Identification and optimisation of a pyrimidopyridone series of IRAK4 inhibitors.

In this article, we report the discovery of a series of pyrimidopyridones as inhibitors of IRAK4 kinase. From a previously disclosed 5-azaquinazoline series, we found that switching the pyridine ring for an N-substituted pyridone gave a novel hinge binding scaffold which retained potency against IRAK4. Importantly, introduction of the carbonyl established an internal hydrogen bond with the 4-NH, establishing a conformational lock and allowing truncation of the large basic substituent to a 1-methylcyclopyl group. Subsequent optimisation, facilitated by X-ray crystal structures, allowed identification of preferred substituents at both the pyridone core and pyrazole. Subsequent combinations of optimal groups allowed control of lipophilicity and identification of potent and selective inhibitors of IRAK4 with better in vitro permeability and lower clearance.

Cluster headache epidemiology including pediatric onset, sex, and ICHD criteria: Results from the International Cluster Headache Questionnaire.

OBJECTIVE: To validate the diagnoses and to investigate epidemiological data from an international, non-clinic-based, and large (n = 1604) survey of participants with cluster headache. BACKGROUND: There are several limitations in current epidemiological data in cluster headache including a lack of large non-clinic-based studies. There is also limited information on several aspects of cluster headache, such as pediatric incidence. METHODS: The International Cluster Headache Questionnaire was an internet-based survey that included questions on cluster headache demographics, criteria from the International Classification of Headache Disorders (ICHD), and medications. RESULTS: A total of 3251 subjects participated in the survey, and 1604 respondents met ICHD criteria for cluster headache. For validation, we interviewed a random sample of 5% (81/1604) of participants and confirmed the diagnosis of cluster headache in 97.5% (79/81). Pediatric onset was found in 27.5% (341/1583) of participants, and only 15.2% (52/341) of participants with pediatric onset were diagnosed before the age of 18. Men were more likely to have episodic cluster headache between ages 10 and 50, but the sex ratio was approximately equal for other ages. An overwhelming majority of respondents had at least one autonomic feature (99.0%, 1588/1604) and had restlessness (96.6%, 1550/1604), but many also had prototypical migrainous features including photophobia or phonophobia (50.1%, 804/1604), pain aggravated by physical activity (31.4%, 503/1604), or nausea and vomiting (27.5%, 441/1604). Interestingly, the first-line medications for acute treatment (oxygen) and preventive treatment (calcium channel blockers) were perceived as significantly less effective in chronic cluster headache (3.2 ± 1.1 and 2.1 ± 1.0 respectively on a 5-point ordinal scale) compared with episodic cluster headache (3.5 ± 1.0 and 2.4 ± 1.1, respectively, p < 0.001 for both comparisons). CONCLUSIONS: Cluster headache often occurs in the pediatric population, although they are typically not diagnosed until adulthood. The onset of cluster headache is the inverse of that in migraine; in migraine women are more likely to have migraine between ages 10 and 50 but the sex ratio is approximately equal otherwise. Prototypical migrainous features are not useful in differentiating cluster headache from migraine. Participant data from a large international study also suggest that chronic cluster headache is not only less responsive to newer treatments (like noninvasive vagus nerve stimulation and galcanezumab), but to traditional first-line treatments as well.

A brief diagnostic screen for cluster headache: Creation and initial validation of the Erwin Test for Cluster Headache.

OBJECTIVE: To use 1) newly generated data, 2) existing evidence, and 3) expert opinion to create and validate a new cluster headache screening tool. METHODS: In phase 1 of the study, we performed a prospective study of an English translation of an Italian screen on 95 participants (45 with cluster headache, 17 with other trigeminal autonomic cephalalgias, 30 with migraine, and 3 with trigeminal neuralgia). In phase 2, we performed a systematic review in PubMed of all studies until September 2019 with diagnostic screening tools for cluster headache. In phase 3, a 6-person panel of cluster headache patients, research coordinators, and headache specialists analyzed the data from the first two phases to generate a new diagnostic screening tool. Finally, in phase 4 this new screen was validated on participants at a single headache center (all diagnoses) and through research recruitment (trigeminal autonomic cephalalgias only, as recruitment was essential but was otherwise low). RESULTS: In total, this study included 319 unique participants including 109 cluster headache participants (95 total participants/45 cluster headache participants in phase 1, and 224 total participants/64 cluster headache participants in phase 4). It also found 123 articles on potential screening tools in our systematic review. In phase 1, analysis of the English translation of an Italian screen generated 7 questions with high sensitivity and specificity against migraine, trigeminal neuralgia, and other trigeminal autonomic cephalalgias, but had grammatical and other limitations as a general screening tool. In phase 2, the systematic review revealed nine studies that met inclusion criteria as diagnostic screening tools for cluster headache, including four where sensitivity and specificity were available for individual questions or small groups of questions. In phase 3, this data was reviewed by the expert panel to generate a brief (6-item), binary (yes/no), written screening test. In phase 4, a total of 224 participants completed the new 6-item screening test (81 migraine, 64 cluster headache, 21 other trigeminal autonomic cephalalgias, 35 secondary headaches, 7 neuralgias, 5 probable migraine, and 11 other headache disorders). Answers to the 6 items were combined in a decision tree algorithm and three items had a sensitivity of 84% (confidence interval or 95% confidence interval 73-92%), specificity of 89% (95% confidence interval 84-94%), positive predictive value of 76% (95% confidence interval 64-85%), and negative predictive value of 93% (95% confidence interval 88-97%) for the diagnosis of cluster headache. These three items focused on headache intensity, duration, and autonomic features. CONCLUSION: The 3-item Erwin Test for Cluster Headache is a promising diagnostic screening tool for cluster headache.

Cluster headache is one of the most intensely painful human conditions: Results from the International Cluster Headache Questionnaire.

OBJECTIVE: To determine the pain intensity of cluster headache through a large survey by comparing it to other painful disorders. Furthermore, to investigate the relationship between maximal pain, autonomic, and other clinical symptoms, as well as demographic attributes of cluster headache. BACKGROUND: The pain of cluster headache is anecdotally considered to be one of the worst pains in existence. The link between pain and autonomic features of cluster headache is understood mechanistically through the trigeminovascular reflex, though it is not clear if this is a graded response. Links between pain and other features of cluster headache are less well understood. METHODS: This Internet-based cross-sectional survey included questions on cluster headache diagnostic criteria, which were used as part of the inclusion/exclusion criteria for the study. Respondents were asked to rate a cluster headache attack on the 0-10 numerical rating scale. Additionally, they were asked if they had experienced a list of other painful conditions such as labor pain or nephrolithiasis; if so they were asked to rate that pain as well. The survey also included demographics, mood scores, and treatment responses. RESULTS: A total of 1604 cluster headache respondents were included in the analysis. Respondents rated cluster headache as significantly (p < 0.001) more intense than every other pain condition examined. Cluster headache attacks were rated as 9.7 ± 0.6 (mean ± standard deviation) on the numerical rating scale, followed by labor pain (7.2 ± 2.0), pancreatitis (7.0 ± 1.5), and nephrolithiasis (6.9 ± 1.9). The majority of cluster headache respondents rated a cluster headache attack at maximal or 10.0 pain (72.1%, 1157/1604). Respondents with maximal pain were statistically significantly more likely to have cranial autonomic features compared to respondents with less pain: conjunctival injection or lacrimation 91% (1057/1157) versus 85% (381/447), eyelid edema 77% (887/1157) versus 66% (293/447), forehead/facial sweating 60% (693/1157) versus 49% (217/447), fullness in the ear 47% (541/1157) versus 35% (155/447), and miosis/ptosis 85% (1124/1157) versus 75% (426/447) (all p values <0.001). Respondents with maximal pain also had other statistically significant findings: more frequent attacks (4.0 ± 2.0 attacks per day vs. 3.5 ± 2.0 attacks per day), higher Hopelessness Depression Symptom Questionnaire scores (24.5 ± 16.9 vs. 21.1 ± 15.2), decreased overall effectiveness from calcium channel blockers (on a 5-point Likert scale), and more likely female: 34% (389/1157) versus 24% (108/447) (all p values <0.001). Pain intensity was not associated with restlessness, headache duration, age of onset, episodic/chronic status, or the effectiveness of any acute or preventive medication other than calcium channel blockers. CONCLUSIONS: Cluster headache is an intensely painful disorder, even in the context of other disorders considered intensely painful. Maximal pain intensity is associated with more cranial autonomic features, suggesting a graded response between pain and autonomic features. Maximal pain intensity is also associated with headache frequency but not duration, suggesting a relationship between pain intensity and mechanisms controlling headache onset, but not between pain intensity and mechanisms controlling headache offset.

Improving metabolic stability and removing aldehyde oxidase liability in a 5-azaquinazoline series of IRAK4 inhibitors.

In this article, we report our efforts towards improving in vitro human clearance in a series of 5-azaquinazolines through a series of C4 truncations and C2 expansions. Extensive DMPK studies enabled us to tackle high Aldehyde Oxidase (AO) metabolism and unexpected discrepancies in human hepatocyte and liver microsomal intrinsic clearance. Our efforts culminated with the discovery of 5-azaquinazoline 35, which also displayed exquisite selectivity for IRAK4, and showed synergistic in vitro activity against MyD88/CD79 double mutant ABC-DLBCL in combination with the covalent BTK inhibitor acalabrutinib.

Effectiveness of Oxygen and Other Acute Treatments for Cluster Headache: Results From the Cluster Headache Questionnaire, an International Survey.

OBJECTIVE: To assess the effectiveness and adverse effects of acute cluster headache medications in a large international sample, including recommended treatments such as oxygen, commonly used medications such as opioids, and emerging medications such as intranasal ketamine. Particular focus is paid to a large subset of respondents 65 years of age or older. BACKGROUND: Large international surveys of cluster headache are rare, as are examinations of treatments and side effects in older cluster headache patients. This article presents data from the Cluster Headache Questionnaire, with respondents from over 50 countries and with the vast majority from the United States, the United Kingdom, and Canada. METHODS: This internet-based survey included questions on cluster headache diagnostic criteria, which were used as part of the inclusion/exclusion criteria for the study, as well as effectiveness of medications, physical and medical complications, psychological and emotional complications, mood scores, and difficulty obtaining medications. The diagnostic questions were also used to create a separate group of respondents with probable cluster headache. Limitations to the methods include the use of nonvalidated questions, the lack of a formal clinical diagnosis of cluster headache, and the grouping of some medications (eg, all triptans as opposed to sumatriptan subcutaneous alone). RESULTS: A total of 3251 subjects participated in the questionnaire, and 2193 respondents met criteria for this study (1604 cluster headache and 589 probable cluster headache). Of the respondents with cluster headache, 68.8% (1104/1604) were male and 78.0% (1245/1596) had episodic cluster headache. Over half of respondents reported complete or very effective treatment for triptans (54%, 639/1139) and oxygen (54%, 582/1082). Between 14 and 25% of respondents reported complete or very effective treatment for ergot derivatives (dihydroergotamine 25%, 42/170; cafergot/ergotamine 17%, 50/303), caffeine and energy drinks (17%, 7/41), and intranasal ketamine (14%, 5/37). Less than 10% reported complete or very effective treatment for opioids (6%, 30/541), intranasal capsaicin (5%, 7/151), and intranasal lidocaine (2%, 5/241). Adverse events were especially low for oxygen (no or minimal physical and medical complications 99%, 1077/1093; no or minimal psychological and emotional complications 97%, 1065/1093), intranasal lidocaine (no or minimal physical and medical complications 97%, 248/257; no or minimal psychological and emotional complications 98%, 251/257), intranasal ketamine (no or minimal physical and medical complications 95%, 38/40; no or minimal psychological and emotional complications 98%, 39/40), intranasal capsaicin (no or minimal physical and medical complications 91%, 145/159; no or minimal psychological and emotional complications 94%, 150/159), and caffeine and energy drinks (no or minimal physical and medical complications 89%, 39/44; no or minimal psychological and emotional complications 91%, 40/44). This is in comparison to ergotamine/cafergot (no or minimal physical and medical complications 83%, 273/327; no or minimal psychological and emotional complications 89%, 290/327), dihydroergotamine (no or minimal physical and medical complications 81%, 143/176; no or minimal psychological and emotional complications 91%, 160/176), opioids (no or minimal physical and medical complications 76%, 416/549; no or minimal psychological and emotional complications 77%, 423/549), or triptans (no or minimal physical and medical complications 73%, 883/1218; no or minimal psychological and emotional complications 85%, 1032/1218). A total of 139 of 1604 cluster headache respondents (8.7%) were age 65 and older and reported similar effectiveness and adverse events to the general population. The 589 respondents with probable cluster headache reported similar medication effectiveness to respondents with a full diagnosis of cluster headache. CONCLUSIONS: Oxygen is reported by survey respondents to be a highly effective treatment with few complications in cluster headache in a large international sample, including those 65 years or over. Triptans are also very effective with some side effects, and newer medications deserve additional study. Patients with probable cluster headache may respond similarly to acute medications as patients with a full diagnosis of cluster headache.

Discovery of N-(4-{[5-Fluoro-7-(2-methoxyethoxy)quinazolin-4-yl]amino}phenyl)-2-[4-(propan-2-yl)-1 H-1,2,3-triazol-1-yl]acetamide (AZD3229), a Potent Pan-KIT Mutant Inhibitor for the Treatment of Gastrointestinal Stromal Tumors.

While the treatment of gastrointestinal stromal tumors (GISTs) has been revolutionized by the application of targeted tyrosine kinase inhibitors capable of inhibiting KIT-driven proliferation, diverse mutations to this kinase drive resistance to established therapies. Here we describe the identification of potent pan-KIT mutant kinase inhibitors that can be dosed without being limited by the tolerability issues seen with multitargeted agents. This effort focused on identification and optimization of an existing kinase scaffold through the use of structure-based design. Starting from a series of previously reported phenoxyquinazoline and quinoline based inhibitors of the tyrosine kinase PDGFRα, potency against a diverse panel of mutant KIT driven Ba/F3 cell lines was optimized, with a particular focus on reducing activity against a KDR driven cell model in order to limit the potential for hypertension commonly seen in second and third line GIST therapies. AZD3229 demonstrates potent single digit nM growth inhibition across a broad cell panel, with good margin to KDR-driven effects. Selectivity over KDR can be rationalized predominantly by the interaction of water molecules with the protein and ligand in the active site, and its kinome selectivity is similar to the best of the approved GIST agents. This compound demonstrates excellent cross-species pharmacokinetics, shows strong pharmacodynamic inhibition of target, and is active in several in vivo models of GIST.

Discovery of a Series of 3-Cinnoline Carboxamides as Orally Bioavailable, Highly Potent, and Selective ATM Inhibitors.

We report the discovery of a novel series of 3-cinnoline carboxamides as highly potent and selective ataxia telangiectasia mutated (ATM) kinase inhibitors. Optimization of this series focusing on potency and physicochemical properties (especially permeability) led to the identification of compound 21, a highly potent ATM inhibitor (ATM cell IC50 0.0028 µM) with excellent kinase selectivity and favorable physicochemical and pharmacokinetics properties. In vivo, 21 in combination with irinotecan showed tumor regression in the SW620 colorectal tumor xenograft model, superior inhibition to irinotecan alone. Compound 21 was selected for preclinical evaluation alongside AZD0156.

AZD9496: An Oral Estrogen Receptor Inhibitor That Blocks the Growth of ER-Positive and ESR1-Mutant Breast Tumors in Preclinical Models.

Fulvestrant is an estrogen receptor (ER) antagonist administered to breast cancer patients by monthly intramuscular injection. Given its present limitations of dosing and route of administration, a more flexible orally available compound has been sought to pursue the potential benefits of this drug in patients with advanced metastatic disease. Here we report the identification and characterization of AZD9496, a nonsteroidal small-molecule inhibitor of ERα, which is a potent and selective antagonist and downregulator of ERα in vitro and in vivo in ER-positive models of breast cancer. Significant tumor growth inhibition was observed as low as 0.5 mg/kg dose in the estrogen-dependent MCF-7 xenograft model, where this effect was accompanied by a dose-dependent decrease in PR protein levels, demonstrating potent antagonist activity. Combining AZD9496 with PI3K pathway and CDK4/6 inhibitors led to further growth-inhibitory effects compared with monotherapy alone. Tumor regressions were also seen in a long-term estrogen-deprived breast model, where significant downregulation of ERα protein was observed. AZD9496 bound and downregulated clinically relevant ESR1 mutants in vitro and inhibited tumor growth in an ESR1-mutant patient-derived xenograft model that included a D538G mutation. Collectively, the pharmacologic evidence showed that AZD9496 is an oral, nonsteroidal, selective estrogen receptor antagonist and downregulator in ER(+) breast cells that could provide meaningful benefit to ER(+) breast cancer patients. AZD9496 is currently being evaluated in a phase I clinical trial. Cancer Res; 76(11); 3307-18. ©2016 AACR.

Optimization of a Novel Binding Motif to (E)-3-(3,5-Difluoro-4-((1R,3R)-2-(2-fluoro-2-methylpropyl)-3-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indol-1-yl)phenyl)acrylic Acid (AZD9496), a Potent and Orally Bioavailable Selective Estrogen Receptor Downregulator and Antagonist.

The discovery of an orally bioavailable selective estrogen receptor downregulator (SERD) with equivalent potency and preclinical pharmacology to the intramuscular SERD fulvestrant is described. A directed screen identified the 1-aryl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole motif as a novel, druglike ER ligand. Aided by crystal structures of novel ligands bound to an ER construct, medicinal chemistry iterations led to (E)-3-(3,5-difluoro-4-((1R,3R)-2-(2-fluoro-2-methylpropyl)-3-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indol-1-yl)phenyl)acrylic acid (30b, AZD9496), a clinical candidate with high oral bioavailability across preclinical species that is currently being evaluated in phase I clinical trials for the treatment of advanced estrogen receptor (ER) positive breast cancer.

Discovery of AZD3147: a potent, selective dual inhibitor of mTORC1 and mTORC2.

High throughput screening followed by a lead generation campaign uncovered a novel series of urea containing morpholinopyrimidine compounds which act as potent and selective dual inhibitors of mTORC1 and mTORC2. We describe the continued compound optimization campaign for this series, in particular focused on identifying compounds with improved cellular potency, improved aqueous solubility, and good stability in human hepatocyte incubations. Knowledge from empirical SAR investigations was combined with an understanding of the molecular interactions in the crystal lattice to improve both cellular potency and solubility, and the composite parameters of LLE and pIC50-pSolubility were used to assess compound quality and progress. Predictive models were employed to efficiently mine the attractive chemical space identified resulting in the discovery of 42 (AZD3147), an extremely potent and selective dual inhibitor of mTORC1 and mTORC2 with physicochemical and pharmacokinetic properties suitable for development as a potential clinical candidate.

Discovery of 4-amino-N-[(1S)-1-(4-chlorophenyl)-3-hydroxypropyl]-1-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide (AZD5363), an orally bioavailable, potent inhibitor of Akt kinases.

Wide-ranging exploration of analogues of an ATP-competitive pyrrolopyrimidine inhibitor of Akt led to the discovery of clinical candidate AZD5363, which showed increased potency, reduced hERG affinity, and higher selectivity against the closely related AGC kinase ROCK. This compound demonstrated good preclinical drug metabolism and pharmacokinetics (DMPK) properties and, after oral dosing, showed pharmacodynamic knockdown of phosphorylation of Akt and downstream biomarkers in vivo, and inhibition of tumor growth in a breast cancer xenograft model.

Design and synthesis of novel lactate dehydrogenase A inhibitors by fragment-based lead generation.

Lactate dehydrogenase A (LDHA) catalyzes the conversion of pyruvate to lactate, utilizing NADH as a cofactor. It has been identified as a potential therapeutic target in the area of cancer metabolism. In this manuscript we report our progress using fragment-based lead generation (FBLG), assisted by X-ray crystallography to develop small molecule LDHA inhibitors. Fragment hits were identified through NMR and SPR screening and optimized into lead compounds with nanomolar binding affinities via fragment linking. Also reported is their modification into cellular active compounds suitable for target validation work.

Development of a practical Buchwald-Hartwig amine arylation protocol using a conveniently prepared (NHC)Pd(R-allyl)Cl catalyst.

Continuing efforts to establish a more general "user-friendly" protocol for the palladium-catalysed arylation of amines (Buchwald-Hartwig reaction) are described herein. Significant advances have been made through the use of the versatile (SIPr)Pd(methallyl)Cl complex in conjunction with the reliable base lithium hexamethyldisilazide (LHMDS).

A new series of neutral 5-substituted 4-anilinoquinazolines as potent, orally active inhibitors of erbB2 receptor tyrosine kinase.

Starting from initial lead 1 containing a basic 5-substituent, optimisation of the glycolamide-derived neutral 5-substituent led to potent inhibitors of erbB2 with good pharmacokinetics. Representative compounds 19 and 21 inhibited phosphorylation of erbB2 in a mouse BT474C xenograft model after oral administration.

Neutral 5-substituted 4-anilinoquinazolines as potent, orally active inhibitors of erbB2 receptor tyrosine kinase.

Neutral 5-substituted 4-anilinoquinazolines addressed high in vivo clearance and phospholipidosis associated with previous basic compounds. A representative compound 8a inhibited tumor growth in a mouse xenograft model when co-administered with the cytochrome P450 inhibitor 1-aminobenzotriazole (ABT), and data are consistent with pharmacology primarily reflecting inhibition of erbB2 receptor tyrosine kinase.

Inhibitors of epidermal growth factor receptor tyrosine kinase: optimisation of potency and in vivo pharmacokinetics.

The structure-activity and structure-property relationships of anilinoquinazoline inhibitors of EGFR were investigated. Strategies to lower volume of distribution and shorten half-life through structure and pKa modulation are discussed.