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Perspective
“Chemokine receptors as therapeutic targets: Why aren’t there more drugs?” Roberto Solari c,n, James E. Pease b, Malcolm Begg a a
Refractory Respiratory Inflammation DPU, Respiratory Therapy Area Unit, GlaxoSmithKline, Gunnels Wood Road, Stevenage, Herts SG1 2NY, UK Leukocyte Biology Section, National Heart and Lung Institute, Imperial College London, South Kensington Campus, London SW7 2AZ, UK c Airway Disease Infection Section, National Heart and Lung Institute, Imperial College, Norfolk Place, London W2 1 PG, UK b
art ic l e i nf o
a b s t r a c t
Article history: Received 1 May 2014 Received in revised form 19 June 2014 Accepted 19 June 2014
Chemokines are a family of around 40 small proteins, which are secreted by a variety of cells, including structural cell types and leukocytes of the immune system. Chemokines bind to their specific 7transmembrane G protein-coupled receptors (GPCRs) and induce a variety of downstream signals which notably modulate polymerization of the actin cytoskeleton and thus drive cellular motility. Excessive or inappropriate release of chemokines is observed in many inflammatory diseases and so there has been a great effort in industry to target chemokine receptors. The large family of GPCRs regulate many physiological cellular processes and they have proved to be highly amenable to pharmacological intervention with small chemicals. Consequently GPCRs make attractive targets for drug discovery and indeed a large number of successful current therapeutics are either agonists or antagonists of GPCRs. The apparent lack of success with chemokine receptors has been frustrating and in this paper we discuss potential reasons for previous failures and also why there is considerable cause for optimism. & 2014 Elsevier B.V. All rights reserved.
Keywords: Chemokine Chemokine receptor Drug discovery Inflammation
1. Reasons for optimism The ability of motile cells such as leukocytes to distinguish between minute differences in chemokine concentrations arising from a site of inflammation results in directed migration or chemotaxis of the cell along the gradient of chemokine. The inadvertent, persistent or over expression of chemokines has been linked with the inflammatory component of several clinically important diseases, notably rheumatoid arthritis, asthma and multiple sclerosis. Chemokines have been validated as targets by genetic deletion or antibody neutralization and this has often been shown to be efficacious in treating disease symptoms in murine models of such diseases. Consequently, the blockade of leukocyte recruitment by chemokine receptors antagonists in the human inflammatory setting has long been seen as an attractive proposition (Viola and Luster, 2008). Unlike most GPCR ligands, chemokines are proteins. Although the blockade of protein:protein interactions by small molecules is usually considered too challenging by the pharmaceutical industry, the fact that chemokine receptors belong to the class A family of GPCRs initially gave cause for optimism and triggered many drug discovery programmes (Table 1). Numerous members of the GPCR superfamily are already the targets of many “blockbuster”
n
Corresponding author. E-mail address: [email protected] (R. Solari).
therapeutics and these receptors typically possess a major and minor hydrophobic binding pocket into which small molecules can enter and antagonize receptor function (Jacoby et al., 2006; Rosenkilde et al., 2010; Congreve et al., 2011). Early forays into chemokine receptor antagonists were rewarded with the discovery of small molecules with low nanomolar activity in vitro, although often a lack of activity at the murine counterpart led to problems in target validation in vivo (Pease and Horuk, 2012). The recent plethora of family A GPCR crystal structures, obtained in the inactive conformation in complex with specific antagonists, has greatly increased our understanding at the molecular level of the process of receptor inactivation (Venkatakrishnan et al., 2013). This leads us to believe that in future, the rational design of chemokine receptor antagonists will become science fact and not science fiction (Congreve et al., 2011). As we chip away at the complex biology of the chemokine system, there appear to be numerous additional disease areas opening up to chemokine receptor antagonists. One such example is the recently reported use of the CXCR4 antagonist Plerixafor to correct the uncontrolled signalling of mutant CXCR4 observed in patients with WHIM syndrome (McDermott et al., 2011). Excitingly, some totally unexpected disease areas are also opening up to chemokine receptor antagonists with enormous potential. For example, the chemokine receptor CCR3, an early target for the pharmaceutical industry in asthma and allergy, has subsequently been shown to play a role in age related macular degeneration (AMD) (Takeda et al., 2009). Expression of the receptor and ligands
http://dx.doi.org/10.1016/j.ejphar.2014.06.060 0014-2999/& 2014 Elsevier B.V. All rights reserved.
Please cite this article as: Solari, R., et al., “Chemokine receptors as therapeutic targets: Why aren’t there more drugs?”. Eur J Pharmacol (2014), http://dx.doi.org/10.1016/j.ejphar.2014.06.060i
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Table 1 Chemokine receptor drugs launched, in clinical trials or terminated. Therapeutic indication
Drug target
Drug name
Company
Clinical phase
Activity
HIV infection Hematopoeitic stem cell mobilization
CCR5 CXCR4
Pfizer Genzyme
Approved Approved
Launched Launched
T-cell lymphoma (and allergic diseases)
CCR4
Launched
III III II
Terminated Ongoing Ongoing
Asthma and allergic rhinitis Asthma Allergic rhinitis Transplant rejection, reperfusion injury COPD COPD, asthma, psoriasis HIV infection Rheumatoid arthritis
CCR5 CCR9 CCR3 (and IL-3/IL5/GMCSF) CCR3 CCR3 CCR3 CXCR1/2 CXCR2 CXCR2 CXCR4 CCR1
Amgen/KyowaHakko Schering-Plough Chemocentryx Pharmaxis
Approved
HIV infection Cröhn's disease, celiac disease Asthma
Maraviroc Plerixafor (Mozobil/ AMD3100) Mogamulizumab (KW-0761) Vicriviroc CCX282 (Traficet) ASM8 GSK766994 GSK766904 AZD3778 Reparixin SCH 527123 SCH-527123 AMD-070 MLN-3897
II II II II II II II II
Terminated Terminated Terminated Terminated Terminated Terminated Terminated Terminated
COPD Multiple sclerosis Neuropathic pain, insulin resistance
CCR1 CCR2 CCR2
AZD4818 INCB3284 BMS-741672
II II II
Terminated Terminated Terminated
Pain, liver disease Allergic rhinitis Multiple sclerosis Rheumatoid arthritis HIV infection Diabetic nephropathy, lupus nephritis, rheumatoid arthritis, psoriasis, restenosis Cancer Atherosclerosis, multiple sclerosis
CCR2 CCR2 CCR2 CCR5 CCR5 MCP-1
PF-4136309 JNJ-17166864 MK0812 AZD-5672 PF-232798 Bindarit
GSK GSK Astra Zeneca Dompe Schering-Plough Pharmacopeia AnorMed Millennium Pharmaceuticals AstraZeneca Incyte Bristol-Meyers Squibb Incyte Johnson & Johnson Merck and Co. AstraZeneca Pfizer Angelini
II II II II II II
Terminated Terminated Terminated Terminated Terminated Terminated
CXCR4 CCR2
BKT-140 MLN-1202
II II
Terminated Terminated
Ulcerative colitis, rheumatoid arthritis Cancer, pulmonary fibrosis Allergy ocular
CXCL10 CCL2 CCL11
MDX-1100 CNTO-888 Bertilimumab
II II II
Terminated Terminated Ongoing
Hematopoeitic stem cell mobilization
CXCR4
II
Ongoing
Cancer
CXCR4
POL5551 and POL6326 CTCE-9908
Biokine Millennium Pharmaceuticals Medarex Centocor Immune Pharmaceuticals Polyphor
I/II
Terminated
Asthma Asthma Asthma Allergic rhinitis Asthma COPD, cystic fibrosis COPD, cystic fibrosis
CCR3 CCR3 CCR3 CCR3 CCR4 CXCR2 CXCR2
GW824575 DPC168 BMS-639623 QAP-642 GSK2239633 GSK656933 SB-265610
I I I I I I I
Terminated Terminated Terminated Terminated Terminated Terminated Terminated
Stem cell transplantation Stem cell transplantation
CXCR4 CXCR4
POL-6326 TG-0054
I I
Terminated Ongoing
Rheumatoid arthritis Rheumatoid arthritis
CCR1 CCR1
CCX-354 MLN-3701
I I
Terminated Ongoing
Multiple sclerosis Multiple sclerosis Asthma HIV infection HIV infection HIV infection HIV infection
CCR2 CCR2 CCR3 CCR5 CCR5 CCR5 CCR5
CCX-140 INCB-8696 AZD1744 GSK706769 INCB-15050 AZD5672 TBR-220
I I I I I I I
Terminated Terminated Terminated Terminated Terminated Terminated Terminated
HIV infection
CCR5
TBR-652
I
Terminated
HIV infection Rheumatoid arthritis Gastrointestinal disease AML HIV infection
CCR5 CCR5 CCR9 CXCR4 CCR5
VCH-286 Maraviroc CCX-025 MDX-1338 CCR5mAb004
I I I I I
Terminated Terminated Terminated Terminated Terminated
Autoimmunity Asthma, chronic obstructive pulmonary disease, rheumatoid arthritis, inflammatory bowel disease and psoriasis
CXCL10
NI-0801
I
Multiple
FX-125L
Ongoing Terminated
Chemokine Therapeutics GSK DuPont BMS Novartis GSK GSK SmithKline Beecham Polyphor Taigen Biotechnology ChemoCentryx Millennium Pharmaceuticals ChemoCentryx Incyte AstraZeneca GlaxoSmithKline Incyte AstraZeneca Takeda Pharmaceuticals Takeda Pharmaceuticals ViroChem Pharma Pfizer ChemoCentryx Medarex Human Genome Sciences NovImmune Funxional Therapeutics
I
Please cite this article as: Solari, R., et al., “Chemokine receptors as therapeutic targets: Why aren’t there more drugs?”. Eur J Pharmacol (2014), http://dx.doi.org/10.1016/j.ejphar.2014.06.060i
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by the choroidal endothelium results in neovascularization and retinal damage. In some mouse models of AMD, CCR3 blockade has excellent efficacy, suggesting a novel clinical use for small molecule CCR3 antagonists. Similarly, increased expression of the key CCR3 ligand CCL11/Eotaxin-1 has been associated with cognitive dysfunction in aging mice (Villeda et al., 2011). Collectively, this suggests that CCR3 antagonism may find favor in two disorders associated with an aging population, far removed from the original intended purpose of such drugs. There is also growing interest in targeting the CXCL8 chemokine for oncology indications (Campbell et al., 2013; Singh et al., 2013).
2. The current state of affairs Arguably the greatest success in chemokine receptor therapeutics is the CCR5 receptor antagonist Maraviroc, which is used in the treatment of HIV. However, despite this success story, other CCR5 antagonists appear to have failed during development (Palani and Tagat, 2006; Pulley, 2007). The only other chemokine receptor ligand marketed is the CXCR4 antagonist/partial agonist Plerixafor which was originally developed to treat HIV infection but was subsequently approved for the mobilization of stem cells from the bone marrow to the bloodstream in cancer patients (Brave et al., 2010). Despite significant investment in medicinal chemistry and pharmacology, other chemokine receptor small molecules agents have failed to make it to market. The reasons for these failures are many and varied (Horuk, 2009) but this does not appear to have stopped drug discovery efforts and there continue to be a number of conventional small molecules progressing through the various phases of pre-clinical and clinical research. The CCR2 antagonist, CCX-140, is in a Phase3 trial for the treatment of Type 2 diabetes and diabetic nephropathy (Hanefeld et al., 2012; Sullivan et al., 2013) and the CCR9 antagonist CCX282 has completed Phase 2 trials for Crohn's disease and ulcerative colitis (Keshav et al., 2013). CCX354, which targets CCR1, has completed a Phase 2 trial for the treatment of rheumatoid arthritis (Dairaghi et al., 2011; Tak et al., 2013) and there have been intense efforts to discover CXCR1/ 2 antagonists, a number of which have made it to clinical trials (ClinicalTrials.gov Identifiers: NCT01068145; NCT01006616; NCT00504439; NCT01209052; Watz et al., 2012; Lazaar et al., 2011; Miller et al., 2014). Finally a novel CCR4 antagonist has also completed a Phase I study (Cahn et al., 2013). However it is currently unclear how many of these programmes are likely to progress much further (Pease and Horuk, 2014). A novel unconventional approach is to move away from making traditional specific receptor antagonists and to discover broad spectrum chemotaxis inhibitors in functional cell based screens (Fox et al., 2009). One example is FX-125L which is an orallyavailable small molecule broad-spectrum chemokine inhibitor under development for the treatment of asthma, chronic obstructive pulmonary disease, rheumatoid arthritis, inflammatory bowel disease and psoriasis (EU Clinical Trials Register EudraCT Number 2011-005036-26). It appears to inhibit the chemoattractant activity of a number of chemokines via an unspecified cell-surface receptor. We are also beginning to see alternative approaches to small molecules. A number of Phase2a studies have recently been completed with ASM-8 in asthma (ClinicalTrials.gov Identifier: NCT00822861). This is a combination of antisense oligonucleotides targeting the common beta chain of the IL3/IL5/GMCSF receptor and the chemokine receptor CCR3 (Imaoka et al., 2011). We are also witnessing the emergence of biological agents such as the CCR4 antagonist Mogamulizumab (KW-0761) that is the first chemokine receptor antibody therapeutic. This humanized
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monoclonal antibody is approved for the treatment of T-cell lymphoma and has the potential to also treat allergic diseases, as it targets CCR4 chemokine receptors selectively expressed in T-helper type 2 (Th2) cells (Subramaniam et al., 2012). Another biological therapy is Bertilimumab, which is a fully-human anti-eotaxin-1 monoclonal antibody under development for the treatment of age-related macular degeneration, asthma, Crohn's disease, pemphigoid and ulcerative colitis (ClinicalTrials.gov Identifier NCT01671956). Chemokine receptor specific nanobodies are also showing efficacy in pre-clinical models so expanding the potential repertoire of biological therapeutics (Maussang et al., 2013). There are also examples of protein epitope mimetic approaches, such as POL5551 and POL6326 which are selective and reversible CXCR4 antagonists, currently in Phase 2 studies for stem cell mobilization in cancer therapy (ClinicalTrials.gov Identifier NCT01105403; Karpova et al., 2013), and this is one of a number of new strategies in development for antagonizing CXCR4 (Rettig et al., 2012). In addition to these conventional and unconventional pharmacology approaches and novel biologicals we are seeing attempts to deliver functional efficacy by agents that drive receptor internalization using a screening process designed to identify molecules that induce chemokine receptor down regulation. TRV-027 is a selective ß-arrestin biased ligand of the angiotensin II type 1 receptor, under development for the treatment of heart failure and CNS indications. It is a competitive antagonist of angiotensin II-stimulated G-protein signaling, but stimulates ß-arrestin recruitment (Violin et al., 2012, 2013). It is now evident that drug discovery strategies for chemokine receptors are highly diverse.
3. The challenges Given the great attraction of chemokine targets and the huge effort expended so far on drug discovery one has to ask why there have been so few successes (Proudfoot et al., 2010; Horuk, 2009). A number of explanations have been proposed but what have we learned from our failures and are there clear ways forward to address these shortcomings (Horuk, 2009; Schall and Proudfoot, 2011)? A major problem is that animal models of inflammatory diseases are poorly predictive of human efficacy (Holmes et al., 2011) and the immune and inflammatory systems of model animal species and man have significant differences which make selecting and validating targets based on animal studies particularly challenging. We have perhaps developed an over reliance on genetic models in mice and simplistic disease models to test drugs. Secondly the chemokine system is complex and potentially highly redundant. Cells can express multiple receptors and each receptor can be promiscuous, binding several ligands. In addition ligands are also promiscuous and can bind several receptors. Chemokine receptors can also form homo- and hetero-dimers and different chemokines acting simultaneously on the same cell can “crosstalk” so greatly further complicating their pharmacology. The chemokines themselves are usually immobilized close to their site of production by binding to glycosaminoglycans (GAGS) as part of the cell guidance mechanism. This species, functional and pharmacological complexity is indeed very challenging. A common interpretation of this apparent redundancy is that multiple chemokines can perform the same physiological function and therefore antagonism of one receptor will never be sufficient. An alternative explanation is that the apparent overlapping functions of chemokines reflect our limited understanding and each chemokine/receptor/cell combination has a specific role in a particular physiologic and pathologic context (Schall and Proudfoot, 2011). In either case what is clear is that we do not have any mechanistic understanding of why the chemokine system has evolved such
Please cite this article as: Solari, R., et al., “Chemokine receptors as therapeutic targets: Why aren’t there more drugs?”. Eur J Pharmacol (2014), http://dx.doi.org/10.1016/j.ejphar.2014.06.060i
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complexity and this makes predicting human efficacy in preclinical models particularly difficult. In terms of receptor pharmacology we are also in our infancy when it comes to our knowledge of these receptors. It has become increasingly clear that chemokine receptors can exhibit complex and biased signalling to different ligands and ligands can be agonists to one receptor and antagonists to another (Kalatskaya et al., 2009; Kenakin and Christopoulos, 2013; Zweemer et al., 2014). In normal physiology, cells need to home to specific sites in the body and in pathology to migrate to sites of inflammation, arrest, function and reverse migrate to resolve inflammation (Robertson et al., 2014). How conventional receptor pharmacology can accommodate all of these spatio-temporal instructions is still poorly appreciated. Finally the chemistry of chemokine receptor antagonists has undoubtedly been problematic. Discovering and developing small molecules with good “drug-like” properties have been a significant problem and many failures may be more compound-related rather than mechanism-related. It is also clear that many small molecules “antagonists” are in fact allosteric modulators and they may bind to multiple sites on the receptor which result in different consequences (Ajram et al., 2014). Another aspect which should be taken into account when targeting chemokine receptors in the disease setting is the potential for changes in receptor expression pattern, particularly following leukocyte activation. For example, it is well documented that stimuli associated with inflammation such as TNF-α can induce the shedding of cell surface chemokine receptors from neutrophils in a metalloproteinase-dependent manner (Asagoe et al., 1998; Khandaker et al., 1999). Similarly, exposure to IFN-γ in vitro can induce the expression of CCR1 and CCR2 on neutrophils (Bonecchi et al., 1999) whilst neutrophils infiltrating inflamed lungs (COPD patients) or joints (rheumatoid arthritis patients) upregulate several CC chemokine receptors not normally associated with these cells (Hartl et al., 2008). Such expression is likely to be clinically relevant, since it was shown ex vivo that stimulation of these receptors can modulate effector functions such as respiratory burst activity. Thus, the chemokine receptor targets need to carefully selected to maximize the therapeutic potential in each disease setting. The final consideration is the potential for toxicity from chemokine antagonism. Chemokine receptor knockouts are mostly healthy and have been useful in determining the roles of different chemokines in various disease models (Power, 2003). This has given encouragement for drug discovery; however, there is also evidence for increased susceptibility to infection and some knockouts have exacerbated symptoms in disease models (Carter, 2002). Although there are few reports of toxicity in clinical trials with chemokine receptor antagonists there are some notable examples. The trial with the CCR5 antagonist Aplaviroc was terminated prematurely due to treatment associated hepatotoxicity although this was considered to be compound related and idiosyncratic (Nichols et al., 2008) and the original anti-HIV trials with Plerixafor were discontinued because of cardiac disturbances .
4. Forward looking opportunities Although we may not yet have all the answers we have learned a great deal from our early efforts at drug discovery and it will certainly be interesting to see in the coming years if novel approaches are more successful than conventional small molecule medicinal chemistry. However, given all the complexities of the chemokine system we are probably not incorporating our current knowledge adequately in our drug discovery strategies. There are many screening platforms we can use to identify novel chemical leads. Often we have used radioligand competition binding or
GTPγS binding assays to membrane preparations from cell lines overexpressing transfected receptors. Clearly such assays will miss out most of the rich receptor functional complexity we have discussed. More recently we have used β-arrestin complementation assays, but although this is performed on a whole cell, only one signalling pathway is being explored and once again the cell line is artificially engineered so missing most of the potential receptor and ligand cross talk. We can run high throughput FLIPR assays to measure Ca2 þ transients, or actin polymerization but once again this is usually in engineered cell lines and only measures one signalling readout. Finally we almost always screen our ligands in solution and not bound to GAGS as they appear in vivo. Surely given all the challenges we face and insights we have we should focus on relevant phenotypic screening assays in relevant primary human cells (Feng et al., 2009). Reductionist assays developed for the convenience of high throughput screening platforms have probably not helped the drug discovery process. We should trust our biological and clinical insights to drive drug discovery strategies rather than relying on the latest fashion in screening technology. Finally we believe one should put much more weight on human clinical data for discovering and validating targets rather than an over dependence on mouse models. If we can incorporate all our learnings to date we stand a much better chance of delivering on the promise of chemokine receptor drugs.
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Perspective
“Chemokine receptors as therapeutic targets: Why aren’t there more drugs?” Roberto Solari c,n, James E. Pease b, Malcolm Begg a a
Refractory Respiratory Inflammation DPU, Respiratory Therapy Area Unit, GlaxoSmithKline, Gunnels Wood Road, Stevenage, Herts SG1 2NY, UK Leukocyte Biology Section, National Heart and Lung Institute, Imperial College London, South Kensington Campus, London SW7 2AZ, UK c Airway Disease Infection Section, National Heart and Lung Institute, Imperial College, Norfolk Place, London W2 1 PG, UK b
art ic l e i nf o
a b s t r a c t
Article history: Received 1 May 2014 Received in revised form 19 June 2014 Accepted 19 June 2014
Chemokines are a family of around 40 small proteins, which are secreted by a variety of cells, including structural cell types and leukocytes of the immune system. Chemokines bind to their specific 7transmembrane G protein-coupled receptors (GPCRs) and induce a variety of downstream signals which notably modulate polymerization of the actin cytoskeleton and thus drive cellular motility. Excessive or inappropriate release of chemokines is observed in many inflammatory diseases and so there has been a great effort in industry to target chemokine receptors. The large family of GPCRs regulate many physiological cellular processes and they have proved to be highly amenable to pharmacological intervention with small chemicals. Consequently GPCRs make attractive targets for drug discovery and indeed a large number of successful current therapeutics are either agonists or antagonists of GPCRs. The apparent lack of success with chemokine receptors has been frustrating and in this paper we discuss potential reasons for previous failures and also why there is considerable cause for optimism. & 2014 Elsevier B.V. All rights reserved.
Keywords: Chemokine Chemokine receptor Drug discovery Inflammation
1. Reasons for optimism The ability of motile cells such as leukocytes to distinguish between minute differences in chemokine concentrations arising from a site of inflammation results in directed migration or chemotaxis of the cell along the gradient of chemokine. The inadvertent, persistent or over expression of chemokines has been linked with the inflammatory component of several clinically important diseases, notably rheumatoid arthritis, asthma and multiple sclerosis. Chemokines have been validated as targets by genetic deletion or antibody neutralization and this has often been shown to be efficacious in treating disease symptoms in murine models of such diseases. Consequently, the blockade of leukocyte recruitment by chemokine receptors antagonists in the human inflammatory setting has long been seen as an attractive proposition (Viola and Luster, 2008). Unlike most GPCR ligands, chemokines are proteins. Although the blockade of protein:protein interactions by small molecules is usually considered too challenging by the pharmaceutical industry, the fact that chemokine receptors belong to the class A family of GPCRs initially gave cause for optimism and triggered many drug discovery programmes (Table 1). Numerous members of the GPCR superfamily are already the targets of many “blockbuster”
n
Corresponding author. E-mail address: [email protected] (R. Solari).
therapeutics and these receptors typically possess a major and minor hydrophobic binding pocket into which small molecules can enter and antagonize receptor function (Jacoby et al., 2006; Rosenkilde et al., 2010; Congreve et al., 2011). Early forays into chemokine receptor antagonists were rewarded with the discovery of small molecules with low nanomolar activity in vitro, although often a lack of activity at the murine counterpart led to problems in target validation in vivo (Pease and Horuk, 2012). The recent plethora of family A GPCR crystal structures, obtained in the inactive conformation in complex with specific antagonists, has greatly increased our understanding at the molecular level of the process of receptor inactivation (Venkatakrishnan et al., 2013). This leads us to believe that in future, the rational design of chemokine receptor antagonists will become science fact and not science fiction (Congreve et al., 2011). As we chip away at the complex biology of the chemokine system, there appear to be numerous additional disease areas opening up to chemokine receptor antagonists. One such example is the recently reported use of the CXCR4 antagonist Plerixafor to correct the uncontrolled signalling of mutant CXCR4 observed in patients with WHIM syndrome (McDermott et al., 2011). Excitingly, some totally unexpected disease areas are also opening up to chemokine receptor antagonists with enormous potential. For example, the chemokine receptor CCR3, an early target for the pharmaceutical industry in asthma and allergy, has subsequently been shown to play a role in age related macular degeneration (AMD) (Takeda et al., 2009). Expression of the receptor and ligands
http://dx.doi.org/10.1016/j.ejphar.2014.06.060 0014-2999/& 2014 Elsevier B.V. All rights reserved.
Please cite this article as: Solari, R., et al., “Chemokine receptors as therapeutic targets: Why aren’t there more drugs?”. Eur J Pharmacol (2014), http://dx.doi.org/10.1016/j.ejphar.2014.06.060i
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Table 1 Chemokine receptor drugs launched, in clinical trials or terminated. Therapeutic indication
Drug target
Drug name
Company
Clinical phase
Activity
HIV infection Hematopoeitic stem cell mobilization
CCR5 CXCR4
Pfizer Genzyme
Approved Approved
Launched Launched
T-cell lymphoma (and allergic diseases)
CCR4
Launched
III III II
Terminated Ongoing Ongoing
Asthma and allergic rhinitis Asthma Allergic rhinitis Transplant rejection, reperfusion injury COPD COPD, asthma, psoriasis HIV infection Rheumatoid arthritis
CCR5 CCR9 CCR3 (and IL-3/IL5/GMCSF) CCR3 CCR3 CCR3 CXCR1/2 CXCR2 CXCR2 CXCR4 CCR1
Amgen/KyowaHakko Schering-Plough Chemocentryx Pharmaxis
Approved
HIV infection Cröhn's disease, celiac disease Asthma
Maraviroc Plerixafor (Mozobil/ AMD3100) Mogamulizumab (KW-0761) Vicriviroc CCX282 (Traficet) ASM8 GSK766994 GSK766904 AZD3778 Reparixin SCH 527123 SCH-527123 AMD-070 MLN-3897
II II II II II II II II
Terminated Terminated Terminated Terminated Terminated Terminated Terminated Terminated
COPD Multiple sclerosis Neuropathic pain, insulin resistance
CCR1 CCR2 CCR2
AZD4818 INCB3284 BMS-741672
II II II
Terminated Terminated Terminated
Pain, liver disease Allergic rhinitis Multiple sclerosis Rheumatoid arthritis HIV infection Diabetic nephropathy, lupus nephritis, rheumatoid arthritis, psoriasis, restenosis Cancer Atherosclerosis, multiple sclerosis
CCR2 CCR2 CCR2 CCR5 CCR5 MCP-1
PF-4136309 JNJ-17166864 MK0812 AZD-5672 PF-232798 Bindarit
GSK GSK Astra Zeneca Dompe Schering-Plough Pharmacopeia AnorMed Millennium Pharmaceuticals AstraZeneca Incyte Bristol-Meyers Squibb Incyte Johnson & Johnson Merck and Co. AstraZeneca Pfizer Angelini
II II II II II II
Terminated Terminated Terminated Terminated Terminated Terminated
CXCR4 CCR2
BKT-140 MLN-1202
II II
Terminated Terminated
Ulcerative colitis, rheumatoid arthritis Cancer, pulmonary fibrosis Allergy ocular
CXCL10 CCL2 CCL11
MDX-1100 CNTO-888 Bertilimumab
II II II
Terminated Terminated Ongoing
Hematopoeitic stem cell mobilization
CXCR4
II
Ongoing
Cancer
CXCR4
POL5551 and POL6326 CTCE-9908
Biokine Millennium Pharmaceuticals Medarex Centocor Immune Pharmaceuticals Polyphor
I/II
Terminated
Asthma Asthma Asthma Allergic rhinitis Asthma COPD, cystic fibrosis COPD, cystic fibrosis
CCR3 CCR3 CCR3 CCR3 CCR4 CXCR2 CXCR2
GW824575 DPC168 BMS-639623 QAP-642 GSK2239633 GSK656933 SB-265610
I I I I I I I
Terminated Terminated Terminated Terminated Terminated Terminated Terminated
Stem cell transplantation Stem cell transplantation
CXCR4 CXCR4
POL-6326 TG-0054
I I
Terminated Ongoing
Rheumatoid arthritis Rheumatoid arthritis
CCR1 CCR1
CCX-354 MLN-3701
I I
Terminated Ongoing
Multiple sclerosis Multiple sclerosis Asthma HIV infection HIV infection HIV infection HIV infection
CCR2 CCR2 CCR3 CCR5 CCR5 CCR5 CCR5
CCX-140 INCB-8696 AZD1744 GSK706769 INCB-15050 AZD5672 TBR-220
I I I I I I I
Terminated Terminated Terminated Terminated Terminated Terminated Terminated
HIV infection
CCR5
TBR-652
I
Terminated
HIV infection Rheumatoid arthritis Gastrointestinal disease AML HIV infection
CCR5 CCR5 CCR9 CXCR4 CCR5
VCH-286 Maraviroc CCX-025 MDX-1338 CCR5mAb004
I I I I I
Terminated Terminated Terminated Terminated Terminated
Autoimmunity Asthma, chronic obstructive pulmonary disease, rheumatoid arthritis, inflammatory bowel disease and psoriasis
CXCL10
NI-0801
I
Multiple
FX-125L
Ongoing Terminated
Chemokine Therapeutics GSK DuPont BMS Novartis GSK GSK SmithKline Beecham Polyphor Taigen Biotechnology ChemoCentryx Millennium Pharmaceuticals ChemoCentryx Incyte AstraZeneca GlaxoSmithKline Incyte AstraZeneca Takeda Pharmaceuticals Takeda Pharmaceuticals ViroChem Pharma Pfizer ChemoCentryx Medarex Human Genome Sciences NovImmune Funxional Therapeutics
I
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by the choroidal endothelium results in neovascularization and retinal damage. In some mouse models of AMD, CCR3 blockade has excellent efficacy, suggesting a novel clinical use for small molecule CCR3 antagonists. Similarly, increased expression of the key CCR3 ligand CCL11/Eotaxin-1 has been associated with cognitive dysfunction in aging mice (Villeda et al., 2011). Collectively, this suggests that CCR3 antagonism may find favor in two disorders associated with an aging population, far removed from the original intended purpose of such drugs. There is also growing interest in targeting the CXCL8 chemokine for oncology indications (Campbell et al., 2013; Singh et al., 2013).
2. The current state of affairs Arguably the greatest success in chemokine receptor therapeutics is the CCR5 receptor antagonist Maraviroc, which is used in the treatment of HIV. However, despite this success story, other CCR5 antagonists appear to have failed during development (Palani and Tagat, 2006; Pulley, 2007). The only other chemokine receptor ligand marketed is the CXCR4 antagonist/partial agonist Plerixafor which was originally developed to treat HIV infection but was subsequently approved for the mobilization of stem cells from the bone marrow to the bloodstream in cancer patients (Brave et al., 2010). Despite significant investment in medicinal chemistry and pharmacology, other chemokine receptor small molecules agents have failed to make it to market. The reasons for these failures are many and varied (Horuk, 2009) but this does not appear to have stopped drug discovery efforts and there continue to be a number of conventional small molecules progressing through the various phases of pre-clinical and clinical research. The CCR2 antagonist, CCX-140, is in a Phase3 trial for the treatment of Type 2 diabetes and diabetic nephropathy (Hanefeld et al., 2012; Sullivan et al., 2013) and the CCR9 antagonist CCX282 has completed Phase 2 trials for Crohn's disease and ulcerative colitis (Keshav et al., 2013). CCX354, which targets CCR1, has completed a Phase 2 trial for the treatment of rheumatoid arthritis (Dairaghi et al., 2011; Tak et al., 2013) and there have been intense efforts to discover CXCR1/ 2 antagonists, a number of which have made it to clinical trials (ClinicalTrials.gov Identifiers: NCT01068145; NCT01006616; NCT00504439; NCT01209052; Watz et al., 2012; Lazaar et al., 2011; Miller et al., 2014). Finally a novel CCR4 antagonist has also completed a Phase I study (Cahn et al., 2013). However it is currently unclear how many of these programmes are likely to progress much further (Pease and Horuk, 2014). A novel unconventional approach is to move away from making traditional specific receptor antagonists and to discover broad spectrum chemotaxis inhibitors in functional cell based screens (Fox et al., 2009). One example is FX-125L which is an orallyavailable small molecule broad-spectrum chemokine inhibitor under development for the treatment of asthma, chronic obstructive pulmonary disease, rheumatoid arthritis, inflammatory bowel disease and psoriasis (EU Clinical Trials Register EudraCT Number 2011-005036-26). It appears to inhibit the chemoattractant activity of a number of chemokines via an unspecified cell-surface receptor. We are also beginning to see alternative approaches to small molecules. A number of Phase2a studies have recently been completed with ASM-8 in asthma (ClinicalTrials.gov Identifier: NCT00822861). This is a combination of antisense oligonucleotides targeting the common beta chain of the IL3/IL5/GMCSF receptor and the chemokine receptor CCR3 (Imaoka et al., 2011). We are also witnessing the emergence of biological agents such as the CCR4 antagonist Mogamulizumab (KW-0761) that is the first chemokine receptor antibody therapeutic. This humanized
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monoclonal antibody is approved for the treatment of T-cell lymphoma and has the potential to also treat allergic diseases, as it targets CCR4 chemokine receptors selectively expressed in T-helper type 2 (Th2) cells (Subramaniam et al., 2012). Another biological therapy is Bertilimumab, which is a fully-human anti-eotaxin-1 monoclonal antibody under development for the treatment of age-related macular degeneration, asthma, Crohn's disease, pemphigoid and ulcerative colitis (ClinicalTrials.gov Identifier NCT01671956). Chemokine receptor specific nanobodies are also showing efficacy in pre-clinical models so expanding the potential repertoire of biological therapeutics (Maussang et al., 2013). There are also examples of protein epitope mimetic approaches, such as POL5551 and POL6326 which are selective and reversible CXCR4 antagonists, currently in Phase 2 studies for stem cell mobilization in cancer therapy (ClinicalTrials.gov Identifier NCT01105403; Karpova et al., 2013), and this is one of a number of new strategies in development for antagonizing CXCR4 (Rettig et al., 2012). In addition to these conventional and unconventional pharmacology approaches and novel biologicals we are seeing attempts to deliver functional efficacy by agents that drive receptor internalization using a screening process designed to identify molecules that induce chemokine receptor down regulation. TRV-027 is a selective ß-arrestin biased ligand of the angiotensin II type 1 receptor, under development for the treatment of heart failure and CNS indications. It is a competitive antagonist of angiotensin II-stimulated G-protein signaling, but stimulates ß-arrestin recruitment (Violin et al., 2012, 2013). It is now evident that drug discovery strategies for chemokine receptors are highly diverse.
3. The challenges Given the great attraction of chemokine targets and the huge effort expended so far on drug discovery one has to ask why there have been so few successes (Proudfoot et al., 2010; Horuk, 2009). A number of explanations have been proposed but what have we learned from our failures and are there clear ways forward to address these shortcomings (Horuk, 2009; Schall and Proudfoot, 2011)? A major problem is that animal models of inflammatory diseases are poorly predictive of human efficacy (Holmes et al., 2011) and the immune and inflammatory systems of model animal species and man have significant differences which make selecting and validating targets based on animal studies particularly challenging. We have perhaps developed an over reliance on genetic models in mice and simplistic disease models to test drugs. Secondly the chemokine system is complex and potentially highly redundant. Cells can express multiple receptors and each receptor can be promiscuous, binding several ligands. In addition ligands are also promiscuous and can bind several receptors. Chemokine receptors can also form homo- and hetero-dimers and different chemokines acting simultaneously on the same cell can “crosstalk” so greatly further complicating their pharmacology. The chemokines themselves are usually immobilized close to their site of production by binding to glycosaminoglycans (GAGS) as part of the cell guidance mechanism. This species, functional and pharmacological complexity is indeed very challenging. A common interpretation of this apparent redundancy is that multiple chemokines can perform the same physiological function and therefore antagonism of one receptor will never be sufficient. An alternative explanation is that the apparent overlapping functions of chemokines reflect our limited understanding and each chemokine/receptor/cell combination has a specific role in a particular physiologic and pathologic context (Schall and Proudfoot, 2011). In either case what is clear is that we do not have any mechanistic understanding of why the chemokine system has evolved such
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complexity and this makes predicting human efficacy in preclinical models particularly difficult. In terms of receptor pharmacology we are also in our infancy when it comes to our knowledge of these receptors. It has become increasingly clear that chemokine receptors can exhibit complex and biased signalling to different ligands and ligands can be agonists to one receptor and antagonists to another (Kalatskaya et al., 2009; Kenakin and Christopoulos, 2013; Zweemer et al., 2014). In normal physiology, cells need to home to specific sites in the body and in pathology to migrate to sites of inflammation, arrest, function and reverse migrate to resolve inflammation (Robertson et al., 2014). How conventional receptor pharmacology can accommodate all of these spatio-temporal instructions is still poorly appreciated. Finally the chemistry of chemokine receptor antagonists has undoubtedly been problematic. Discovering and developing small molecules with good “drug-like” properties have been a significant problem and many failures may be more compound-related rather than mechanism-related. It is also clear that many small molecules “antagonists” are in fact allosteric modulators and they may bind to multiple sites on the receptor which result in different consequences (Ajram et al., 2014). Another aspect which should be taken into account when targeting chemokine receptors in the disease setting is the potential for changes in receptor expression pattern, particularly following leukocyte activation. For example, it is well documented that stimuli associated with inflammation such as TNF-α can induce the shedding of cell surface chemokine receptors from neutrophils in a metalloproteinase-dependent manner (Asagoe et al., 1998; Khandaker et al., 1999). Similarly, exposure to IFN-γ in vitro can induce the expression of CCR1 and CCR2 on neutrophils (Bonecchi et al., 1999) whilst neutrophils infiltrating inflamed lungs (COPD patients) or joints (rheumatoid arthritis patients) upregulate several CC chemokine receptors not normally associated with these cells (Hartl et al., 2008). Such expression is likely to be clinically relevant, since it was shown ex vivo that stimulation of these receptors can modulate effector functions such as respiratory burst activity. Thus, the chemokine receptor targets need to carefully selected to maximize the therapeutic potential in each disease setting. The final consideration is the potential for toxicity from chemokine antagonism. Chemokine receptor knockouts are mostly healthy and have been useful in determining the roles of different chemokines in various disease models (Power, 2003). This has given encouragement for drug discovery; however, there is also evidence for increased susceptibility to infection and some knockouts have exacerbated symptoms in disease models (Carter, 2002). Although there are few reports of toxicity in clinical trials with chemokine receptor antagonists there are some notable examples. The trial with the CCR5 antagonist Aplaviroc was terminated prematurely due to treatment associated hepatotoxicity although this was considered to be compound related and idiosyncratic (Nichols et al., 2008) and the original anti-HIV trials with Plerixafor were discontinued because of cardiac disturbances .
4. Forward looking opportunities Although we may not yet have all the answers we have learned a great deal from our early efforts at drug discovery and it will certainly be interesting to see in the coming years if novel approaches are more successful than conventional small molecule medicinal chemistry. However, given all the complexities of the chemokine system we are probably not incorporating our current knowledge adequately in our drug discovery strategies. There are many screening platforms we can use to identify novel chemical leads. Often we have used radioligand competition binding or
GTPγS binding assays to membrane preparations from cell lines overexpressing transfected receptors. Clearly such assays will miss out most of the rich receptor functional complexity we have discussed. More recently we have used β-arrestin complementation assays, but although this is performed on a whole cell, only one signalling pathway is being explored and once again the cell line is artificially engineered so missing most of the potential receptor and ligand cross talk. We can run high throughput FLIPR assays to measure Ca2 þ transients, or actin polymerization but once again this is usually in engineered cell lines and only measures one signalling readout. Finally we almost always screen our ligands in solution and not bound to GAGS as they appear in vivo. Surely given all the challenges we face and insights we have we should focus on relevant phenotypic screening assays in relevant primary human cells (Feng et al., 2009). Reductionist assays developed for the convenience of high throughput screening platforms have probably not helped the drug discovery process. We should trust our biological and clinical insights to drive drug discovery strategies rather than relying on the latest fashion in screening technology. Finally we believe one should put much more weight on human clinical data for discovering and validating targets rather than an over dependence on mouse models. If we can incorporate all our learnings to date we stand a much better chance of delivering on the promise of chemokine receptor drugs.
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