Filling a nick in NIK: Extending the half-life of a NIK inhibitor through structure-based drug design.

Inhibition of NF-κB inducing kinase (NIK) has been pursued as a promising therapeutic target for autoimmune disorders due to its highly regulated role in key steps of the NF-κB signaling pathway. Previously reported NIK inhibitors from our group were shown to be potent, selective, and efficacious, but had higher human dose projections than desirable for immunology indications. Herein we report the clearance-driven optimization of a NIK inhibitor guided by metabolite identification studies and structure-based drug design. This led to the identification of an azabicyclo[3.1.0]hexanone motif that attenuated in vitro and in vivo clearance while maintaining NIK potency and increasing selectivity over other kinases, resulting in a greater than ten-fold reduction in predicted human dose.

Strategies to Mitigate the Bioactivation of Aryl Amines.

The bioactivation of xenobiotics to yield reactive metabolites can lead to tolerability and toxicity concerns within a drug discovery program. Development of strategies for mitigating the metabolic liability of commonly encountered toxicophores, such as anilines, relies on an understanding of the relative tendency of these functionalities to undergo bioactivation. In this report, we present the first systematic study of the structure-activity relationships of the bioactivation of aryl amine fragments (molecular weight < 250 Da) using a glutathione (GSH) trapping assay in the presence of human liver microsomes and the reduced form of nicotinamide adenine dinucleotide phosphate. This study demonstrates that conversion of anilines to nitrogen-containing heteroarylamines results in a lower abundance of GSH conjugates in the order phenyl > pyrimidine ≈ pyridine > pyridazine. Introduction of electron-withdrawing functionality on the aromatic ring had a less pronounced effect on the extent of GSH conjugation. Examination of more drug-like compounds sourced from in-house drug discovery programs revealed similar trends in bioactivation between matched pairs containing (hetero)aryl amines. This study provides medicinal chemists with insights and qualitative guidance for the minimization of risks related to aryl amine metabolism.

Enantioselective Synthesis of Azamerone.

A concise and selective synthesis of the dichlorinated meroterpenoid azamerone is described. The paucity of tactics for the synthesis of natural-product-relevant chiral organochlorides motivated the development of unique strategies for accessing these motifs in enantioenriched forms. The route features a novel enantioselective chloroetherification reaction, a Pd-catalyzed cross-coupling between a quinone diazide and a boronic hemiester, and a late-stage tetrazine [4+2]-cycloaddition/oxidation cascade.

Catalytic Enantioselective Dihalogenation in Total Synthesis.

To date, more than 5000 biogenic halogenated molecules have been characterized. This number continues to increase as chemists explore chloride- and bromide-rich marine environments in search of novel bioactive natural products. Naturally occurring organohalogens span nearly all biosynthetic structural classes, exhibit a range of unique biological activities, and have been the subject of numerous investigations. Despite the abundance of and interest in halogenated molecules, enantioselective methods capable of forging carbon-halogen bonds in synthetically relevant contexts remain scarce. Accordingly, syntheses of organohalogens often rely on multistep functional group interconversions to establish carbon-halogen stereocenters. Our group has developed an enantioselective dihalogenation reaction and utilized it in the only reported examples of catalytic enantioselective halogenation in natural product synthesis. In this Account, we describe our laboratory's development of a method for catalytic, enantioselective dihalogenation and the application of this method to the synthesis of both mono- and polyhalogenated natural products. In the first part, we describe the initial discovery of a TADDOL-mediated dibromination of cinnamyl alcohols. Extension of this reaction to a second-generation system capable of selective bromochlorination, dichlorination, and dibromination is then detailed. This system is capable of controlling the enantioselectivity of dihalide formation, chemoselectivity for polyolefinic substrates, and regioselectivity in the case of bromochlorination. The ability of this method to exert control over regioselectivity of halide delivery permits selective halogenation of electronically nonbiased olefins required for total synthesis. In the second part, we demonstrate how the described dihalogenation has provided efficient access to a host of structurally diverse natural products. The most direct application of this methodology is in the synthesis of naturally occurring vicinal dihalides. Chiral vicinal bromochlorides represent a class of >175 natural products; syntheses of five members of this class, including its flagship member, (+)-halomon, have been accomplished through use of the catalytic, enantioselective bromochlorination. Likewise, enantioselective dichlorination has provided selective access to two members of the chlorosulfolipids, a class of linear, acyclic polychlorides. Synthesis of chiral monohalides has been achieved through solvolysis of enantioenriched bromochlorides; this approach has resulted in the synthesis of five bromocyclohexane-containing natural products through an enantiospecific bromopolyene cyclization. In reviewing these syntheses, a framework for the synthesis of chiral organohalogens mediated by catalytic, enantioselective dihalogenation has emerged. The development of a selective dihalogenation method has been highly enabling in the synthesis of halogenated natural products. In this Account, we detail all examples of catalytic, enantioselective halogenation in total synthesis and encourage the further development of synthetically useful halogenation methodologies.

Catalytic Enantioselective Dihalogenation and the Selective Synthesis of (-)-Deschloromytilipin A and (-)-Danicalipin A.

A titanium-based catalytic enantioselective dichlorination of simple allylic alcohols is described. This dichlorination reaction provides stereoselective access to all common dichloroalcohol building blocks used in syntheses of chlorosulfolipid natural products. An enantioselective synthesis of ent-(-)-deschloromytilipin A and a concise, eight-step synthesis of ent-(-)-danicalipin A are executed and employ the dichlorination reaction as the first step. Extension of this system to enantioselective dibromination and its use in the synthesis of pentabromide stereoarrays relevant to bromosulfolipids is reported. The described dichlorination and dibromination reactions are capable of exerting diastereocontrol in complex settings allowing X-ray crystal structure analysis of natural and unnatural diastereomers of polyhalogenated stereohexads.

When Cofactors Aren't X Factors: Functional Groups That Are Labile in Human Liver Microsomes in the Absence of NADPH.

The metabolic stability of compounds is often assessed at an early stage in drug discovery programs by profiling with hepatic microsomes. Exclusion of the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) in these assays provides insight into non-cytochrome P450 (CYP)-mediated metabolism. This report uses a matched molecular pair (MMP) application to assess which chemical substituents are commonly susceptible to non-NADPH-mediated metabolism by microsomes. The analysis found the overall prevalence of metabolism in the absence of NADPH to be low, with esters, amides, aldehydes, and oxetanes being among the most commonly susceptible functional groups. Given that non-CYP enzymes, such as esterases, may be expressed extrahepatically and lead to lower confidence in predicted pharmacokinetic profiles, an awareness of the functional groups that commonly undergo non-NADPH-mediated metabolism-as well as options for their replacement based on experimental MMP data-may help researchers derisk metabolic stability issues at an earlier stage in drug discovery.

LogD Contributions of Substituents Commonly Used in Medicinal Chemistry.

The importance of physicochemical property calculation and measurement is well-established in drug discovery. In particular, lipophilicity predictions play a central role in target design and prioritization. While significant progress has been made in our ability to calculate both logP and logD, the quality of these predictions is limited by the size and diversity of the underlying data set. Access to diverse data sets and advanced models is often limited to large organizations or consortia, and they are not available to many students and practitioners of medicinal chemistry. A molecular matched pair analysis of median ΔlogD 7.4 contributions for substituents commonly used in drug discovery programs at Genentech is reported. The results of this ΔlogD analysis are compiled into a single table, which we anticipate will be of use to practicing medicinal chemists.

Halogenation-Dependent Effects of the Chlorosulfolipids of Ochromonas danica on Lipid Bilayers.

The chlorosulfolipids are amphiphilic natural products with stereochemically complex patterns of chlorination and sulfation. Despite their role in toxic shellfish poisoning, potential pharmacological activities, and unknown biological roles, they remain understudied due to the difficulties in purifying them from natural sources. The structure of these molecules, with a charged sulfate group in the middle of the hydrophobic chain, appears incompatible with the conventional lipid bilayer structure. Questions about chlorosulfolipids remain unanswered partly due to the unavailability of structural analogues with which to conduct structure-function studies. We approach this problem by combining enantioselective total synthesis and membrane biophysics. Using a combination of Langmuir pressure-area isotherms of lipid monolayers, fluorescence imaging of vesicles, mass spectrometry imaging, natural product isolation, small-angle X-ray scattering, and cryogenic electron microscopy, we show that danicalipin A (1) likely inserts into lipid bilayers in the headgroup region and alters their structure and phase behavior. Specifically, danicalipin A (1) thins the bilayer and fluidizes it, allowing even saturated lipid to form fluid bilayers. Lipid monolayers show similar fluidizing upon insertion of danicalipin A (1). Furthermore, we show that the halogenation of the molecule is critical for its membrane activity, likely due to sterically controlled conformational changes. Synthetic unchlorinated and monochlorinated analogues do not thin and fluidize lipid bilayers to the same extent as the natural product. Overall, this study sheds light on how amphiphilic small molecules interact with lipid bilayers and the importance of stereochemistry and halogenation for this interaction.