The design, synthesis and optimisaion of CCR4 antagonists for the treatment of asthma

This thesis was previously held under moratorium in Chemistry Department (GSK) from 25th August 2015 until 1st October 2017.The 7-transmembrane chemokine receptor, CCR4 is differentially expressed on T helper type 2 (TH2) cells; together with its endogenous ligands, CCL17 and CCL22, it promotes activation and recruitment of TH2 cells into the lungs. Infiltration of TH2 cells into the airways is an important characteristic of allergic asthma, so inhibition of CCR4 binding to its endogenous ligands might provide a potential therapeutic target for treating the disease.In order to identify a potential back-up to the clinical development candidate, N-(3-((3-(5-chlorothiophene-2-sulfonamido)-4-methoxy-1H-indazol-1-yl)methyl)benzyl)-2-hydroxy-2-methylpropanamide (6), a library of 2,3-dichlorobenzene sulfonamides were designed. Synthesis and evaluation of the 'hits' in the primary antagonist assay (GTPγS) led to the identification of three novel templates with good potency and ligand efficiency (LE). The benzimidazolone template, exemplified by 88 (2,3-dichloro-N-(6-methoxy-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl) benzenesulfonamide), demonstrated a good PK profile (low clearance, moderate to high oral bioavailability) in both rat and dog, and had better potency in the human whole blood (hWB) assay compared to the other two series, and hence was explored further. Investigation of substituents of the benzimidazolone series, and incorporation of nitrogen into the core led to the discovery of 239 (2,3-dichloro-N-(6-methoxy-2-oxo-2,3-dihydro-1H-imidazo[4,5-b]pyridin-5-yl)benzenesulfonamide), which had a pA2 value of 6.0 in the hWB assay and a much improved LE (0.43) compared to indazole 6 (LE = 0.30). The azabenzimidazolone 239 provides a new starting point for further lead optimisation to develop a novel allosteric CCR4 antagonist from which a potential oral asthma drug could emerge.A phenylpyrazole sulfonamide (N-(2-(1H-pyrazol-1-yl)phenyl)phenylsulfonamide) was also identified, where X-ray diffraction studies showed that one of the nitrogens of the pyrazole ring forms an intramolecular hydrogen bond with the sulfonamide NH and promotes a conformation that was thought to be preferred in the active site. A SAR study around the phenylpyrazole core provided pyridine analogue 115 (N-(3-(1H-pyrazol-1-yl)pyridin-2-yl)-5-chlorothiophene-2-sulfonamide) with very good potency, high LE (0.47) and excellent physicochemical properties (chromlogD7.4 = 2.7, MW = 340, solubility = 133 μg/mL). Pyridylpyrazole 115 also represent good starting points for a lead optimisation programme.In addition, a novel series of 2,8-diazaspiro[4.5]decan-8-yl)pyrimidin-4-amine inhibitors, were identified using computational modelling studies, which bind to a different allosteric region on the CCR4 receptor compared to indazole 6. Evaluation of SAR led to identification of several analogues that exhibited hWB activity superior or comparable to that of 6. The most potent compound, 282 (N-(2,4-dichlorobenzyl)-2-(2-(pyrrolidin-2-ylmethyl)-2,8-diazaspiro[4.5]decan-8-yl)pyrimidin-4-amine, hWB pA2 = 6.7) demonstrated anti-inflammatory activity in acute in vivo murine allergic inflammation models. After an ovalbumin challenge in mice, compound 282 showed not only significant reduction in eosinophil and lymphocyte infiltration into the bronchoalveolar lavage (BAL) fluid but also displayed a profound effect on airway hyperreactivity, returning the levels back to the control line. The in vivo results indicate that targeting CCR4 with a small molecule allosteric antagonist could be a potential novel way of treating allergic diseases such as asthma. Interestingly, representative compounds from the spiro-pyrimidine series were found to induce receptor internalisation of CCR4 in HUT78 cell with approximately 50% reduction of cell surface receptor. It is suggested that in vivo activity of 282 might be due to its ability to induce internalisation of the receptor.The 7-transmembrane chemokine receptor, CCR4 is differentially expressed on T helper type 2 (TH2) cells; together with its endogenous ligands, CCL17 and CCL22, it promotes activation and recruitment of TH2 cells into the lungs. Infiltration of TH2 cells into the airways is an important characteristic of allergic asthma, so inhibition of CCR4 binding to its endogenous ligands might provide a potential therapeutic target for treating the disease.In order to identify a potential back-up to the clinical development candidate, N-(3-((3-(5-chlorothiophene-2-sulfonamido)-4-methoxy-1H-indazol-1-yl)methyl)benzyl)-2-hydroxy-2-methylpropanamide (6), a library of 2,3-dichlorobenzene sulfonamides were designed. Synthesis and evaluation of the 'hits' in the primary antagonist assay (GTPγS) led to the identification of three novel templates with good potency and ligand efficiency (LE). The benzimidazolone template, exemplified by 88 (2,3-dichloro-N-(6-methoxy-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl) benzenesulfonamide), demonstrated a good PK profile (low clearance, moderate to high oral bioavailability) in both rat and dog, and had better potency in the human whole blood (hWB) assay compared to the other two series, and hence was explored further. Investigation of substituents of the benzimidazolone series, and incorporation of nitrogen into the core led to the discovery of 239 (2,3-dichloro-N-(6-methoxy-2-oxo-2,3-dihydro-1H-imidazo[4,5-b]pyridin-5-yl)benzenesulfonamide), which had a pA2 value of 6.0 in the hWB assay and a much improved LE (0.43) compared to indazole 6 (LE = 0.30). The azabenzimidazolone 239 provides a new starting point for further lead optimisation to develop a novel allosteric CCR4 antagonist from which a potential oral asthma drug could emerge.A phenylpyrazole sulfonamide (N-(2-(1H-pyrazol-1-yl)phenyl)phenylsulfonamide) was also identified, where X-ray diffraction studies showed that one of the nitrogens of the pyrazole ring forms an intramolecular hydrogen bond with the sulfonamide NH and promotes a conformation that was thought to be preferred in the active site. A SAR study around the phenylpyrazole core provided pyridine analogue 115 (N-(3-(1H-pyrazol-1-yl)pyridin-2-yl)-5-chlorothiophene-2-sulfonamide) with very good potency, high LE (0.47) and excellent physicochemical properties (chromlogD7.4 = 2.7, MW = 340, solubility = 133 μg/mL). Pyridylpyrazole 115 also represent good starting points for a lead optimisation programme.In addition, a novel series of 2,8-diazaspiro[4.5]decan-8-yl)pyrimidin-4-amine inhibitors, were identified using computational modelling studies, which bind to a different allosteric region on the CCR4 receptor compared to indazole 6. Evaluation of SAR led to identification of several analogues that exhibited hWB activity superior or comparable to that of 6. The most potent compound, 282 (N-(2,4-dichlorobenzyl)-2-(2-(pyrrolidin-2-ylmethyl)-2,8-diazaspiro[4.5]decan-8-yl)pyrimidin-4-amine, hWB pA2 = 6.7) demonstrated anti-inflammatory activity in acute in vivo murine allergic inflammation models. After an ovalbumin challenge in mice, compound 282 showed not only significant reduction in eosinophil and lymphocyte infiltration into the bronchoalveolar lavage (BAL) fluid but also displayed a profound effect on airway hyperreactivity, returning the levels back to the control line. The in vivo results indicate that targeting CCR4 with a small molecule allosteric antagonist could be a potential novel way of treating allergic diseases such as asthma. Interestingly, representative compounds from the spiro-pyrimidine series were found to induce receptor internalisation of CCR4 in HUT78 cell with approximately 50% reduction of cell surface receptor. It is suggested that in vivo activity of 282 might be due to its ability to induce internalisation of the receptor