ABSTRACT
The discovery of novel 4-hydroxy-2-(heterocyclic)pyrimidine-5-carboxamide inhibitors of hypoxia-inducible factor (HIF) prolyl hydroxylases (PHD) is described. These are potent, selective, orally bioavailable across several species, and active in stimulating erythropoiesis. Mouse and rat studies showed hematological changes with elevations of plasma EPO and circulating reticulocytes following single oral dose administration, while 4-week q.d. po administration in rat elevated hemoglobin levels. A major focus of the optimization process was to decrease the long half-life observed in higher species with early compounds. These efforts led to the identification of 28 (MK-8617), which has advanced to human clinical trials for anemia.
Subject(s)
Anemia/drug therapy , Drug Discovery , Enzyme Inhibitors/pharmacology , Hypoxia-Inducible Factor-Proline Dioxygenases/antagonists & inhibitors , Pyridazines/pharmacology , Pyrimidines/pharmacology , Administration, Oral , Anemia/enzymology , Animals , Dose-Response Relationship, Drug , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/chemistry , Humans , Hypoxia-Inducible Factor-Proline Dioxygenases/metabolism , Mice , Mice, Inbred C57BL , Models, Molecular , Molecular Structure , Pyridazines/administration & dosage , Pyridazines/chemistry , Pyrimidines/administration & dosage , Pyrimidines/chemistry , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipABSTRACT
Hit-to-lead efforts resulted in the discovery of compound 19, a potent CYP11B2 inhibitor that displays high selectivity vs related CYPs, good pharmacokinetic properties in rat and rhesus, and lead-like physical properties. In a rhesus pharmacodynamic model, compound 19 displays robust, dose-dependent aldosterone lowering efficacy, with no apparent effect on cortisol levels.
ABSTRACT
We report the discovery of a benzimidazole series of CYP11B2 inhibitors. Hit-to-lead and lead optimization studies identified compounds such as 32, which displays potent CYP11B2 inhibition, high selectivity versus related CYP targets, and good pharmacokinetic properties in rat and rhesus. In a rhesus pharmacodynamic model, 32 produces dose-dependent aldosterone lowering efficacy, with no apparent effect on cortisol levels.
ABSTRACT
We report herein the identification of MK-4409, a potent and selective fatty acid amide hydrolase (FAAH) inhibitor. Starting from a high throughput screening (HTS) hit, medicinal chemistry efforts focused on optimizing of FAAH inhibition in vitro potency, improving the pharmacokinetic (PK) profile, and increasing in vivo efficacy in rodent inflammatory and neuropathic pain assays.
ABSTRACT
We report herein the discovery of a fatty acid amide hydrolase (FAAH) positron emission tomography (PET) tracer. Starting from a pyrazole lead, medicinal chemistry efforts directed toward reducing lipophilicity led to the synthesis of a series of imidazole analogues. Compound 6 was chosen for further profiling due to its appropriate physical chemical properties and excellent FAAH inhibition potency across species. [(11)C]-6 (MK-3168) exhibited good brain uptake and FAAH-specific signal in rhesus monkeys and is a suitable PET tracer for imaging FAAH in the brain.
ABSTRACT
The discovery of 1,3,8-triazaspiro[4.5]decane-2,4-diones (spirohydantoins) as a structural class of pan-inhibitors of the prolyl hydroxylase (PHD) family of enzymes for the treatment of anemia is described. The initial hit class, spirooxindoles, was identified through affinity selection mass spectrometry (AS-MS) and optimized for PHD2 inhibition and optimal PK/PD profile (short-acting PHDi inhibitors). 1,3,8-Triazaspiro[4.5]decane-2,4-diones (spirohydantoins) were optimized as an advanced lead class derived from the original spiroindole hit. A new set of general conditions for C-N coupling, developed using a high-throughput experimentation (HTE) technique, enabled a full SAR analysis of the spirohydantoins. This rapid and directed SAR exploration has resulted in the first reported examples of hydantoin derivatives with good PK in preclinical species. Potassium channel off-target activity (hERG) was successfully eliminated through the systematic introduction of acidic functionality to the molecular structure. Undesired upregulation of alanine aminotransferese (ALT) liver enzymes was mitigated and a robust on-/off-target margin was achieved. Spirohydantoins represent a class of highly efficacious, short-acting PHD1-3 inhibitors causing a robust erythropoietin (EPO) upregulation in vivo in multiple preclinical species. This profile deems spirohydantoins as attractive short-acting PHDi inhibitors with the potential for treatment of anemia.
Subject(s)
Anemia/drug therapy , Aza Compounds/chemical synthesis , Hydantoins/chemical synthesis , Hypoxia-Inducible Factor 1/metabolism , Procollagen-Proline Dioxygenase/antagonists & inhibitors , Spiro Compounds/chemical synthesis , Animals , Aza Compounds/pharmacokinetics , Aza Compounds/pharmacology , Dogs , ERG1 Potassium Channel , Erythropoietin/biosynthesis , Ether-A-Go-Go Potassium Channels/metabolism , High-Throughput Screening Assays , Humans , Hydantoins/pharmacokinetics , Hydantoins/pharmacology , Hypoxia-Inducible Factor-Proline Dioxygenases , Indoles/chemical synthesis , Indoles/pharmacokinetics , Indoles/pharmacology , Liver/drug effects , Liver/enzymology , Macaca mulatta , Mass Spectrometry , Mice , Mice, Inbred C57BL , Protein Binding , Rats , Spiro Compounds/pharmacokinetics , Spiro Compounds/pharmacology , Structure-Activity Relationship , Up-RegulationABSTRACT
Structure-activity relationship (SAR) studies of 3-arylpropionic acids-a class of novel S1P(1) selective agonists-by introducing substitution to the propionic acid chain and replacing the adjacent phenyl ring with pyridine led to a series of modified 3-arylpropionic acids with enhanced half-life in rat. These analogs (e.g., cyclopropanecarboxylic acids) exhibited longer half-life in rat than did unmodified 3-arylpropionic acids. This result suggests that metabolic oxidation at the propionic acid chain, particularly at the C3 benzylic position of 3-arylpropionic acids, is probably responsible for their short half-life in rodent.
Subject(s)
Propionates/chemical synthesis , Propionates/pharmacology , Receptors, Lysosphingolipid/agonists , Animals , Biological Availability , CHO Cells , Cricetinae , Cricetulus , Guanosine 5'-O-(3-Thiotriphosphate)/metabolism , Half-Life , Lymphocyte Count , Lymphocytes/drug effects , Lymphocytes/metabolism , Male , Mice , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipABSTRACT
Novel series of sphingosine-1-phosphate (S1P) receptor agonists were developed through a systematic SAR aimed to achieve high selectivity for a single member of the S1P family of receptors, S1P1. The optimized structure represents a highly S1P1-selective and efficacious agonist: S1P1/S1P2, S1P1/S1P3, S1P1/S1P4>10,000-fold, S1P1/S1P5>600-fold, while EC50 (S1P1) <0.2 nM. In vivo experiments are consistent with S1P1 receptor agonism alone being sufficient for achieving desired lymphocyte-lowering effect.
Subject(s)
Immunosuppressive Agents/chemical synthesis , Immunosuppressive Agents/pharmacology , Lymphocytes/drug effects , Receptors, Lysosphingolipid/agonists , Animals , CHO Cells , Cricetinae , Lymphocyte Count , Lymphocytes/cytology , Structure-Activity RelationshipABSTRACT
A series of 3-arylpropionic acids were synthesized as S1P1 receptor agonists. Structure-activity relationship studies on the pendant phenyl ring revealed several structural features offering selectivity of S1P1 binding against S1P2-5. These highly selective S1P1 agonists induced peripheral blood lymphocyte lowering in mice and one of them was found to be efficacious in a rat skin transplantation model, supporting that S1P1 agonism is primarily responsible for the immunosuppressive efficacy observed in preclinical animal models.
Subject(s)
Immunosuppressive Agents/pharmacology , Phenylpropionates/chemical synthesis , Phenylpropionates/pharmacology , Receptors, Lysosphingolipid/agonists , Animals , CHO Cells , Cricetinae , Ligands , Lymphocyte Count/veterinary , Mice , Rats , Skin Transplantation , Structure-Activity RelationshipABSTRACT
A series of 2,5-cis-disubstituted pyrrolidines were synthesized and evaluated as S1P receptor agonists. Compounds 15-21 were identified with good selectivity over S1P3 which lowered circulating lymphocytes after oral administration in mice.
Subject(s)
Carboxylic Acids/chemistry , Carboxylic Acids/pharmacology , Pyrrolidines/chemistry , Receptors, Lysosphingolipid/agonists , Administration, Oral , Animals , Carboxylic Acids/chemical synthesis , Molecular Structure , Rats , Receptors, Lysosphingolipid/metabolism , Structure-Activity RelationshipABSTRACT
A series of 2-aryl(pyrrolidin-4-yl)acetic acids were synthesized and their biological activities were evaluated as agonists of S1P receptors. These analogs were able to induce lowering of lymphocyte counts in the peripheral blood of mice and were found to have good overall pharmacokinetic properties in rat.
Subject(s)
Acetates/pharmacology , Pyrrolidines/pharmacology , Receptors, Lysosphingolipid/agonists , Acetates/chemical synthesis , Acetates/chemistry , Animals , Lymphocyte Count , Lysophospholipids/antagonists & inhibitors , Mice , Molecular Structure , Protein Binding/drug effects , Pyrrolidines/chemical synthesis , Pyrrolidines/chemistry , Rats , Sphingosine/analogs & derivatives , Sphingosine/antagonists & inhibitors , Stereoisomerism , Structure-Activity RelationshipABSTRACT
A class of 3,5-diphenyl-1,2,4-oxadiazole based compounds have been identified as potent sphingosine-1-phosphate-1 (S1P1) receptor agonists with minimal affinity for the S1P2 and S1P3 receptor subtypes. Analogue 26 (S1P1 IC50 = 0.6 nM) has an excellent pharmacokinetics profile in the rat and dog and is efficacious in a rat skin transplant model, indicating that S1P3 receptor agonism is not a component of immunosuppressive efficacy.
Subject(s)
Immunosuppressive Agents/chemical synthesis , Oxadiazoles/chemical synthesis , Receptors, Lysosphingolipid/agonists , Animals , CHO Cells , Cricetinae , Cricetulus , Dogs , Graft Survival , Immunosuppressive Agents/pharmacokinetics , Immunosuppressive Agents/pharmacology , Lymphocyte Count , Oxadiazoles/pharmacokinetics , Oxadiazoles/pharmacology , Radioligand Assay , Rats , Rats, Inbred Lew , Rats, Sprague-Dawley , Skin Transplantation , Structure-Activity RelationshipABSTRACT
FTY720 is an immunosuppressive agent that modulates lymphocyte trafficking. It is phosphorylated in vivo to FTY720-phosphate (FTY-P) and binds to a family of G protein-coupled receptors recognizing sphingosine 1-phosphate (S1P) as the natural ligand. It has previously been reported that FTY-P blocks egress of lymphocytes from the thymus and lymph nodes, resulting in peripheral blood lymphopenia. We now report that FTY-P also causes displacement of marginal zone (MZ) B cells to the splenic follicles, an effect that is similar to that observed after in vivo administration of lipopolysaccharide. This effect is specific to B cells in the MZ, as treatment with FTY-P does not cause redistribution of the resident macrophage population. A small but statistically significant decrease in the expression of beta1 integrin on MZ B cells was observed with FTY-P treatment. The redistribution of MZ B cells from the MZ sinuses does not abolish the ability of these cells to respond to the T-independent antigen, trinitrophenol-Ficoll. It has been proposed that the displacement of MZ B cells to the follicles is an indication of cell activation. Consistent with this, FTY-P caused an increase in percentage of MZ B cells expressing activation markers CD9, CD1d, and CD24. These results suggest that S1P receptors on MZ B cells are responsible for their mobilization to follicles.
Subject(s)
B-Lymphocytes/physiology , Cell Movement/drug effects , Germinal Center/metabolism , Immunosuppressive Agents/administration & dosage , Propylene Glycols/administration & dosage , Receptors, Lysosphingolipid/antagonists & inhibitors , Animals , Antigens, CD/biosynthesis , B-Lymphocytes/cytology , Cell Movement/physiology , Female , Fingolimod Hydrochloride , Gene Expression Regulation/drug effects , Gene Expression Regulation/physiology , Germinal Center/cytology , Injections, Intraperitoneal , Lipopolysaccharides/administration & dosage , Lymphocyte Activation/drug effects , Lymphocyte Activation/physiology , Lymphoid Tissue/cytology , Lymphoid Tissue/metabolism , Macrophages/cytology , Macrophages/metabolism , Mice , Receptors, Lysosphingolipid/metabolism , Sphingosine/analogs & derivativesABSTRACT
Moderately potent, selective S1P(1) receptor agonists identified from high-throughput screening have been adapted into lipophilic tails for a class of orally bioavailable amino acid-based S1P(1) agonists represented by 7. Many of the new compounds are potent S1P(1) agonists that select against the S1P(2), S1P(3), and S1P(4) (although not S1P(5)) receptor subtypes. Analogues 18 and 24 are highly orally bioavailable and possess excellent pharmacokinetic profiles in the rat, dog, and rhesus monkey.
Subject(s)
Azetidines/pharmacology , Immunosuppressive Agents/pharmacology , Receptors, Lysosphingolipid/agonists , Administration, Oral , Animals , Azetidines/pharmacokinetics , Biological Availability , CHO Cells , Cricetinae , Dogs , Drug Design , Guanosine 5'-O-(3-Thiotriphosphate)/metabolism , Humans , Immunosuppressive Agents/pharmacokinetics , Lymphocytes/drug effects , Macaca mulatta , Mice , Rats , Rats, Inbred Lew , Structure-Activity RelationshipABSTRACT
Sphingosine-1-phosphate (S1P), a lipid signaling molecule that regulates many cellular functions, is synthesized from sphingosine and ATP by the action of sphingosine kinase. Two such kinases have been identified, SPHK1 and SPHK2. To begin to investigate the physiological functions of sphingosine kinase and S1P signaling, we generated mice deficient in SPHK1. Sphk1 null mice were viable, fertile, and without any obvious abnormalities. Total SPHK activity in most Sphk1-/-tissues was substantially, but not completely, reduced indicating the presence of multiple sphingosine kinases. S1P levels in most tissues from the Sphk1-/- mice were not markedly decreased. In serum, however, there was a significant decrease in the S1P level. Although S1P signaling regulates lymphocyte trafficking, lymphocyte distribution was unaffected in lymphoid organs of Sphk1-/- mice. The immunosuppressant FTY720 was phosphorylated and elicited lymphopenia in the Sphk1 null mice showing that SPHK1 is not required for the functional activation of this sphingosine analogue prodrug. The results with these Sphk1 null mice reveal that some key physiologic processes that require S1P receptor signaling, such as vascular development and proper lymphocyte distribution, can occur in the absence of SPHK1.
Subject(s)
Lymphopenia/etiology , Phosphotransferases (Alcohol Group Acceptor)/deficiency , Propylene Glycols/toxicity , Sphingosine/analogs & derivatives , Animals , Base Sequence , DNA/genetics , Fingolimod Hydrochloride , Immunosuppressive Agents/metabolism , Immunosuppressive Agents/toxicity , Lymphocytes/drug effects , Lymphocytes/physiology , Lymphopenia/chemically induced , Lymphopenia/enzymology , Lysophospholipids/metabolism , Mice , Mice, Knockout , Phosphorylation , Phosphotransferases (Alcohol Group Acceptor)/genetics , Phosphotransferases (Alcohol Group Acceptor)/physiology , Prodrugs/metabolism , Prodrugs/toxicity , Propylene Glycols/metabolism , Receptors, Lysosphingolipid/metabolism , Signal Transduction , Sphingosine/metabolismABSTRACT
A series of conformationally constrained 3-(N-alkylamino)propylphosphonic acids were systematically synthesized and their activities as S1P receptor agonists were evaluated. Several pyrrolidine and cyclohexane analogs had S1P receptor profiles comparable to the acyclic lead compound, 3-(N-tetradecylamino)propylphosphonic acid (3), lowered circulating lymphocytes in mice after iv administration and were thus identified as being suitable for further investigations.
Subject(s)
Drug Design , Organophosphonates/chemical synthesis , Receptors, Lysosphingolipid/agonists , Animals , CHO Cells , Cricetinae , Humans , Molecular Conformation , Organophosphonates/chemistry , Organophosphonates/pharmacology , Structure-Activity RelationshipABSTRACT
The novel immunosuppressive agent FTY720 (1) is phosphorylated in vivo in a variety of species yielding an active metabolite that is an agonist of four of the five known G-protein-coupled sphingosine-1-phosphate (S1P) receptors. A synthesis amenable to producing gram quantities of the stereoisomeric phosphate esters, a determination of their absolute stereochemistry via an enantioselective synthesis and their characterization as S1P receptor agonists and antagonists is reported.
Subject(s)
Immunosuppressive Agents/chemical synthesis , Organophosphates/chemical synthesis , Propylene Glycols/chemical synthesis , Animals , CHO Cells , Cricetinae , Fingolimod Hydrochloride , Humans , Molecular Conformation , Sphingosine/analogs & derivativesABSTRACT
3-(N-Alkyl)aminopropylphosphonic acids are potent agonists of four of the five known sphingosine-1-phosphate (S1P) receptor subtypes.
Subject(s)
Receptors, G-Protein-Coupled/agonists , Receptors, Lysosphingolipid/agonists , Animals , CHO Cells , Cricetinae , Humans , Inhibitory Concentration 50 , Ligands , Organophosphonates/chemical synthesis , Organophosphorus Compounds/chemical synthesis , Phosphorus Radioisotopes , Structure-Activity RelationshipABSTRACT
Structurally modified 3-(N-benzylamino)propylphosphonic acid S1P receptor agonists that maintain affinity for S1P1, and have decreased affinity for S1P3 are efficacious, but exhibit decreased acute cardiovascular toxicity in rodents than do nonselective agonists.
Subject(s)
Benzyl Compounds/pharmacology , Cardiovascular System/drug effects , Myocardium/metabolism , Organophosphorus Compounds/pharmacology , Receptors, G-Protein-Coupled/agonists , Receptors, Lysosphingolipid/agonists , Receptors, Lysosphingolipid/antagonists & inhibitors , Animals , Benzyl Compounds/chemistry , Binding Sites , CHO Cells , Cricetinae , Humans , Inhibitory Concentration 50 , Kinetics , Lysophospholipids/chemistry , Lysophospholipids/pharmacology , Mice , Organophosphorus Compounds/chemistry , Phosphorus Radioisotopes , Rats , Receptors, G-Protein-Coupled/metabolism , Sphingosine/analogs & derivatives , Sphingosine/chemistry , Sphingosine/pharmacologyABSTRACT
Alteration in lymphocyte trafficking and prevention of graft rejection in rodents observed on exposure to FTY720 (1) or its corresponding phosphate ester 2 can be induced by the systemic administration of potent sphingosine-1-phosphate receptor agonists exemplified by 19. The similar S1P receptor profiles of 2 and 19 coupled with their comparable potency in vivo supports a connection between S1P receptor agonism and immunosuppressive efficacy.