versus time data.
Results: Median CD4+ cell CCR5 receptor occupancy following 7 days of 600 mg twice daily 873140 dosing in 8 HIV- subjects was 98%. Post-treatment analysis on days 8, 9, 11, and 13 suggested t1/2 receptor occupancy on CD4+ cells was 127 hours post-last dose. HIV+ subjects received 200 mg once daily, 200 mg twice daily, 400 mg once daily, or 600 mg twice daily over 10 days. Median CD4+ cell CCR5 receptor occupancy on day 10 was > 95% across doses. A cross-dose combined post-treatment analysis at days 11, 12, 15, 17, 19, and 24 estimated a t1/2 receptor occupancy on CD4+ cells at 122 hours post last dose. Higher doses tended to result in longer duration of occupancy (t1/2 receptor occupancy of 69 hours [95% CI 53 to 97 hours] for 200 mg once daily vs 152 hours [95% CI 107 to 264 hours] for 600 mg twice daily). The t1/2 receptor occupancy decline was comparable between total lymphocytes and CD4+ cells for both HIV- and HIV+ subjects.
Conclusions: Studies using CCR5-specific monoclonal antibodies demonstrate substantial and prolonged in vivo blood CCR5 receptor occupancy by 873140. At sample times post final dose, when plasma drug levels were undetectable, significant CCR5 receptor occupancy (> 50%) was observed for approximately 5 days. The prolonged CCR5 receptor occupancy suggests a potential mechanism for the sustained antiretroviral effect seen following 873140 administration in HIV+ subjects. Taken together, the data support further evaluation of 873140 in HIV-infected individuals.
Resistance to HIV Chemokine Receptor Antagonists
CROI abstract #59
Christos J Petropoulos*1, W Huang1, J Toma1, S Fransen1, S Bonhoeffer2, and J Whitcomb1
1ViroLogic, Inc, South San Francisco, CA, USA and 2ETH Zurich, Switzerland
HIV-1 entry inhibitors represent a diverse new class of antiretroviral agents. Virus entry is a multi-step process involving several virus envelope proteins (gp120SU, gp41TM) and host cell receptors (CD4, CCR5, CXCR4). The cascade of protein-protein interactions and conformational changes that mediate virus entry represent novel targets that are functionally distinct from conventional enzymatic targets. Consequently, resistance to entry inhibitors can differ significantly from that of protease and reverse transcriptase inhibitors, and may emerge via alternative mechanisms depending on the specific molecular interaction that is targeted. Viruses with reduced susceptibility to fusion inhibitors (enfuvirtide) display log-sigmoid inhibition curves that typically reach 100% inhibition at high drug concentrations, consistent with a competitive mechanism of inhibition and escape. Resistance to fusion inhibitors is best described by increases in the IC50. In contrast, viruses with reduced susceptibility to inhibitors that antagonize envelope-co-receptor interactions often exhibit incomplete inhibition even at elevated drug concentrations. The inability to inhibit 100% of virus replication at high drug concentrations is consistent with an allosteric mechanism of inhibition and escape. Viruses that develop resistance to co-receptor antagonists likely acquire the ability to bind and utilize receptor-inhibitor complexes. Consequently, resistance to allosteric co-receptor inhibitors is best described by the extent of incomplete inhibition observed at high drug concentrations.
Structurally-related HIV Co-receptor Antagonists Bind to Similar Regions of CCR5 but Have Differential Activities against UK-427,857-resistant Primary Isolates CROI abstract #96.
Mike Westby*, C Smith-Burchnell, D Hamilton, J Mori, M Macartney, N Robas, B Irvine, M Fidock, F Perruccio, J Mills, K Burt, C Barber, P Stephenson, P Dorr, and M Perros
Pfizer Global Res and Devt, Sandwich, UK
Background: CCR5-tropic HIV-1 variants, selected during prolonged serial passage of virus in the presence of HIV co-receptor antagonists, appear to use compound-bound receptor to infect target cells. Previously published studies suggest that all small molecule CCR5 antagonists bind to a similar region of the receptor, raising the possibility of generalized class-resistance.
Methods: We have located the binding site of UK-427,857 and structurally related compounds within CCR5, using a panel of site-directed mutants and a 3-dimensional model based on homology with bovine rhodopsin. We also measured their potency against R5 viruses, including CCR5-tropic variants of primary HIV-1 isolates that are highly resistant to UK-427,857 (427res).
Results: All the antagonists bind CCR5 in a pocket formed by the trans-membrane helices and extracellular loop 2 (ECL2), centered on an ionic interaction with E283 and a hydrophobic interaction with Y108. The compounds have low nanomolar potency against the R5 lab-adapted HIV-1 strain, Ba-L, and are active against the wild type R5 primary isolates, CC1/85 and RU570. Compounds that contain a triazole moiety were inactive against 427res variants of CC1/85 and RU570. In contrast, compounds where the triazole is replaced by an imidazopiperidine retained activity.
Conclusions: Substitution of a key functional group in a series of structurally related HIV co-receptor antagonists leads to biologically significant changes in the way 427res viruses interact with compound-bound CCR5. It appears that subtle differences in the occupation of the binding pocket, in particular around the ECL2 interface, enable some compounds to block replication of 427res strains. Encouragingly, our data indicate that resistance to an HIV co-receptor antagonist will not necessarily lead to drug-class resistance.