Multitarget Therapeutic Leads for Alzheimer’s Disease: Quinolizidinyl Derivatives of Bi‐ and Tricyclic Systems as Dual Inhibitors of Cholinesterases and β‐Amyloid (Aβ) Aggregation

Multitarget therapeutic leads for Alzheimer’s disease were designed on the models of compounds capable of maintaining or restoring cell protein homeostasis and of inhibiting β‐amyloid (Aβ) oligomerization. Thirty‐seven thioxanthen‐9‐one, xanthen‐9‐one, naphto‐ and anthraquinone derivatives were tested for the direct inhibition of Aβ(1–40) aggregation and for the inhibition of electric eel acetylcholinesterase (eeAChE) and horse serum butyrylcholinesterase (hsBChE). These compounds are characterized by basic side chains, mainly quinolizidinylalkyl moieties, linked to various bi‐ and tri‐cyclic (hetero)aromatic systems. With very few exceptions, these compounds displayed inhibitory activity on both AChE and BChE and on the spontaneous aggregation of β‐amyloid. In most cases, IC₅₀ values were in the low micromolar and sub‐micromolar range, but some compounds even reached nanomolar potency. The time course of amyloid aggregation in the presence of the most active derivative (IC₅₀=0.84 μM ) revealed that these compounds might act as destabilizers of mature fibrils rather than mere inhibitors of fibrillization. Many compounds inhibited one or both cholinesterases and Aβ aggregation with similar potency, a fundamental requisite for the possible development of therapeutics exhibiting a multitarget mechanism of action. The described compounds thus represent interesting leads for the development of multitarget AD therapeutics.     

A Combinatorial Virtual Screening Approach Driving the Synthesis of 2,4-Thiazolidinedione-Based Molecules as New Dual mPGES-1/5-LO Inhibitors

Dual inhibition of microsomal prostaglandin E₂ synthase-1 (mPGES-1) and 5-lipoxygenase (5-LO), two key enzymes involved in pro-inflammatory eicosanoid biosynthesis, represents a new strategy for treating inflammatory disorders. Herein we report the discovery of 2,4-thiazolidinedione-based mPGES-1/5-LO dual inhibitors following a multidisciplinary protocol, involving virtual combinatorial screening, chemical synthesis, and validation of the biological activities for the selected compounds. Following the multicomponent-based chemical route for the decoration of the 2,4-thiazolidinedione core, a large library of virtual compounds was built (∼2.0×10⁴ items) and submitted to virtual screening. Nine selected molecules were synthesized and biologically evaluated, disclosing among them four compounds able to reduce the activity of both enzymes in the mid- and low- micromolar range of activities. These results are of interest for further expanding the chemical diversity around the 2,4-thiazolidinedione central core, facilitating the identification of novel anti-inflammatory agents endowed with a promising and safer pharmacological profile.     

Fluorophore‐Labeled Cyclooxygenase‐2 Inhibitors for the Imaging of Cyclooxygenase‐2 Overexpression in Cancer: Synthesis and Biological Studies

A group of cyclooxygenase‐2 (COX‐2)‐specific fluorescent cancer biomarkers were synthesized by linking the anti‐inflammatory drugs ibuprofen, (S)‐naproxen, and celecoxib to the 7‐nitrobenzofurazan (NBD) fluorophore. In vitro COX‐1/COX‐2 inhibition studies indicated that all of these fluorescent conjugates are COX‐2 inhibitors (IC₅₀ range: 0.19–23.0 μM ) with an appreciable COX‐2 selectivity index (SI≥4.3–444). In this study the celecoxib–NBD conjugate N‐(2‐((7‐nitrobenzo[c][1,2,5]oxadiazol‐4‐yl)amino)ethyl)‐4‐(5‐(p‐tolyl)‐3‐(trifluoromethyl)‐1H‐pyrazol‐1‐yl)benzenesulfonamide ( 14 ), which displayed the highest COX‐2 inhibitory potency and selectivity (COX‐2 IC₅₀=0.19 μM ; SI=443.6), was identified as an impending COX‐2‐specific biomarker for the fluorescence imaging of cancer using a COX‐2‐expressing human colon cancer cell line (HCA‐7).     

Structure‐Based Design of Microsomal Prostaglandin E2 Synthase‐1 (mPGES‐1) Inhibitors using a Virtual Fragment Growing Optimization Scheme

A small library of 2,3‐dihydroxybenzamide‐ and N‐(2,3‐dihydroxyphenyl)‐4‐sulfonamide‐based microsomal prostaglandin E₂ synthase‐1 (mPGES‐1) inhibitors was identified following a step‐by‐step optimization of small aromatic fragments selected to interact in focused regions in the active site of mPGES‐1. During the virtual optimization process, the 2,3‐dihydroxybenzamide moiety was first selected as a backbone of the proposed new chemical entities; the identified compounds were then synthesized and biologically evaluated, identifying derivatives with very promising inhibitory activities in the micromolar range. Subsequent structure‐guided replacement of the 2,3‐dihydroxybenzamide by the N‐(2,3‐dihydroxyphenyl)sulfonamide moiety led to the identification of N‐(2,3‐dihydroxyphenyl)‐4‐biphenylsulfonamide ( 6 ), the most potent small molecule of the series (IC₅₀=0.53±0.04 μm ). The simple synthetic procedure and the possibility of enhancing the potency of this class of inhibitors through additional structural modifications pave the way for further development of new molecules with mPGES‐1‐inhibitory activity, with potential application as anti‐inflammatory and anticancer agents.     

From Ergolines to Indoles: Improved Inhibitors of the Human H3 Receptor for the Treatment of Narcolepsy

Ergolines were recently identified as a novel class of H3 receptor (H3R) inverse agonists. Although their optimization led to drug candidates with encouraging properties for the treatment of narcolepsy, brain penetration remained low. To overcome this issue, ergoline 1 ((6aR,9R,10aR)‐4‐(2‐(dimethylamino)ethyl)‐N‐phenyl‐9‐(pyrrolidine‐1‐carbonyl)‐6,6a,8,9,10,10a‐hexahydroindolo[4,3‐fg]quinoline‐7(4H)‐carboxamide)) was transformed into a series of indole derivatives with high H3R affinity. These new molecules were profiled by simultaneous determination of their brain receptor occupancy (RO) levels and pharmacodynamic (PD) effects in mice. These efforts culminated in the discovery of 15 m ((R)‐1‐isopropyl‐5‐(1‐(2‐(2‐methylpyrrolidin‐1‐yl)ethyl)‐1H‐indol‐4‐yl)pyridin‐2(1H)‐one), which has an ideal profile showing a strong correlation of PD effects with RO, and no measurable safety liabilities. Its desirably short duration of action was confirmed by electroencephalography (EEG) measurements in rats.     

NVP‐BHG712: Effects of Regioisomers on the Affinity and Selectivity toward the EPHrin Family

Erythropoietin‐producing hepatocellular (EPH) receptors are transmembrane receptor tyrosine kinases. Their extracellular domains bind specifically to ephrin A/B ligands, and this binding modulates intracellular kinase activity. EPHs are key players in bidirectional intercellular signaling, controlling cell morphology, adhesion, and migration. They are increasingly recognized as cancer drug targets. We analyzed the binding of NVP‐BHG712 (NVP) to EPHA2 and EPHB4. Unexpectedly, all tested commercially available NVP samples turned out to be a regioisomer (NVPiso) of the inhibitor, initially described in a Novartis patent application. They only differ by the localization of a single methyl group on either one of two adjacent nitrogen atoms. The two compounds of identical mass revealed different binding modes. Furthermore, both in vitro and in vivo experiments showed that the isomers differ in their kinase affinity and selectivity.     

Synthesis of bicyclic alkene-/alkane-bridged nisin mimics by ring-closing metathesis and their biochemical evaluation as lipid II binders: toward the design of potential novel antibiotics

This report describes the design, synthesis, and biochemical evaluation of alkene- and alkane-bridged AB(C)-ring mimics of the lantibiotic nisin. Nisin belongs to a class of natural antimicrobial peptides, and has a unique mode of action: its AB(C)-ring system binds to the pyrophosphate moiety of lipid II. This mode of action was the rationale for the design of smaller nisin-derived peptides to obtain novel potential antibiotics. As a conformational constraint the thioether bridge was mimicked by an alkene- or alkane isostere. The peptides of the linear individual ring precursors were synthesized on solid support or in solution, and cyclized by ring-closing metathesis in solution with overall yields of between 36 and 89 %. The individual alkene-bridged macrocycles were assembled in solution by using carbodiimide-based synthesis protocols for the corresponding AB(C)-ring mimics. These compounds were tested for their binding affinity toward lipid II by evaluation of their potency to inhibit nisin-induced carboxyfluorescein release from large unilamellar vesicles. It was found that these AB(C)-ring mimics were not able to induce membrane leakage; however, they acted by inhibiting nisin-induced carboxyfluorescein release; this indicates their affinity toward lipid II. These results imply that an alkene or alkane moiety is a suitable thioether bridge mimic.     

We Need 2C but Not 2B: Developing Serotonin 2C (5‐HT2C) Receptor Agonists for the Treatment of CNS Disorders

The serotonin 2C (5‐HT_2C) receptor has been identified as a potential drug target for the treatment of a variety of central nervous system (CNS) disorders, such as obesity, substance abuse, and schizophrenia. In this Viewpoint article, recent progress in developing selective 5‐HT_2C agonists for use in treating these disorders is summarized, including the work of our group. Challenges in this field and the possible future directions are described. Homology modeling as a method to predict the binding modes of 5‐HT_2C ligands to the receptor is also discussed. Compared to known ligands, the improved pharmacological profiles of the 2‐phenylcyclopropylmethylamine‐based 5‐HT_2C agonists make them preferred candidates for further studies.     

Progress in the Development of Eukaryotic Elongation Factor 2 Kinase (eEF2K) Natural Product and Synthetic Small Molecule Inhibitors for Cancer Chemotherapy

Eukaryotic elongation factor 2 kinase (eEF2K or Ca²⁺/calmodulin-dependent protein kinase, CAMKIII) is a new member of an atypical α-kinase family different from conventional protein kinases that is now considered as a potential target for the treatment of cancer. This protein regulates the phosphorylation of eukaryotic elongation factor 2 (eEF2) to restrain activity and inhibit the elongation stage of protein synthesis. Mounting evidence shows that eEF2K regulates the cell cycle, autophagy, apoptosis, angiogenesis, invasion, and metastasis in several types of cancers. The expression of eEF2K promotes survival of cancer cells, and the level of this protein is increased in many cancer cells to adapt them to the microenvironment conditions including hypoxia, nutrient depletion, and acidosis. The physiological function of eEF2K and its role in the development and progression of cancer are here reviewed in detail. In addition, a summary of progress for in vitro eEF2K inhibitors from anti-cancer drug discovery research in recent years, along with their structure–activity relationships (SARs) and synthetic routes or natural sources, is also described. Special attention is given to those inhibitors that have been already validated in vivo, with the overall aim to provide reference context for the further development of new first-in-class anti-cancer drugs that target eEF2K.     

N‐Benzyl‐4‐((heteroaryl)methyl)benzamides: A New Class of Direct NADH‐Dependent 2‐trans Enoyl–Acyl Carrier Protein Reductase (InhA) Inhibitors with Antitubercular Activity

Isoniazid (INH) remains one of the cornerstones of antitubercular chemotherapy for drug‐sensitive strains of M. tuberculosis bacteria. However, the increasing prevalence of multidrug‐resistant (MDR) and extensively drug‐resistant (XDR) strains containing mutations in the KatG enzyme, which is responsible for the activation of INH into its antitubercular form, have rendered this drug of little or no use in many cases of drug‐resistant tuberculosis. Presented herein is a novel family of antitubercular direct NADH‐dependent 2‐trans enoyl–acyl carrier protein reductase (InhA) inhibitors based on an N‐benzyl‐4‐((heteroaryl)methyl)benzamide template; unlike INH, these do not require prior activation by KatG. Given their direct InhA target engagement, these compounds should be able to circumvent KatG‐related resistance in the clinic. The lead molecules were shown to be potent inhibitors of InhA and showed activity against M. tuberculosis bacteria. This new family of inhibitors was found to be chemically tractable, as exemplified by the facile synthesis of analogues and the establishment of structure–activity relationships. Furthermore, a co‐crystal structure of the initial hit with the enzyme is disclosed, providing valuable information toward the design of new InhA inhibitors for the treatment of MDR/XDR tuberculosis.     

Improved 2,4‐Diarylthiazole‐Based Antiprion Agents: Switching the Sense of the Amide Group at C5 Leads to an Increase in Potency

Amide derivatives of 2,4‐diarylthiazole‐5‐carboxylic acids were synthesised and tested for efficacy in a cell line model of prion disease. A number of compounds demonstrating antiprion activity were thereby identified from the screening libraries, showing improved potency and reproducibility of results relative to amide derivatives of the related 2,4‐diphenyl‐5‐aminothiazole, which have been documented previously. Thus, 'switching' the sense of the amide bond at thiazole C5 revealed a more promising lead series of potential prion disease therapeutics. Furthermore, 3,5‐diaryl‐1,2,4‐thiadiazoles isolated as by‐products during library synthesis provided a handful of additional examples possessing an antiprion effect, thereby augmenting the set of newly identified active compounds. Evaluation of binding to cellular prion protein (PrP^C) showed only weak affinities at best, suggesting that the newly identified antiprion agents do not mediate their biological effect through direct interaction with PrP^C.     

Medicinal Hypervalent Tellurium Prodrugs Bearing Different Ligands: A Comparative Study of the Chemical Profiles of AS101 and Its Halido Replaced Analogues

Ammonium trichloro (dioxoethylene-O,O′) tellurate (AS101) is a potent immunomodulator prodrug that, in recent years, entered various clinical trials and was tested for a variety of potential therapeutic applications. It has been demonstrated that AS101 quickly activates in aqueous milieu, producing TeOCl₃⁻, which likely represents the pharmacologically active species. Here we report on the study of the activation process of AS101 and of two its analogues. After the synthesis and characterization of AS101 and its derivatives, we have carried out a comparative study through a combined experimental and computational analysis. Based on the obtained results, we describe here, for the first time, the detailed reaction that AS101 and its bromido- and iodido-replaced analogues undergo in presence of water, allowing the conversion of the original molecule to the likely true pharmacophore. Interestingly, moving down in the halogens’ group we observed a higher tendency to react, attributable to the ligands’ effect. The chemical and mechanistic implications of these meaningful differences are discussed.     

Synthesis and Antitumor Activity of Ether Glycerophospholipids Bearing a Carbamate Moiety at the sn‐2 Position: Selective Sensitivity Against Prostate Cancer Cell Lines

Analogues of 1‐O‐hexadecyl‐sn‐3‐glycerophosphonocholine (compounds 1 – 4 ) or sn‐3‐glycerophosphocholine (compound 5 ) bearing a carbamate or dicarbamate moiety at the sn‐2 position were synthesized and evaluated for their antiproliferative activity against cancer cells derived from a variety of tissues. Although all of the compounds are antiproliferative, surprisingly the carbamates ( 1 and 2 ) are more effective against the hormone‐independent cell lines DU145 and PC3 than toward other cancer cell lines we examined. This selectivity was not observed with the dicarbamates ( 3 and 4 ). Phosphocholine carbamate analogue 5 is as effective against the prostate cancer cell lines as the corresponding phosphonocholine analogue 1 . Cell death induced by 2′‐(trimethylammonio)ethyl 4‐hexadecyloxy‐3(R)‐N‐methylcarbamoyl‐1‐butanephosphonate (carbamate analogue 2 ) appeared to be mediated by apoptosis, as assessed by caspase activation and loss of mitochondrial membrane potential. The in vivo activity of 2 was evaluated in a murine prostate cancer xenograft model. Oral and intravenous administration showed that 2 is effective in inhibiting the growth of PC3 tumors in Rag2M mice. Our studies show that the glycerolipid carbamates reported herein represent a class of prostate‐cancer‐selective cytotoxic agents.     

Discovery of 5′′‐Chloro‐N‐[(5,6‐dimethoxypyridin‐2‐yl)methyl]‐2,2′:5′,3′′‐terpyridine‐3′‐carboxamide (MK‐1064): A Selective Orexin 2 Receptor Antagonist (2‐SORA) for the Treatment of Insomnia

The field of small‐molecule orexin antagonist research has evolved rapidly in the last 15 years from the discovery of the orexin peptides to clinical proof‐of‐concept for the treatment of insomnia. Clinical programs have focused on the development of antagonists that reversibly block the action of endogenous peptides at both the orexin 1 and orexin 2 receptors (OX₁R and OX₂R), termed dual orexin receptor antagonists (DORAs), affording late‐stage development candidates including Merck’s suvorexant (new drug application filed 2012). Full characterization of the pharmacology associated with antagonism of either OX₁R or OX₂R alone has been hampered by the dearth of suitable subtype‐selective, orally bioavailable ligands. Herein, we report the development of a selective orexin 2 antagonist (2‐SORA) series to afford a potent, orally bioavailable 2‐SORA ligand. Several challenging medicinal chemistry issues were identified and overcome during the development of these 2,5‐disubstituted nicotinamides, including reversible CYP inhibition, physiochemical properties, P‐glycoprotein efflux and bioactivation. This article highlights structural modifications the team utilized to drive compound design, as well as in vivo characterization of our 2‐SORA clinical candidate, 5′′‐chloro‐N‐[(5,6‐dimethoxypyridin‐2‐yl)methyl]‐2,2′:5′,3′′‐terpyridine‐3′‐carboxamide (MK‐1064), in mouse, rat, dog, and rhesus sleep models.     

The Lab Oddity Prevails: Discovery of Pan‐CDK Inhibitor (R)‐S‐Cyclopropyl‐S‐(4‐{[4‐{[(1R,2R)‐2‐hydroxy‐1‐methylpropyl]oxy}‐5‐(trifluoromethyl)pyrimidin‐2‐yl]amino}phenyl)sulfoximide (BAY 1000394) for the Treatment of Cancer

Lead optimization of a high‐throughput screening hit led to the rapid identification of aminopyrimidine ZK 304709, a multitargeted CDK and VEGF‐R inhibitor that displayed a promising preclinical profile. Nevertheless, ZK 304709 failed in phase I studies due to dose‐limited absorption and high inter‐patient variability, which was attributed to limited aqueous solubility and off‐target activity against carbonic anhydrases. Further lead optimization efforts to address the off‐target activity profile finally resulted in the introduction of a sulfoximine group, which is still a rather unusual approach in medicinal chemistry. However, the sulfoximine series of compounds quickly revealed very interesting properties, culminating in the identification of the nanomolar pan‐CDK inhibitor BAY 1000394, which is currently being investigated in phase I clinical trials.