ABSTRACT
Interleukin-1 receptor associated kinase 4 (IRAK4) is an essential mediator of the IL-1R and TLR signaling pathways, both of which have been implicated in multiple autoimmune conditions. Hence, blocking the activity of IRAK4 represents an attractive approach for the treatment of autoimmune diseases. The activity of this serine/threonine kinase is dependent on its kinase and scaffolding activities; thus, degradation represents a potentially superior approach to inhibition. Herein, we detail the exploration of structure-activity relationships that ultimately led to the identification of KT-474, a potent, selective, and orally bioavailable heterobifunctional IRAK4 degrader. This represents the first heterobifunctional degrader evaluated in a nononcology indication and dosed to healthy human volunteers. This molecule successfully completed phase I studies in healthy adult volunteers and patients with atopic dermatitis or hidradenitis suppurativa. Phase II clinical trials in both of these indications have been initiated.
ABSTRACT
Developing therapies for the activated B-cell like (ABC) subtype of diffuse large B-cell lymphomas (DLBCL) remains an area of unmet medical need. A subset of ABC DLBCL tumors is driven by activating mutations in myeloid differentiation primary response protein 88 (MYD88), which lead to constitutive activation of interleukin-1 receptor associated kinase 4 (IRAK4) and cellular proliferation. IRAK4 signaling is driven by its catalytic and scaffolding functions, necessitating complete removal of this protein and its escape mechanisms for complete therapeutic suppression. Herein, we describe the identification and characterization of a dual-functioning molecule, KT-413 and show it efficiently degrades IRAK4 and the transcription factors Ikaros and Aiolos. KT-413 achieves concurrent degradation of these proteins by functioning as both a heterobifunctional degrader and a molecular glue. Based on the demonstrated activity and safety of KT-413 in preclinical studies, a phase 1 clinical trial in B-cell lymphomas, including MYD88 mutant ABC DLBCL, is currently underway.
Subject(s)
Interleukin-1 Receptor-Associated Kinases , Lymphoma, Large B-Cell, Diffuse , Mutation , Myeloid Differentiation Factor 88 , Interleukin-1 Receptor-Associated Kinases/metabolism , Interleukin-1 Receptor-Associated Kinases/antagonists & inhibitors , Myeloid Differentiation Factor 88/metabolism , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/genetics , Lymphoma, Large B-Cell, Diffuse/metabolism , Lymphoma, Large B-Cell, Diffuse/pathology , Humans , Animals , Cell Line, Tumor , Drug Discovery , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/therapeutic use , Mice , Imidazoles/chemistry , Imidazoles/pharmacology , Imidazoles/metabolism , Proteolysis/drug effects , Structure-Activity RelationshipABSTRACT
Toll-like receptor-driven and interleukin-1 (IL-1) receptor-driven inflammation mediated by IL-1 receptor-associated kinase 4 (IRAK4) is involved in the pathophysiology of hidradenitis suppurativa (HS) and atopic dermatitis (AD). KT-474 (SAR444656), an IRAK4 degrader, was studied in a randomized, double-blind, placebo-controlled phase 1 trial where the primary objective was safety and tolerability. Secondary objectives included pharmacokinetics, pharmacodynamics and clinical activity in patients with moderate to severe HS and in patients with moderate to severe AD. KT-474 was administered as a single dose and then daily for 14 d in 105 healthy volunteers (HVs), followed by dosing for 28 d in an open-label cohort of 21 patients. Degradation of IRAK4 was observed in HV blood, with mean reductions after a single dose of ≥93% at 600-1,600 mg and after 14 daily doses of ≥95% at 50-200 mg. In patients, similar IRAK4 degradation was achieved in blood, and IRAK4 was normalized in skin lesions where it was overexpressed relative to HVs. Reduction of disease-relevant inflammatory biomarkers was demonstrated in the blood and skin of patients with HS and patients with AD and was associated with improvement in skin lesions and symptoms. There were no drug-related infections. These results, from what, to our knowledge, is the first published clinical trial using a heterobifunctional degrader, provide initial proof of concept for KT-474 in HS and AD to be further confirmed in larger trials. ClinicalTrials.gov identifier: NCT04772885 .
Subject(s)
Dermatitis, Atopic , Hidradenitis Suppurativa , Humans , Hidradenitis Suppurativa/drug therapy , Dermatitis, Atopic/drug therapy , Interleukin-1 Receptor-Associated Kinases , Treatment Outcome , Skin/pathology , Double-Blind Method , Severity of Illness IndexABSTRACT
Quantitative understanding about the dynamics of drug-target interactions in biological systems is essential, especially in rare disease programs with small patient populations. Follistatin, by antagonism of myostatin and activin, which are negative regulators of skeletal muscle and inflammatory response, is a promising therapeutic target for Duchenne Muscular Dystrophy. In this study, we constructed a quantitative systems pharmacology model for FS-EEE-Fc, a follistatin recombinant protein to investigate its efficacy from dual target binding, and, subsequently, to project its human efficacious dose. Based on model simulations, with an assumed efficacy threshold of 7-10% muscle volume increase, 3-5 mg/kg weekly dosing of FS-EEE-Fc is predicted to achieve meaningful clinical outcome. In conclusion, the study demonstrated an application of mechanism driven approach at early stage of a rare disease drug development to support lead compound optimization, enable human dose, pharmacokinetics, and efficacy predictions.
Subject(s)
Follistatin/pharmacokinetics , Models, Biological , Muscle, Skeletal/drug effects , Muscular Dystrophy, Duchenne/drug therapy , Neuromuscular Agents/pharmacokinetics , Systems Biology , Activin Receptors, Type II/antagonists & inhibitors , Activin Receptors, Type II/metabolism , Animals , Drug Dosage Calculations , Follistatin/administration & dosage , Humans , Ligands , Muscle, Skeletal/growth & development , Muscle, Skeletal/metabolism , Muscular Dystrophy, Duchenne/metabolism , Muscular Dystrophy, Duchenne/physiopathology , Myostatin/antagonists & inhibitors , Myostatin/metabolism , Neuromuscular Agents/administration & dosage , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/pharmacokinetics , Signal TransductionABSTRACT
BACKGROUND: Myostatin antagonists are being developed as therapies for Duchenne muscular dystrophy due to their strong hypertrophic effects on skeletal muscle. Engineered follistatin has the potential to combine the hypertrophy of myostatin antagonism with the anti-inflammatory and anti-fibrotic effects of activin A antagonism. METHODS: Engineered follistatin was administered to C57BL/6 mice for 4 weeks, and muscle mass and myofiber size was measured. In the mdx model, engineered follistatin was dosed for 12 weeks in two studies comparing to an Fc fusion of the activin IIB receptor or an anti-myostatin antibody. Functional measurements of grip strength and tetanic force were combined with tissue analysis for markers of necrosis, inflammation, and fibrosis to evaluate improvement in dystrophic pathology. RESULTS: In wild-type and mdx mice, dose-dependent increases in muscle mass and quadriceps myofiber size were observed for engineered follistatin. In mdx, increases in grip strength and tetanic force were combined with improvements in muscle markers for necrosis, inflammation, and fibrosis. Improvements in dystrophic pathology were greater for engineered follistatin than the anti-myostatin antibody. CONCLUSIONS: Engineered follistatin generated hypertrophy and anti-fibrotic effects in the mdx model.
Subject(s)
Activins/antagonists & inhibitors , Follistatin/therapeutic use , Muscular Dystrophies/drug therapy , Myostatin/antagonists & inhibitors , Animals , Follistatin/administration & dosage , Hand Strength , Male , Mice , Mice, Inbred C57BL , Mice, Inbred mdx , Muscle Contraction , Recombinant Proteins/administration & dosage , Recombinant Proteins/therapeutic useABSTRACT
Kynurenic acid (KYNA) plays a significant role in maintaining normal brain function, and abnormalities in KYNA levels have been associated with various central nervous system disorders. Confirmation of its causality in human diseases requires safe and effective modulation of central KYNA levels in the clinic. The kynurenine aminotransferases (KAT) II enzyme represents an attractive target for pharmacologic modulation of central KYNA levels; however, KAT II and KYNA turnover kinetics, which could contribute to the duration of pharmacologic effect, have not been reported. In this study, the kinetics of central KYNA-lowering effect in rats and nonhuman primates (NHPs, Cynomolgus macaques) was investigated using multiple KAT II irreversible inhibitors as pharmacologic probes. Mechanistic pharmacokinetic-pharmacodynamic analysis of in vivo responses to irreversible inhibition quantitatively revealed that 1) KAT II turnover is relatively slow [16-76 hours' half-life (t1/2)], whereas KYNA is cleared more rapidly from the brain (<1 hour t1/2) in both rats and NHPs, 2) KAT II turnover is slower in NHPs than in rats (76 hours vs. 16 hours t1/2, respectively), and 3) the percent contribution of KAT II to KYNA formation is constant (â¼80%) across rats and NHPs. Additionally, modeling results enabled establishment of in vitro-in vivo correlation for both enzyme turnover rates and drug potencies. In summary, quantitative translational analysis confirmed the feasibility of central KYNA modulation in humans. Model-based analysis, where system-specific properties and drug-specific properties are mechanistically separated from in vivo responses, enabled quantitative understanding of the KAT II-KYNA pathway, as well as assisted development of promising candidates to test KYNA hypothesis in humans.
Subject(s)
Brain/metabolism , Enzyme Inhibitors/administration & dosage , Kynurenic Acid/analysis , Transaminases/metabolism , Animals , Brain Chemistry/drug effects , Cells, Cultured , Chromatography, Liquid , Enzyme Inhibitors/pharmacology , Female , Half-Life , Humans , Macaca fascicularis , Male , Pyrazoles/administration & dosage , Pyrazoles/pharmacology , Rats , Tandem Mass Spectrometry , Transaminases/antagonists & inhibitorsABSTRACT
Follistatin (FS) is an important regulatory protein, a natural antagonist for transforming growth factor-ß family members activin and myostatin. The diverse biologic roles of the activin and myostatin signaling pathways make FS a promising therapeutic target for treating human diseases exhibiting inflammation, fibrosis, and muscle disorders, such as Duchenne muscular dystrophy. However, rapid heparin-mediated hepatic clearance of FS limits its therapeutic potential. We targeted the heparin-binding loop of FS for site-directed mutagenesis to improve clearance parameters. By generating a series of FS variants with one, two, or three negative amino acid substitutions, we demonstrated a direct and proportional relationship between the degree of heparin-binding affinity in vitro and the exposure in vivo. The triple mutation K(76,81,82)E abolished heparin-binding affinity, resulting in â¼20-fold improved in vivo exposure. This triple mutant retains full functional activity and an antibody-like pharmacokinetic profile, and shows a superior developability profile in physical stability and cell productivity compared with FS variants, which substitute the entire heparin-binding loop with alternative sequences. Our surgical approach to mutagenesis should also reduce the immunogenicity risk. To further lower this risk, we introduced a novel glycosylation site into the heparin-binding loop. This hyperglycosylated variant showed a 10-fold improved exposure and decreased clearance in mice compared with an IgG1 Fc fusion protein containing the native FS sequence. Collectively, our data highlight the importance of improving pharmacokinetic properties by manipulating heparin-binding affinity and glycosylation content and provide a valuable guideline to design desirable therapeutic FS molecules.
Subject(s)
Follistatin/genetics , Follistatin/pharmacokinetics , Protein Engineering , Recombinant Proteins/genetics , Recombinant Proteins/pharmacokinetics , Amino Acid Motifs , Amino Acid Sequence , Animals , Female , Follistatin/metabolism , Follistatin/therapeutic use , Glycosylation , Heparin/metabolism , Humans , Mice , Mutation , Recombinant Proteins/metabolism , Recombinant Proteins/therapeutic use , Tissue DistributionABSTRACT
This work provides a perspective on the qualification and verification of physiologically based pharmacokinetic (PBPK) platforms/models intended for regulatory submission based on the collective experience of the Simcyp Consortium members. Examples of regulatory submission of PBPK analyses across various intended applications are presented and discussed. European Medicines Agency (EMA) and US Food and Drug Administration (FDA) recent draft guidelines regarding PBPK analyses and reporting are encouraging, and to advance the use and acceptability of PBPK analyses, more clarity and flexibility are warranted.
Subject(s)
Computer Simulation , Drug Approval , Models, Biological , Pharmacokinetics , Europe , Humans , United States , United States Food and Drug AdministrationABSTRACT
BACKGROUND: Guanfacine extended-release (GXR) is an orally administered, non-stimulant treatment for children and adolescents with attention-deficit/hyperactivity disorder (ADHD) and is primarily metabolized by the 3A4 isozyme of cytochrome P450 (CYP3A4). The results of clinical pharmacokinetic (PK) studies indicate that guanfacine is sensitive to drug-drug interactions (DDIs) perpetrated by strong inhibitors and inducers of CYP3A4. OBJECTIVE: The aim was to provide guidance on the possible requirement for GXR dose adjustment in children and adolescents with ADHD by predicting DDIs following co-administration with moderate CYP3A4 inhibitors and inducers. METHODS: A physiologically based PK model for GXR orally administered to healthy adults was developed based on physicochemical, in vitro and clinical PK data. The model was validated using clinical PK data for co-administration of GXR with ketoconazole (strong CYP3A4 inhibitor) or rifampicin (strong CYP3A4 inducer). RESULTS: Model predictions indicated that co-administration of GXR with the moderate CYP3A4 inhibitors erythromycin 500 mg three times a day or fluconazole 200 mg daily (q.d.) increased the guanfacine area under the plasma concentration-time curve (AUC) by 2.31-fold or 1.98-fold, respectively, compared with GXR monotherapy. The moderate CYP3A4 inducer efavirenz 400 mg or 600 mg q.d. was predicted to reduce guanfacine AUC to 58 or 33% of its value for GXR monotherapy, respectively. CONCLUSION: Without the requirement for additional clinical studies, the following GXR dose recommendations were developed and approved for US labeling for use in children and adolescents with ADHD: (1) decrease GXR to 50% of the usual target dose when it is co-administered with strong or moderate CYP3A4 inhibitors; (2) consider titrating GXR up to double the usual target dose over 1-2 weeks when it is co-administered with strong or moderate CYP3A4 inducers.
Subject(s)
Attention Deficit Disorder with Hyperactivity/drug therapy , Cytochrome P-450 CYP3A Inducers/administration & dosage , Cytochrome P-450 CYP3A Inhibitors/administration & dosage , Cytochrome P-450 CYP3A/metabolism , Guanfacine/administration & dosage , Adolescent , Adrenergic alpha-2 Receptor Agonists/therapeutic use , Alkynes , Anti-Retroviral Agents/pharmacokinetics , Antifungal Agents/pharmacokinetics , Benzoxazines/pharmacokinetics , Child , Cyclopropanes , Cytochrome P-450 CYP3A Inducers/pharmacokinetics , Cytochrome P-450 CYP3A Inhibitors/pharmacokinetics , Delayed-Action Preparations , Drug Interactions , Erythromycin/pharmacokinetics , Fluconazole/pharmacokinetics , Guanfacine/pharmacokinetics , Humans , Models, BiologicalABSTRACT
The article "Development of Guanfacine Extended-Release Dosing Strategies in Children and Adolescents with ADHD Using a PhysiologicallyBased Pharmacokinetic Model to Predict Drug-Drug Interactions with Moderate CYP3A4 Inhibitors or Inducers", written by Aiqun Li, Karen Yeo, Devin Welty, Haojing Rong, was originally published electronically on the publisher's internet portal (currently SpringerLink) on 02nd November, 2017 without open access.
ABSTRACT
C5-substituted 2,4-diaminoquinazoline inhibitors of the decapping scavenger enzyme DcpS (DAQ-DcpSi) have been developed for the treatment of spinal muscular atrophy (SMA), which is caused by genetic deficiency in the Survival Motor Neuron (SMN) protein. These compounds are claimed to act as SMN2 transcriptional activators but data underlying that claim are equivocal. In addition it is unclear whether the claimed effects on SMN2 are a direct consequence of DcpS inhibitor or might be a consequence of lysosomotropism, which is known to be neuroprotective. DAQ-DcpSi effects were characterized in cells in vitro utilizing DcpS knockdown and 7-methyl analogues as probes for DcpS vs non-DcpS-mediated effects. We also performed analysis of Smn transcript levels, RNA-Seq analysis of the transcriptome and SMN protein in order to identify affected pathways underlying the therapeutic effect, and studied lysosomotropic and non-lysosomotropic DAQ-DCpSi effects in 2B/- SMA mice. Treatment of cells caused modest and transient SMN2 mRNA increases with either no change or a decrease in SMNΔ7 and no change in SMN1 transcripts or SMN protein. RNA-Seq analysis of DAQ-DcpSi-treated N2a cells revealed significant changes in expression (both up and down) of approximately 2,000 genes across a broad range of pathways. Treatment of 2B/- SMA mice with both lysomotropic and non-lysosomotropic DAQ-DcpSi compounds had similar effects on disease phenotype indicating that the therapeutic mechanism of action is not a consequence of lysosomotropism. In striking contrast to the findings in vitro, Smn transcripts were robustly changed in tissues but there was no increase in SMN protein levels in spinal cord. We conclude that DAQ-DcpSi have reproducible benefit in SMA mice and a broad spectrum of biological effects in vitro and in vivo, but these are complex, context specific, and not the result of simple SMN2 transcriptional activation.
Subject(s)
Endoribonucleases/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Muscular Atrophy, Spinal/drug therapy , Muscular Atrophy, Spinal/enzymology , Quinazolines/pharmacology , Animals , Cell Line , Disease Models, Animal , Enzyme Inhibitors/chemistry , Female , Gene Knockdown Techniques , HEK293 Cells , Humans , Male , Mice , Mice, Knockout , Muscular Atrophy, Spinal/genetics , Promoter Regions, Genetic , Quinazolines/chemistry , RNA, Messenger/genetics , RNA, Messenger/metabolism , Survival of Motor Neuron 2 Protein/deficiency , Survival of Motor Neuron 2 Protein/genetics , Survival of Motor Neuron 2 Protein/metabolismABSTRACT
The C-5 substituted 2,4-diaminoquinazoline RG3039 (compound 1), a member of a chemical series that was identified and optimized using an SMN2 promoter screen, prolongs survival and improves motor function in a mouse model of spinal muscular atrophy (SMA). It is a potent inhibitor of the mRNA decapping scavenger enzyme (DcpS), but the mechanism whereby DcpS inhibition leads to therapeutic benefit is unclear. Compound 1 is a dibasic lipophilic molecule that is predicted to accumulate in lysosomes. To understand if the in vivo efficacy is due to DcpS inhibition or other effects resulting from the physicochemical properties of the chemotype, we undertook structure based molecular design to identify DcpS inhibitors with improved physicochemical properties. Herein we describe the design, synthesis, and in vitro pharmacological characterization of these DcpS inhibitors along with the in vivo mouse CNS PK profile of PF-DcpSi (compound 24), one of the analogs found to be efficacious in SMA mouse model.
Subject(s)
Drug Design , Endoribonucleases/antagonists & inhibitors , Muscular Atrophy, Spinal/drug therapy , Quinazolines/chemistry , Quinazolines/therapeutic use , RNA, Messenger/antagonists & inhibitors , Animals , Disease Models, Animal , Endoribonucleases/genetics , Endoribonucleases/metabolism , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacokinetics , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/therapeutic use , HEK293 Cells , Humans , Mice , Molecular Docking Simulation , Muscular Atrophy, Spinal/genetics , Muscular Atrophy, Spinal/metabolism , Quinazolines/pharmacokinetics , Quinazolines/pharmacology , RNA, Messenger/genetics , Survival of Motor Neuron 2 ProteinABSTRACT
PURPOSE: Growth and differentiation factor 8 (GDF-8) is a negative regulator of skeletal muscle mass and targeted by inhibitors to treat diseases associated with muscle loss. In order to enable clinical and translational investigations of GDF-8 inhibitors, specific and sensitive measurements of GDF-8 are necessary. EXPERIMENTAL DESIGN: An immunoaffinity LC-MS/MS assay for quantification of GDF-8 in serum was developed, qualified and implemented. The workflow includes offline enrichment of GDF-8 using an anti-GDF-8 antibody, followed by isolation using magnetic beads, trypsin digestion, and quantification using 2D nanoflow LC-MS/MS. RESULTS: This assay was qualified in human serum with a lower LOQ of 1.0 ng/mL based on the intact protein. GDF-8 was quantified in serum from juvenile and adult humans as well as mouse, rat, and cynomolgus monkey. Additionally, the assay was utilized to demonstrate an increase of total GDF-8 in serum following administration of an anti-GDF-8 monoclonal antibody therapeutic in cynomolgus monkeys. CONCLUSIONS AND CLINICAL RELEVANCE: A specific and sensitive method was developed for the measurement of GDF-8 in juvenile and adult serum samples as well as preclinical species. The confident quantification of GDF-8 now enables a greater understanding of any association between changes GDF-8 levels and muscle mass.
Subject(s)
Antibodies, Monoclonal/chemistry , Biological Assay/standards , Myostatin/blood , Amino Acid Sequence , Animals , Chromatography, Affinity/methods , Chromatography, Liquid/methods , Gene Expression , Humans , Immunomagnetic Separation/methods , Macaca fascicularis , Mice , Myostatin/genetics , Peptides/chemical synthesis , Rats , Tandem Mass Spectrometry/methodsABSTRACT
Human plasma-derived α1-antitrypsin (AAT) delivered by intravenous infusion is used as augmentation therapy in patients with emphysema who have a genetic mutation resulting in deficiency of AAT. Inhalation is an alternative route of administration that can potentially increase the efficacy and convenience of treatment. This study was conducted to determine whether delivery to the lungs, initially via the intratracheal (IT) route of administration, would deliver efficacious levels of a recombinant AAT (rAAT) to the site of action in the lungs in mice. 125I-radiolabeled rAAT, fluorophore-conjugated rAAT (rAAT-Alexa488), and NE680 (neutrophil elastase 680, a silent fluorescent substrate of neutrophil elastase which fluoresces in the near-infrared range upon activation by neutrophil elastase) were used to characterize the pharmacokinetics and tissue distribution profile, distribution of rAAT within the lung, and efficacy of rAAT to inhibit neutrophil elastase at the site of action, respectively. The study has demonstrated that rAAT was able to gain access to locations where neutrophil elastase was localized. The histochemical quantification of rAAT activity relative to dose at the site of action provided here will improve confidence in predicting the human dose via the inhalation route.
ABSTRACT
Facilitating activation, or delaying inactivation, of the native Kv7 channel reduces neuronal excitability, which may be beneficial in controlling spontaneous electrical activity during epileptic seizures. In an effort to identify a compound with such properties, the structure-activity relationship (SAR) and in vitro ADME for a series of heterocyclic Kv7.2-7.5 channel openers was explored. PF-05020182 (2) demonstrated suitable properties for further testing in vivo where it dose-dependently decreased the number of animals exhibiting full tonic extension convulsions in response to corneal stimulation in the maximal electroshock (MES) assay. In addition, PF-05020182 (2) significantly inhibited convulsions in the MES assay at doses tested, consistent with in vitro activity measure. The physiochemical properties, in vitro and in vivo activities of PF-05020182 (2) support further development as an adjunctive treatment of refractory epilepsy.
Subject(s)
Drug Discovery , Epilepsy/drug therapy , Ion Channel Gating/drug effects , KCNQ2 Potassium Channel/metabolism , Piperidines/pharmacology , Pyrimidines/pharmacology , Animals , Cell Line , Dose-Response Relationship, Drug , Electroshock , Humans , KCNQ2 Potassium Channel/agonists , Microsomes/drug effects , Molecular Structure , Piperidines/administration & dosage , Piperidines/chemistry , Pyrimidines/administration & dosage , Pyrimidines/chemistry , Rats , Structure-Activity RelationshipABSTRACT
This Perspective provides important concepts about the blood-brain barrier (BBB) in drug discovery and how they should be applied effectively in designing successful CNS drugs. Key parameters for brain penetration are discussed, including unbound brain concentration, unbound brain-to-plasma ratio, BBB permeability, fraction unbound in brain and plasma, and transporters. Results from a retrospective analysis of 32 Pfizer CNS clinical drug candidates are described. Frequently encountered misconceptions about brain penetration in drug discovery programs are clarified. Strategies and guidance are provided to enhance or minimize brain exposure for CNS or peripheral targets, respectively. Recommendations for screening methodologies and a cascade in assessing brain penetration potential are presented.
Subject(s)
Brain/metabolism , Central Nervous System Agents/pharmacokinetics , ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , ATP Binding Cassette Transporter, Subfamily G, Member 2 , ATP-Binding Cassette Transporters/metabolism , Animals , Biological Transport , Blood Proteins/metabolism , Blood-Brain Barrier/metabolism , Central Nervous System Agents/blood , Central Nervous System Agents/cerebrospinal fluid , Drug Discovery , Humans , Models, Biological , Neoplasm Proteins/metabolism , Permeability , Protein BindingABSTRACT
Positive allosteric modulators ('potentiators') of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) have been shown to display a mechanism-based exposure-response continuum in preclinical species with procognitive electrophysiological and behavioral effects ('efficacy') at low exposures and motor coordination disruptions at progressively higher exposures. Due to the dose-capping nature of such motor coordination deficits, an exposure threshold-mediated adverse event (C(AE) ), the adequacy of separation between the maximal total plasma compound concentration (C(max) ) at a predicted clinically efficacious oral dose and this adverse event (AE) was explored in early drug research with three AMPAR potentiators considered potential candidates for clinical trials. In vitro metabolism studies in human liver microsomes and human hepatocytes demonstrated the metabolic clearance for each compound was predominately due to cytochromes P450 (CYP). Thus, for each compound's anticipated clinically efficacious dose, human C(max) variability following oral administration was assessed using Simcyp software, which combines its virtual human populations database using extensive demographic, physiological and genomic information with routinely collected compound-specific in vitro biochemical data to simulate and predict drug disposition. Using a combination of experimentally determined recombinant human CYP intrinsic clearances for CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A4, human binding factors, expected fraction absorbed and estimated steady-state volume of distribution, Simcyp simulations demonstrated that two of the three potentiators had acceptable projected C(max) variability (i.e. the 95th percentile C(max) did not breach C(AE) ). This evaluation aided in the selection of compounds for preclinical progression, and represents a novel application of pharmacologically based pharmacokinetic (PBPK) software approaches to predict interpatient variability.
Subject(s)
Drug Discovery/trends , Drug-Related Side Effects and Adverse Reactions/metabolism , Pharmaceutical Preparations/administration & dosage , Pharmaceutical Preparations/metabolism , Software/trends , Administration, Oral , Cytochrome P-450 Enzyme System/administration & dosage , Cytochrome P-450 Enzyme System/pharmacokinetics , Drug Discovery/methods , Drug-Related Side Effects and Adverse Reactions/diagnosis , Forecasting , Humans , Microsomes, Liver/drug effects , Microsomes, Liver/metabolism , Pharmacokinetics , Protein Binding/physiology , Receptors, AMPA/metabolismABSTRACT
The potent 5-lipoxygenase-activating protein (FLAP) inhibitor 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxypyridin-3-yl)benzyl]-5-(5-methylpyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethylpropionic acid 11cc is described (AM803, now GSK2190915). Building upon AM103 (1) (Hutchinson et al. J. Med Chem.2009, 52, 5803-5815; Stock et al. Bioorg. Med. Chem. Lett. 2010, 20, 213-217; Stock et al. Bioorg. Med. Chem. Lett.2010, 20, 4598-4601), SAR studies centering around the pyridine moiety led to the discovery of compounds that exhibit significantly increased potency in a human whole blood assay measuring LTB(4) inhibition with longer drug preincubation times (15 min vs 5 h). Further studies identified 11cc with a potency of 2.9 nM in FLAP binding, an IC(50) of 76 nM for inhibition of LTB(4) in human blood (5 h incubation) and excellent preclinical toxicology and pharmacokinetics in rat and dog. 11cc also demonstrated an extended pharmacodynamic effect in a rodent bronchoalveolar lavage (BAL) model. This compound has successfully completed phase 1 clinical studies in healthy volunteers and is currently undergoing phase 2 trials in asthmatic patients.
Subject(s)
5-Lipoxygenase-Activating Protein Inhibitors/chemical synthesis , Anti-Asthmatic Agents/chemical synthesis , Indoles/chemical synthesis , Pentanoic Acids/chemical synthesis , 5-Lipoxygenase-Activating Protein Inhibitors/pharmacokinetics , 5-Lipoxygenase-Activating Protein Inhibitors/pharmacology , Administration, Oral , Animals , Anti-Asthmatic Agents/pharmacokinetics , Anti-Asthmatic Agents/pharmacology , Bronchoalveolar Lavage , Cytochrome P-450 Enzyme Inhibitors , Dogs , Female , Humans , In Vitro Techniques , Indoles/pharmacokinetics , Indoles/pharmacology , Male , Pentanoic Acids/pharmacokinetics , Pentanoic Acids/pharmacology , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipABSTRACT
A series of potent 5-lipoxygenase-activating protein (FLAP) inhibitors are herein described. SAR studies focused on the discovery of novel alicyclic moieties appended to an indole core to optimize potency, physical properties and off-target activities. Subsequent SAR on the N-benzyl substituent of the indole led to the discovery of compound 39 (AM679) which showed potent inhibition of leukotrienes in human blood and in a rodent bronchoalvelolar lavage (BAL) challenge model.
Subject(s)
Carrier Proteins/antagonists & inhibitors , Indoles/chemistry , Lipoxygenase Inhibitors/chemistry , Membrane Proteins/antagonists & inhibitors , Pentanoic Acids/chemistry , 5-Lipoxygenase-Activating Proteins , Animals , Carrier Proteins/metabolism , Humans , Indoles/chemical synthesis , Indoles/pharmacology , Leukotrienes/blood , Leukotrienes/metabolism , Lipoxygenase Inhibitors/chemical synthesis , Lipoxygenase Inhibitors/pharmacology , Membrane Proteins/metabolism , Mice , Models, Animal , Pentanoic Acids/chemical synthesis , Pentanoic Acids/pharmacology , Rats , Structure-Activity RelationshipABSTRACT
The potent and selective 5-lipoxygenase-activating protein leukotriene synthesis inhibitor 3-[3-tert-butylsulfanyl-1-[4-(6-methoxy-pyridin-3-yl)-benzyl]-5-(pyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethyl-propionic acid (11j) is described. Lead optimization was designed to afford compounds with superior in vitro and in vivo inhibition of leukotriene synthesis in addition to having excellent pharmacokinetics and safety in rats and dogs. The key structural features of these new compounds are incorporation of heterocycles on the indole N-benzyl substituent and replacement of the quinoline group resulting in compounds with excellent in vitro and in vivo activities, superior pharmacokinetics, and improved physical properties. The methoxypyridine derivative 11j has an IC(50) of 4.2 nM in a 5-lipoxygenase-activating protein (FLAP) binding assay, an IC(50) of 349 nM in the human blood LTB(4) inhibition assay, and is efficacious in a murine ovalbumin model of allergen-induced asthma. Compound 11j was selected for clinical development and has successfully completed phase 1 trials in healthy volunteers.