Characterization of a Stable 2,2'-Azobis(2-Methylpropanenitrile) Degradant and Its Use to Monitor the Oxidative Environment During Forced Degradation Studies by Liquid Chromatography/Mass Spectrometry.

Azo compounds are commonly used to study radical-mediated degradation of pharmaceutical compounds. The favorable chemical and physical properties of 2,2'-azobis(2-methylpropanenitrile) (AIBN) have made it one of the most widely used compound for these type of studies. This article describes the characterization of a stable product, N-(1-cyano-1-methylethyl)-2-methylpropanamide, formed during the decomposition of AIBN.

A selective, orally bioavailable 1,2,4-triazolo[1,5-a]pyridine-based inhibitor of Janus kinase 2 for use in anticancer therapy: discovery of CEP-33779.

Members of the JAK family of nonreceptor tyrosine kinases play a critical role in the growth and progression of many cancers and in inflammatory diseases. JAK2 has emerged as a leading therapeutic target for oncology, providing a rationale for the development of a selective JAK2 inhibitor. A program to optimize selective JAK2 inhibitors to combat cancer while reducing the risk of immune suppression associated with JAK3 inhibition was undertaken.

Design, synthesis, and anaplastic lymphoma kinase (ALK) inhibitory activity for a novel series of 2,4,8,22-tetraazatetracyclo[14.3.1.1³,⁷.1⁹,¹³]docosa-1(20),3(22),4,6,9(21),10,12,16,18-nonaene macrocycles.

A novel set of 2,4,8,22-tetraazatetracyclo[14.3.1.

Strategies to mitigate the bioactivation of 2-anilino-7-aryl-pyrrolo[2,1-f][1,2,4]triazines: identification of orally bioavailable, efficacious ALK inhibitors.

Chemical strategies to mitigate cytochrome P450-mediated bioactivation of novel 2,7-disubstituted pyrrolo[2,1-f][1,2,4]triazine ALK inhibitors are described along with synthesis and biological activity. Piperidine-derived analogues showing minimal microsomal reactive metabolite formation were discovered. Potent, selective, and metabolically stable ALK inhibitors from this class were identified, and an orally bioavailable compound (32) with antitumor efficacy in ALK-driven xenografts in mouse models was extensively characterized.

2,7-Pyrrolo[2,1-f][1,2,4]triazines as JAK2 inhibitors: modification of target structure to minimize reactive metabolite formation.

The JAK2/STAT pathway has important roles in hematopoiesis. With the discovery of the JAK2 V617F mutation and its presence in many patients with myeloproliferative neoplasms, research in the JAK2 inhibitor arena has dramatically increased. We report a novel series of potent JAK2 inhibitors containing a 2,7-pyrrolotriazine core.

2,7-Disubstituted-pyrrolotriazine kinase inhibitors with an unusually high degree of reactive metabolite formation.

There are numerous published studies establishing a link between reactive metabolite formation and toxicity of various drugs. Although the correlation between idiosyncratic reactions and reactive metabolite formation is not 1:1, the association between the two is such that many pharmaceutical companies now monitor for reactive metabolites as a standard part of drug candidate testing and selection. The most common method involves in vitro human microsomal incubations in the presence of a thiol trapping agent, such as glutathione (GSH), followed by LC/MS analysis.

2,4-Diaminopyrimidine inhibitors of c-Met kinase bearing benzoxazepine anilines.

Elaboration of the SAR around a series of 2,4-diaminopyrimidines led to a number of c-Met inhibitors in which kinase selectivity was modulated by substituents appended on the C4-aminobenzamide ring and the nature of the C2-aminoaryl ring. Further lead optimization of the C2-aminoaryl group led to benzoxazepine analogs whose pharmaceutical properties were modulated by the nature of the substituent on the benzoxazepine nitrogen. Tumor stasis (with partial regressions) were observed when an orally bioavailable analog was evaluated in a GTL-16 tumor xenograft mouse model.

Modification of CellSensor irf1-bla TF-1 and irf1-bla HEL assays for direct comparison of wild-type JAK2 and JAK2 V617F inhibition.

The Janus kinase (JAK)-signal transducer and activator of transcription pathway is an important therapeutic target because of its role in the regulation of cell growth. Aberrant, constitutive activation of JAK2 signaling has been implicated in myeloproliferative disorders with a single, activating somatic V617F mutation in the JH2 pseudokinase domain of JAK2 as the prevalent molecular lesion. Invitrogen has developed the CellSensor(®) cell lines interferon regulatory factor-1 (irf1)-beta-lactamase (bla) TF-1 and irf1-bla HEL for use in evaluating inhibitors of wild-type JAK2 and mutant JAK2 V617F, respectively.

Acyclic, orally bioavailable ketone-based cathepsin K inhibitors.

Starting from a potent ketone-based inhibitor with poor drug properties, incorporation of P(2)-P(3) elements from a ketoamide-based inhibitor led to the identification of a hybrid series of ketone-based cathepsin K inhibitors with better oral bioavailability than the starting ketone.

Novel, potent P2-P3 pyrrolidine derivatives of ketoamide-based cathepsin K inhibitors.

Starting from a potent pantolactone ketoamide cathepsin K inhibitor discovered from structural screening, conversion of the lactone scaffold to a pyrrolidine scaffold allowed exploration of the S(3) subsite of cathepsin K. Manipulation of P3 and P1' groups afforded potent inhibitors with drug-like properties.

Semicarbazone-based inhibitors of cathepsin K, are they prodrugs for aldehyde inhibitors?

Starting from potent aldehyde inhibitors with poor drug properties, derivatization to semicarbazones led to the identification of a series of semicarbazone-based cathepsin K inhibitors with greater solubility and better pharmacokinetic profiles than their parent aldehydes. Furthermore, a representative semicarbazone inhibitor attenuated bone resorption in an ex vivo rat calvarial bone resorption model. However, based on enzyme inhibition comparisons at neutral pH, semicarbazone hydrolysis rates, and 13C NMR experiments, these semicarbazones probably function as prodrugs of aldehydes.

Elements of diabetic nephropathy in a patient with GLUT 2 deficiency.

The Fanconi-Bickel syndrome is caused by homozygosity or compound heterozygosity for mutations of the facilitated glucose transporter 2 gene (GLUT2). Glycogen accumulates in renal tubular cells and they fail to reabsorb multiple filtered solutes because of impairment in GLUT2-mediated efflux of glucose. We describe a 10-year-old male child with GLUT2 deficiency who produced massive amounts of 3-deoxyfructose (3-DF) in the kidneys.

P2-P3 conformationally constrained ketoamide-based inhibitors of cathepsin K.

An orally bioavailable series of ketoamide-based cathepsin K inhibitors with good pharmacokinetic properties has been identified. Starting from a potent inhibitor endowed with poor drug properties, conformational constraint of the P(2)-P(3) linker and modifications to P(1') elements led to an enhancement in potency, solubility, clearance, and bioavailability. These optimized inhibitors attenuated bone resorption in a rat TPTX hypocalcemic bone resorption model.

A structural screening approach to ketoamide-based inhibitors of cathepsin K.

Several novel ketoamide-based inhibitors of cathepsin K have been identified. Starting from a modestly potent inhibitor, structural screening of P2 elements led to 100-fold enhancements in inhibitory activity. Modifications to one of these leads resulted in an orally bioavailable cathepsin K inhibitor.

Potent and selective P2-P3 ketoamide inhibitors of cathepsin K with good pharmacokinetic properties via favorable P1', P1, and/or P3 substitutions.

A series of ketoamides were synthesized and evaluated for inhibitory activity against cathepsin K. Exploration of the interactions between achiral P(2) substituents and the cysteine protease based on molecular modelling suggestions resulted in potent cathepsin K inhibitors that demonstrated high selectivity versus cathepsins B, H, and L. Subsequent modifications of the P(3), P(1), and P(1') moieties afforded orally bioavailable inhibitors.

Orally bioavailable small molecule ketoamide-based inhibitors of cathepsin K.

An orally available series of ketoamide-based inhibitors of cathepsin K has been identified. Starting from a potent inhibitor with poor oral bioavailability, modifications to P1 and P1' elements led to enhancements in solubility and permeability. These improvements resulted in orally available cathepsin K inhibitors.

Design of small molecule ketoamide-based inhibitors of cathepsin K.

A novel series of ketoamide-based inhibitors of cathepsin K has been identified. Modifications to P(2) and P(3) elements were crucial to enhancing inhibitory activity. Although not optimized, a selected inhibitor was effective in attenuating type I collagen hydrolysis in a surrogate assay of bone resorption.