Justin T. Douglas

Opioid receptor probes derived from cycloaddition of the hallucinogen natural product salvinorin A.

As part of our continuing efforts toward more fully understanding the structure-activity relationships of the neoclerodane diterpene salvinorin A, we report the synthesis and biological characterization of unique cycloadducts through [4+2] Diels-Alder cycloaddition. Microwave-assisted methods were developed and successfully employed, aiding in functionalizing the chemically sensitive salvinorin A scaffold. This demonstrates the first reported results for both cycloaddition of the furan ring and functionalization via microwave-assisted methodology of the salvinorin A skeleton. The cycloadducts yielded herein introduce electron-withdrawing substituents and bulky aromatic groups into the C-12 position. Kappa opioid (KOP) receptor space was explored through aromatization of the bent oxanorbornadiene system possessed by the cycloadducts to a planar phenyl ring system. Although dimethyl- and diethylcarboxylate analogues 5 and 6 retain some affinity and selectivity for KOP receptors and are full agonists, their aromatized counterparts 13 and 14 have reduced affinity for KOP receptors. The methods developed herein signify a novel approach toward rapidly probing the structure-activity relationships of furan-containing natural products.

Design, Synthesis and Biological Evaluation of Aminoalkylindole Derivatives as Cannabinoid Receptor Ligands with Potential for Treatment of Alcohol Abuse

Attenuation of increased endocannabinoid signaling with a CB1R neutral antagonist might offer a new therapeutic direction for treatment of alcohol abuse. We have recently reported that a monohydroxylated metabolite of the synthetic aminoalkylindole cannabinoid JHW-073 (3) exhibits neutral antagonist activity at CB1Rs and thus may serve as a promising lead for the development of novel alcohol abuse therapies. In the current study, we show that systematic modification of an aminoalkylindole scaffold identified two new compounds with dual CB1R antagonist/CB2R agonist activity. Similar to the CB1R antagonist/inverse agonist rimonabant, analogues 27 and 30 decrease oral alcohol self-administration without affecting total fluid intake and block the development of alcohol-conditioned place preference. Collectively, these initial findings suggest that design and systematic modification of aminoalkylindoles such as 3 may lead to development of novel cannabinoid ligands with dual CB1R antagonist/CB2R agonist activity with potential for use as treatments of alcohol abuse.

Mechanistic Origin of the Stereodivergence in Decarboxylative Allylation

A stereochemical test has been used to probe the mechanism of decarboxylative allylation. This probe suggests that the mechanism of DcA reactions can change based on the substitution pattern at the alpha-carbon of the nucleophile; however, reaction via stabilized malonate nucleophiles is the lower energy pathway. Lastly, this mechanistic proposal has predictive power and can be used to explain chemoselectivities in decarboxylative reactions that were previously confounding.

Copper‐Mediated Deoxygenative Trifluoromethylation of Benzylic Xanthates: Generation of a CCF3 Bond from an O‐Based Electrophile

The conversion of an alcohol-based functional group, into a trifluoromethyl analogue is a desirable transformation. However, few methods are capable of converting O-based electrophiles into trifluoromethanes. The copper-mediated trifluoromethylation of benzylic xanthates using Umemotos reagent as the source of CF3 to form C-CF3 bonds is described. The method is compatible with an array of benzylic xanthates bearing useful functional groups. A preliminary mechanistic investigation suggests that the C-CF3 bond forms by reaction of the substrate with in situ generated CuCF3 and CuOTf. Further evidence suggests that the reaction could proceed via a radical cation intermediate.

Mechanism of Decarboxylation of Pyruvic Acid in the Presence of Hydrogen Peroxide.

The purpose of this work was to probe the rate and mechanism of rapid decarboxylation of pyruvic acid in the presence of hydrogen peroxide (H2O2) to acetic acid and carbon dioxide over the pH range 2-9 at 25 °C, utilizing UV spectrophotometry, high performance liquid chromatography (HPLC), and proton and carbon nuclear magnetic resonance spectrometry ((1)H, (13)C-NMR). Changes in UV absorbance at 220 nm were used to determine the kinetics as the reaction was too fast to follow by HPLC or NMR in much of the pH range. The rate constants for the reaction were determined in the presence of molar excess of H2O2 resulting in pseudo first-order kinetics. No buffer catalysis was observed. The calculated second-order rate constants for the reaction followed a sigmoidal shape with pH-independent regions below pH 3 and above pH 7 but increased between pH 4 and 6. Between pH 4 and 9, the results were in agreement with a change from rate-determining nucleophilic attack of the deprotonated peroxide species, HOO(-), on the α-carbonyl group followed by rapid decarboxylation at pH values below 6 to rate-determining decarboxylation above pH 7. The addition of H2O2 to ethyl pyruvate was also characterized.

1,3-Allylic Strain as a Strategic Diversification Element for Constructing Libraries of Substituted 2-Arylpiperidines†

Screening approaches to probe and drug discovery require access to high-quality small molecule libraries. One contemporary challenge in providing such access is the construction of libraries that maximize the coverage of chemical (functional group), stereochemical, and spatial diversity in a given chemotype.[1] Although the problem of functional group diversity has been addressed since the earliest days of combinatorial chemistry and parallel synthesis, the incorporation of stereochemical diversity and, more broadly, shape diversity has required the development of new strategies. These include the use of spatially diverse scaffolds or the pre-construction of stereochemically diverse building blocks that are then combined to afford final products (“build–couple–pair” [2] is an example of this). In this paper, we describe a conformational switching approach toward shape-diverse piperidine libraries in which the presence or absence of 1,3-allylic strain[3] is leveraged to enhance both (1) scaffold diversity by regiodivergent opening of epoxide intermediates and (2) the conformational space of the final library through the simple expedient of changing the nature of nitrogen substitution.[4] The concept is illustrated Figure 1 for a series of 2-aryl substituted piperidines. For a given epoxide isomer, the conformation of the piperidine ring will depend on whether the N1 atom is sp3 hybridized (Ar preferring an equatorial position due to minimization of 1,3-diaxial interactions) or sp2 (Ar axial, due to A1,3 strain in the equatorial isomer[5]). Nucleophilic addition to the epoxide would then take place according to the Furst–Plattner principle[6] (trans–diaxial opening), leading to two constitutional isomers from this epoxide intermediate (2,4- vs. 2,5-cis Ar/Nu relationships). Stereochemical diversity would then follow by applying the same principles to the alternative epoxide diastereomer, affording the analogous 2,4- and 2,5-trans isomers. Once prepared – and likely following the downstream introduction of functional group diversity – the library could then be N-substituted by a different set of alkyl or acyl substituents, leading to a doubling of the library members through conformational diversity. Figure 1 Use of A1,3 strain to control constitutional (2,4- vs. 2,5-Ar/Nu relationship), stereochemical (cis vs. trans), and conformational diversity in piperidine libraries. Functional group diversity arises from variations in Ar, Nu, and R. We chose to demonstrate this approach by preparing a library based on the triazole-containing piperidines shown in Figure 2 (selected because the arylpiperidine chemotype appears in a number of bioactive compounds and is therefore a desirable library scaffold for broad screening[7]). To a first approximation, the expected conformations in one such library are shown (four isomers bearing two different N-groups), demonstrating the range of conformational and configurational space covered by these compounds. Figure 2 Conformational diversity of targeted library containing sp3 and sp2 hybridized N-substituted versions of scaffolds A and B. In all, eight idealized conformers (four amines and four amides, all chair piperidines) are shown in the two overlays. Details, ... To demonstrate the value of 1,3-allylic strain in scaffold preparation, we first carried out the stereochemically- and constitutionally-differentiated scaffold syntheses shown in Scheme 1. Four 2-aryl-1,2,3,6-tetrahydropyridines were constructed from substituted benzaldehydes[8] in two steps via bisallylation/N-acylation followed by RCM. The N-acylated derivatives underwent highly stereoselective epoxidation reactions, presumably because the top faces as drawn were blocked by the aromatic groups in the most stable conformations.[5] Using m-CPBA or methyl(trifluoromethyl)dioxirane,[9] epoxidation from the less-hindered bottom alkene face afforded anti epoxides 3 (after DBU-mediated Fmoc removal) and 5.[10] Alternatively, NBS/H2O produced an uncharacterized bromohydrin intermediate,[11] resulting from trans-diaxial addition of H2O to the bromonium ion formed on the less-hindered bottom alkene face. Treatment with base delivered the corresponding synepoxides 4 and 6.[12] Scheme 1 Preparation of scaffold precursors via (a) tetrahydropiperidine synthesis and (b) stereoselective epoxidation reactions and allylic strain control of regiochemistry of epoxide ring opening (general conditions for azidation: NaN3, NH4Cl, MeOH/H2O, 60 °C). ... Both NH and N-acylated epoxy piperidines were prepared as each was expected to react through the divergent conformations shown in Scheme 1 (b) to provide regioisomeric nucleophilic addition products according to the well-established Furst–Plattner principle of trans–diaxial epoxide ring opening.[6a] In this way, allylic strain controlled both the stereochemical and regiochemical outcome of scaffold synthesis, delivering all four isomers of the trans-4,5-disubstituted 2-arylpiperidines desired for the targeted library. In only one stereochemical series was no selectivity obtained: epoxide 6 gave an essentially 1:1 mixture due to competing electronic and stereoelectronic preferences for this epoxide (see Supporting Information for details). However, even in this case, we were able to isolate >700 mg quantities of the desired NH azido alcohols in high purity (as was the case for all 16 of the desired scaffolds) for further diversification. The scaffolds were decorated by removal of the Boc group (if present) followed by either reductive alkylation or N-acylation, and finally, a Cu-induced triazole formation step (Scheme 2).[13] For this initial, speculative screening library, we chose a relatively small number of substituents that represent a range of aliphatic and aromatic diversity at the newly-introduced positions. The total number of compounds targeted in these steps, where R1 and R2 were one of the three substituents shown, was 16 × 9 × 2 = 288 compounds. Following parallel synthesis, the library was prepared for screening by mass-directed HPLC purification, yielding ultimately 268 compounds in >90% purity (UV) and >10 mg quantities. All compounds were characterized by high-resolution mass spectrometry. Scheme 2 Scaffold diversification. A subset of 32 compounds (16 amines and the 16 corresponding amides), comprising four substitution patterns for each of the four stereochemical and constitutional isomers, was subjected to conformational analysis by 1H NMR coupling constant analysis. The data revealed that each functionalized amino-piperidine isomer adopted the same conformation in solution, regardless of the substituents appended to the core. The same held true for the corresponding amides. The conformational profiles of the 8 stereochemical families of the triazol-containing piperidines (4 amines and 4 amides) are shown in Figure 3.[14] Figure 3a shows overlays of each amino-piperidine (blue) paired with the corresponding amido-piperidine (green) counterpart, highlighting the conformational differences achieved through the introduction of 1,3-allylic strain. Three of the four families of amino-piperidines adopted chair forms placing the 2-aryl substituents equatorial. The 2,4-trans family of amino-piperidines exhibited a slight distortion from ideal chair conformation, while still placing the 2-aryl substituents in a pseudoequatorial. The corresponding 2,5-cis and 2,4-trans families of amido-piperidines adopted the opposite chair forms, placing the 2-aromatic substituents axial. However, rather than adopting chair forms placing all three non-hydrogen substituents (Ph, OH, triazole) in axial positions, the 2,5-trans amido-piperidines exhibited twist-like conformations, and the 2,4-cis amido-piperidines adopted boat conformations. Thus, each of the eight compounds in the family presents the piperidine substituents in a unique three-dimensional array. Taken together, these compounds (and by extension the entire library that they represent) comprise a shape-diverse collection with predictable three-dimensional shapes for use in biological screening and structure-activity relationship (SAR) development (Figure 3b). Figure 3 (a) Overlays contrasting amino-piperidine conformation (blue) with the corresponding amido-piperidine conformation (green) for each isomeric pair (NMR). (b) Overlay showing the chemical space occupied by the entire library. For each case: Ar = Ph, R1 ... Through these efforts, we have demonstrated a useful protocol for maximizing the stereochemical diversity in a piperidine library using a limited number of scaffolds and building blocks. This approach features the use of 1,3-allylic strain for controlling both the ring opening of epoxide precursors and, thus, constitutional isomerism, as well as the conformations of the final library members. In so doing, a library of 268 drug-like compounds having predictable conformations has been prepared. The compounds prepared in this work are being scrutinized by high-throughput screening whereas the concepts utilized in the present case are currently being applied to the construction of other heterocyclic libraries.

Potential Drug Abuse Therapeutics Derived from the Hallucinogenic Natural Product Salvinorin A.

Previous structure-activity relationship studies of salvinorin A have shown that modification of the acetate functionality off the C-2 position to a methoxy methyl or methoxy ethyl ether moiety leads to increased potency at KOP receptors. However, the reason for this increase remains unclear. Here we report our efforts towards the synthesis and evaluation of C-2 constrained analogs of salvinorin A. These analogs were evaluated at opioid receptors in radioligand binding experiments as well as in the GTP-γ-S functional assay. One compound, 5, was found to have affinity and potency at κ opioid (KOP) receptors comparable to salvinorin A. In further studies, 5 was found to attenuate cocaine-induced drug seeking behavior in rats comparably to salvinorin A. This finding represents the first example of a salvinorin A analog that has demonstrated anti-addictive capabilities.

Metabolomic Analysis Reveals Novel Isoniazid Metabolites and Hydrazones in Human Urine

Isoniazid (INH) is a first-line drug for tuberculosis control; the side effects of INH are thought to be associated with its metabolism, and this study was designed to globally characterize isoniazid metabolism. Metabolomic strategies were used to profile isoniazid metabolism in humans. Eight known and seven novel INH metabolites and hydrazones were identified in human urine. The novel products included two hydroxylated INH metabolites and five hydrazones. The two novel metabolites were determined as 2-oxo-1,2-dihydro-pyridine-4-carbohydrazide and isoniazid N-oxide. Five novel hydrazones were produced by condensation of isoniazid with keto acids that are intermediates in the metabolism of essential amino acids, namely, leucine and/or isoleucine, lysine, tyrosine, tryptophan, and phenylalanine. This study enhances our knowledge of isoniazid metabolism and disposition and may offer new avenues for investigating INH-induced toxicity.

Dual Catalytic Decarboxylative Allylations of α-Amino Acids and Their Divergent Mechanisms.

The room temperature radical decarboxylative allylation of N-protected α-amino acids and esters has been accomplished via a combination of palladium and photoredox catalysis to provide homoallylic amines. Mechanistic investigations revealed that the stability of the α-amino radical, which is formed by decarboxylation, dictates the predominant reaction pathway between competing mechanisms.

Determination of pKa and Hydration Constants for a Series of α-Keto-Carboxylic Acids Using Nuclear Magnetic Resonance Spectrometry

The determination of the acid-base dissociation constants, and thus the pKa values, of α-keto acids such as pyruvic acid is complex because of the existence of these acids in their hydrated and nonhydrated or oxo state. Equilibria involved in the hydration and dehydration of the α-keto group of pyruvic acid and three other α-keto acids, 3-methyl-2-oxobutanoic acid, 4-methyl-2-oxopentanoic acid, and 2-oxo-2-phenylacetic acid, were investigated by proton and carbon nuclear magnetic resonance spectrometry, at constant ionic strength, 0.15, and 25 °C. Dissociation constants for the oxo (pKa(oxo)) and hydrated (pKa(hyd)) acids of each compound were estimated from the change in the degree of hydration with changes in pH and directly from the changes in chemical shifts of various hydrogen and carbons nuclei with pH. α-Keto acids showed greater hydration in their acidic forms than their carboxylate forms. The degree of hydration was sensitive to steric and electronic/resonance factors. As expected, the oxo forms of the acids were stronger acids compared with their hydrated analogs, and their dissociation constants were also sensitive to steric and electronic factors.