Design, In silico Screening, Synthesis, Characterisation and DFT-based Electronic Properties of Dihydropyridine-based Molecule as L-type Calcium Channel Blocker.

BACKGROUND: People of all nationalities and social classes are now affected by the growing issue of hypertension. Over time, there has been a consistent rise in the fatality rate. A range of therapeutic compounds, on the other hand, are often used to handle hypertension. OBJECTIVES: The objectives of this study are first to design potential antihypertensive drugs based on the DHP scaffold, secondly, to analyse drug-likeness properties of the ligands and investigate their molecular mechanisms of binding to the model protein Cav1.2 and finally to synthesise the best ligand. MATERIALS AND METHODS: Due to the lack of 3D structures for human Cav1.2, the protein structure was modelled using a homology modelling approach. A protein-ligand complex's strength and binding interaction were investigated using molecular docking and molecular dynamics techniques. DFT-based electronic properties of the ligand were calculated using the M06-2X/ def2- TZVP level of theory. The SwissADME website was used to study the ADMET properties. RESULTS: In this study, a series of DHP compounds (19 compounds) were properly designed to act as calcium channel blockers. Among these compounds, compound 16 showed excellent binding scores (-11.6 kcal/mol). This compound was synthesised with good yield and characterised. To assess the structural features of the synthesised molecule quantum chemical calculations were performed. CONCLUSION: Based on molecular docking, molecular dynamics simulations, and drug-likeness properties of compound 16 can be used as a potential calcium channel blocker.

Discovery of selective Orai channel blockers bearing an indazole or a pyrazole scaffold.

The calcium release activated calcium (CRAC) channel is highly expressed in T lymphocytes and plays a critical role in regulating T cell proliferation and functions including activation of the transcription factor nuclear factor of activated T cells (NFAT), cytokine production and cytotoxicity. The CRAC channel consists of the Orai pore subunit and STIM (stromal interacting molecule) endoplasmic reticulum calcium sensor. Loss of CRAC channel mediated calcium signaling has been identified as an underlying cause of severe combined immunodeficiency (SCID), leading to drastically weakened immunity against infections. Gain-of-function mutations in Orai and STIM have been associated with tubular aggregated myopathy (TAM), a skeletal muscle disease. While a number of small molecules have shown activity in inhibiting the CRAC signaling pathway, the usefulness of those tool compounds is limited by their off-target activity against TRPM4 and TRPM7 ion channels, high lipophilicity, and a lack of understanding of their mechanism of action. We report structure-activity relationship (SAR) studies that resulted in the characterization of compound 4k [1-(cyclopropylmethyl)-N-(3-fluoropyridin-4-yl)-1H-indazole-3-carboxamie] as a fast onset, reversible, and selective CRAC channel blocker. 4k fully blocked the CRAC current (IC50: 4.9 µM) and the nuclear translocation of NFAT at 30 and 10 µM, respectively, without affecting the electrophysiological function of TRPM4 and TRPM7 channels. Computational modeling appears to support its direction binding to Orai proteins that form the transmembrane CRACchannel.

Chiral superficially porous stationary phases for enantiomeric separation of condensed 1,4-dihydropyridine derivatives.

1,4-dihydropyridine (DHP) scaffold occupies a prominent position among all heterocyclic compounds owing to its versatile pharmacological properties, particularly its well-known calcium channel blocking activity. In the quest of developing new calcium channel blockers, fifty seven 5-oxo-hexahydroquinoline (HHQ) derivatives carrying DHP framework in a condensed ring system were recently synthesized as racemic mixtures. Due to their potential as drug candidates, enantiomers arising from the asymmetric center at the C-4 position of the HHQ ring were separated. Four modern columns packed with 2.7 µm superficially porous particles bonded with a chiral selector were used. The chiral selectors were three macrocyclic glycopeptide selectors: vancomycin, teicoplanin, and a macrocyclic derivative called nico. The fourth bonded selector was the dinitrobenzamido-tetrahydrophenanthrenyl derivative called Whelko. The four chromatographic modes were assayed with the mobile phase compositions: reversed phase with acetonitrile/buffer 30/70 %v/v, normal phase with hexane/ethanol 80/20 %v/v, and subcritical fluid chromatography with CO2/methanol 80/20 %v/v at 25 °C. The WhelkoShell column was the most effective in separating this set of 57 compounds. Several enantioresolution factors passed 20 with enantioselectivity ratios higher than 4. Molecular modeling showed that the compounds had a T-shape that fitted well the molecular structure of the WhelkoShell selector in the normal or subcritical modes. Additionally, seven compounds had a second chiral center. The NicoShell column was able to separate all four stereoisomers of these compounds in the reversed phase mode. The preparative production of pure enantiomers of these compounds would be straightforward using the WhelkoShell column in the subcritical mode.

Establishing the Thermodynamic Cards of Dipine Models' Oxidative Metabolism on 21 Potential Elementary Steps.

Dipines are a type of important antihypertensive drug as L-calcium channel blockers, whose core skeleton is the 1,4-dihydropyridine structure. Since the dihydropyridine ring is a key structural factor for biological activity, the thermodynamics of the aromatization dihydropyridine ring is a significant feature parameter for understanding the mechanism and pathways of dipine metabolism in vivo. Herein, 4-substituted-phenyl-2,6-dimethyl-3,5-diethyl-formate-1,4-dihydropyridines are refined as the structurally closest dipine models to investigate the thermodynamic potential of dipine oxidative metabolism. In this work, the thermodynamic cards of dipine models' aromatization on 21 potential elementary steps in acetonitrile have been established. Based on the thermodynamic cards, the thermodynamic properties of dipine models and related intermediates acting as electrons, hydrides, hydrogen atoms, protons, and two hydrogen ions (atoms) donors are discussed. Moreover, the thermodynamic cards are applied to evaluate the redox properties, and judge or reveal the possible oxidative mechanism of dipine models.

Descending to inhibit: Antagonist-induced downward shift of VSD II in TPC2.

In this issue of Structure, Chi et al.1 report structural and functional studies that reveal the inhibition mechanism of the lysosomal two-pore channel TPC2 by the antagonist SG-094, which is of interest for drug development. Antagonist binding induces the downward displacement of the voltage-sensor domain II (VSD II), which is accompanied by asymmetric conformational rearrangements of the entire channel.

Synthesis, biosimulation and pharmacological evaluation of benzimidazole derivatives with antihypertensive multitarget effect.

In this study, we synthesized a series of seven benzimidazole derivatives incorporating the structural acidic framework of angiotensin II (Ang II) type 1 receptor (AT1R) antagonists (ARA-II) employing a three-step reaction sequence. The chemical structures were confirmed by 1H NMR, 13C NMR and mass spectral data. Through biosimulation, compounds 1-7 were identified as computational safe hits, thus, best candidates underwent ex vivo testing against two distinct mechanisms implicated in hypertension: antagonism of the Ang II type 1 receptor and the blockade of calcium channel. Molecular docking studies helped to understand at the molecular level the dual vasorelaxant effects with the recognition sites of the AT1R and the L-type calcium channel. In an in vivo spontaneously hypertensive rat model (SHR), intraperitoneally administration of compound 1 at 20 mg/kg resulted in a 25 % reduction in systolic blood pressure, demonstrating both ex vivo vasorelaxant action and in vivo antihypertensive multitarget efficacy. ©2024 Elsevier.

Cocrystal Prediction of Nifedipine Based on the Graph Neural Network and Molecular Electrostatic Potential Surface.

Nifedipine (NIF) is a dihydropyridine calcium channel blocker primarily used to treat conditions such as hypertension and angina. However, its low solubility and low bioavailability limit its effectiveness in clinical practice. Here, we developed a cocrystal prediction model based on Graph Neural Networks (CocrystalGNN) for the screening of cocrystals with NIF. And scoring 50 coformers using CocrystalGNN. To validate the reliability of the model, we used another prediction method, Molecular Electrostatic Potential Surface (MEPS), to verify the prediction results. Subsequently, we performed a second validation using experiments. The results indicate that our model achieved high performance. Ultimately, cocrystals of NIF were successfully obtained and all cocrystals exhibited better solubility and dissolution characteristics compared to the parent drug. This study lays a solid foundation for combining virtual prediction with experimental screening to discover novel water-insoluble drug cocrystals.

Improving the bioavailability and therapeutic efficacy of felodipine for the control of diabetes-associated atherosclerosis: In vitro and in vivo characterization.

Felodipine has proven to be effective as an atherosclerosis therapy because it increases blood flow to the vessel wall. However, the poor solubility, low bioavailability, and hepatic first-pass metabolism of oral felodipine compromise its therapeutic effectiveness. The study's goal is to create a nasal pH-sensitive hydrogel of felodipine-loaded invasomes (IPHFI) that will improve felodipine's release, permeation, bioavailability, and efficacy as a potential diabetes-associated atherosclerosis therapy. According to the pre-formulation study, the felodipine-loaded invasomes formulation composed of phospholipid (3%w/v), cholesterol (0.16%w/v), ethanol (3%v/v) and cineole (1%v/v) was chosen as the optimum formulation. The optimum formulation was characterized in vitro and then mixed with a mixture of chitosan and glyceryl monooleate to make the IPHFI formulation. The IPHFI formulation enhanced the release and permeation of felodipine by 2.99 and 3-fold, respectively. To assess the efficacy and bioavailability of the IPHFI formulation, it was studied in vivo using an experimental atherosclerosis rat model. Compared to oral free felodipine, the nasal administration of the IPHFI formulation increased the bioavailability by 3.37-fold and decreased the serum cholesterol, triglycerides, LDL, and calcification score by 1.56, 1.53, 1.80, and 1.18 ratios, respectively. Thus, nasal IPHFI formulation may represent a promising diabetes-associated atherosclerosis therapy.

Structural basis for inhibition of the lysosomal two-pore channel TPC2 by a small molecule antagonist.

Two pore channels are lysosomal cation channels with crucial roles in tumor angiogenesis and viral release from endosomes. Inhibition of the two-pore channel 2 (TPC2) has emerged as potential therapeutic strategy for the treatment of cancers and viral infections, including Ebola and COVID-19. Here, we demonstrate that antagonist SG-094, a synthetic analog of the Chinese alkaloid medicine tetrandrine with increased potency and reduced toxicity, induces asymmetrical structural changes leading to a single binding pocket at only one intersubunit interface within the asymmetrical dimer. Supported by functional characterization of mutants by Ca2+ imaging and patch clamp experiments, we identify key residues in S1 and S4 involved in compound binding to the voltage sensing domain II. SG-094 arrests IIS4 in a downward shifted state which prevents pore opening via the IIS4/S5 linker, hence resembling gating modifiers of canonical VGICs. These findings may guide the rational development of new therapeutics antagonizing TPC2 activity.

Novel Scorpion Toxin ω-Buthitoxin-Hf1a Selectively Inhibits Calcium Influx via CaV3.3 and CaV3.2 and Alleviates Allodynia in a Mouse Model of Acute Postsurgical Pain.

Venom peptides have evolved to target a wide range of membrane proteins through diverse mechanisms of action and structures, providing promising therapeutic leads for diseases, including pain, epilepsy, and cancer, as well as unique probes of ion channel structure-function. In this work, a high-throughput FLIPR window current screening assay on T-type CaV3.2 guided the isolation of a novel peptide named ω-Buthitoxin-Hf1a from scorpion Hottentotta franzwerneri crude venom. At only 10 amino acid residues with one disulfide bond, it is not only the smallest venom peptide known to target T-type CaVs but also the smallest structured scorpion venom peptide yet discovered. Synthetic Hf1a peptides were prepared with C-terminal amidation (Hf1a-NH2) or a free C-terminus (Hf1a-OH). Electrophysiological characterization revealed Hf1a-NH2 to be a concentration-dependent partial inhibitor of CaV3.2 (IC50 = 1.18 µM) and CaV3.3 (IC50 = 0.49 µM) depolarized currents but was ineffective at CaV3.1. Hf1a-OH did not show activity against any of the three T-type subtypes. Additionally, neither form showed activity against N-type CaV2.2 or L-type calcium channels. The three-dimensional structure of Hf1a-NH2 was determined using NMR spectroscopy and used in docking studies to predict its binding site at CaV3.2 and CaV3.3. As both CaV3.2 and CaV3.3 have been implicated in peripheral pain signaling, the analgesic potential of Hf1a-NH2 was explored in vivo in a mouse model of incision-induced acute post-surgical pain. Consistent with this role, Hf1a-NH2 produced antiallodynia in both mechanical and thermal pain.

Structural basis for human Cav3.2 inhibition by selective antagonists.

The Cav3.2 subtype of T-type calcium channels has been targeted for developing analgesics and anti-epileptics for its role in pain and epilepsy. Here we present the cryo-EM structures of Cav3.2 alone and in complex with four T-type calcium channel selective antagonists with overall resolutions ranging from 2.8 Å to 3.2 Å. The four compounds display two binding poses. ACT-709478 and TTA-A2 both place their cyclopropylphenyl-containing ends in the central cavity to directly obstruct ion flow, meanwhile extending their polar tails into the IV-I fenestration. TTA-P2 and ML218 project their 3,5-dichlorobenzamide groups into the II-III fenestration and place their hydrophobic tails in the cavity to impede ion permeation. The fenestration-penetrating mode immediately affords an explanation for the state-dependent activities of these antagonists. Structure-guided mutational analysis identifies several key residues that determine the T-type preference of these drugs. The structures also suggest the role of an endogenous lipid in stabilizing drug binding in the central cavity.

Discovery and Structure-Activity Relationship of a Ryanodine Receptor 2 Inhibitor.

Ryanodine receptor 2 (RyR2) is a large Ca2+-release channel in the sarcoplasmic reticulum (SR) of cardiac muscle cells. It serves to release Ca2+ from the SR into the cytosol to initiate muscle contraction. RyR2 overactivation is associated with arrhythmogenic cardiac disease, but few specific inhibitors have been reported so far. Here, we identified an RyR2-selective inhibitor 1 from the chemical compound library and synthesized it from glycolic acid. Synthesis of various derivatives to investigate the structure-activity relationship of each substructure afforded another two RyR2-selective inhibitors 6 and 7, among which 6 was the most potent. Notably, compound 6 also inhibited Ca2+ release in cells expressing the RyR2 mutants R2474S, R4497C and K4750Q, which are associated with cardiac arrhythmias such as catecholaminergic polymorphic ventricular tachycardia (CPVT). This inhibitor is expected to be a useful tool for research on the structure and dynamics of RyR2, as well as a lead compound for the development of drug candidates to treat RyR2-related cardiac disease.

Discovery of α-Obscurine Derivatives as Novel Cav3.1 Calcium Channel Blockers.

Voltage-gated calcium channels (VGCCs), particularly T-type calcium channels (TTCCs), are crucial for various physiological processes and have been implicated in pain, epilepsy, and cancer. Despite the clinical trials of TTCC blockers like Z944 and MK8998, none are currently available on the market. This study investigates the efficacy of Lycopodium alkaloids, particularly as natural product-based TTCC blockers. We synthesized eighteen derivatives from α-obscurine, a lycodine-type alkaloid, and identified five derivatives with significant Cav3.1 blockade activity. The most potent derivative, compound 7, exhibited an IC50 value of 0.19±0.03 µM and was further analyzed through molecular docking, revealing key interactions with Cav3.1. These findings provide a foundation for the structural optimization of Cav3.1 calcium channel blockers and present compound 7 as a promising lead compound for drug development and a tool for chemical biology research.

Design, Synthesis, Molecular Modeling, and Biological Evaluation of Novel Pyrimidine Derivatives as Potential Calcium Channel Blockers.

Pyrimidines play an important role in modern medical fields. They have a wide spectrum of biological activities such as antimicrobial, anticancer, anti-allergic, anti-leishmanial, antioxidant agents and others. Moreover, in recent years, 3,4-dihydropyrimidin-2(1H)ones have attracted researchers to synthesize them via Biginelli reaction and evaluate their antihypertensive activities as bioisosters of Nifedipine, which is a famous calcium channel blocker. Our new target compounds were prepared through one-pot reaction of thiourea 1, ethyl acetoacetate 2 and/or 1H-indole-2-carbaldehyde, 2-chloroquinoline-3-carbaldehyde, 1,3-diphenyl-1H-pyrazole-4-carbaldehyde, 3a-c in acid medium (HCl) yielding pyrimidines 4a-c, which in turn were hydrolyzed to carboxylic acid derivatives 5a-c which were chlorinated by SOCl2 to give acyl chlorides 6a-c. Finally, the latter were reacted with some selected aromatic amines, namely, aniline, p-toluidine and p-nitroaniline, producing amides 7a-c, 8a-c, and 9a-c. The purity of the prepared compounds was examined via TLC monitoring, and structures were confirmed by different spectroscopic techniques such as IR, 1HNMR, 13CNMR, and mass spectroscopy. The in vivo evaluation of the antihypertensive activity revealed that compounds 4c, 7a, 7c, 8c, 9b and 9c had comparable antihypertensive properties with Nifedipine. On the other hand, the in vitro calcium channel blocking activity was evaluated by IC50 measurement and results revealed that compounds 4c, 7a, 7b, 7c, 8c, 9a, 9b, and 9c had comparable calcium channel blocking activity with the reference Nifedipine. Based on the aforementioned biological results, we selected compounds 8c and 9c to be docked onto Ryanodine and dihydropyridine receptors. Furthermore, we developed a structure-activity relationship. The designed compounds in this study show promising activity profiles in reducing blood pressure and as calcium channel blockers, and could be considered as new potential antihypertensive and/or antianginal agents.

Structure-activity relationship of dihydropyridines for rhabdomyosarcoma.

Childhood muscle-related cancer rhabdomyosarcoma is a rare disease with a 50-year unmet clinical need for the patients presented with advanced disease. The rarity of ∼350 cases per year in North America generally diminishes the viability of large-scale, pharmaceutical industry driven drug development efforts for rhabdomyosarcoma. In this study, we performed a large-scale screen of 640,000 compounds to identify the dihydropyridine (DHP) class of anti-hypertensives as a priority compound hit. A structure-activity relationship was uncovered with increasing cell growth inhibition as side chain length increases at the ortho and para positions of the parent DHP molecule. Growth inhibition was consistent across n = 21 rhabdomyosarcoma cell line models. Anti-tumor activity in vitro was paralleled by studies in vivo. The unexpected finding was that the action of DHPs appears to be other than on the DHP receptor (i.e., L-type voltage-gated calcium channel). These findings provide the basis of a medicinal chemistry program to develop dihydropyridine derivatives that retain anti-rhabdomyosarcoma activity without anti-hypertensive effects.

Synthesis of Dihydropyrimidines: Isosteres of Nifedipine and Evaluation of Their Calcium Channel Blocking Efficiency.

Hypertension and cardiovascular diseases related to it remain the leading medical challenges globally. Several drugs have been synthesized and commercialized to manage hypertension. Some of these drugs have a dihydropyrimidine skeleton structure, act as efficient calcium channel blockers, and affect the calcium ions' intake in vascular smooth muscle, hence managing hypertension. The synthesis of such moieties is crucial, and documenting their structure-activity relationship, their evolved and advanced synthetic procedures, and future opportunities in this area is currently a priority. Tremendous efforts have been made after the discovery of the Biginelli condensation reaction in the synthesis of dihydropyrimidines. From the specific selection of Biginelli adducts to the variation in the formed intermediates to achieve target compounds containing heterocylic rings, aldehydes, a variety of ketones, halogens, and many other desired functionalities, extensive studies have been carried out. Several substitutions at the C3, C4, and C5 positions of dihydropyrimidines have been explored, aiming to produce feasible derivatives with acceptable yields as well as antihypertensive activity. The current review aims to cover this requirement in detail.

Artificial intelligence-driven identification of morin analogues acting as CaV1.2 channel blockers: Synthesis and biological evaluation.

Morin is a vasorelaxant flavonoid, whose activity is ascribable to CaV1.2 channel blockade that, however, is weak as compared to that of clinically used therapeutic agents. A conventional strategy to circumvent this drawback is to synthesize new derivatives differently decorated and, in this context, morin-derivatives able to interact with CaV1.2 channels were found by employing the potential of PLATO in target fishing and reverse screening. Three different derivatives (5a-c) were selected as promising tools, synthesized, and investigated in in vitro functional studies using rat aorta rings and rat tail artery myocytes. 5a-c were found more effective vasorelaxant agents than the naturally occurring parent compound and antagonized both electro- and pharmaco-mechanical coupling in an endothelium-independent manner. 5a, the series' most potent, reduced also Ca2+ mobilization from intracellular store sites. Furthermore, 5a≈5c > 5b inhibited Ba2+ current through CaV1.2 channels. However, compound 5a caused also a concentration-dependent inhibition of KCa1.1 channel currents.

Focusing on C-4 position of Hantzsch 1,4-dihydropyridines: Molecular modifications, enantioseparation, and binding mechanism to L- and T-type calcium channels.

1,4-Dihydropyridines (DHPs) represent the blockbuster class of L-type calcium channel blockers that have tremendous therapeutic value against cardiovascular conditions. Due to their abilities to additionally target other subtypes of calcium channels, DHPs are also considered promising molecules for the treatment of neurological and psychiatric disorders. Having been in the market for more than forty years, DHP is one of the most modified scaffolds for the development of novel molecules acting on calcium channels. Taking the chemical structures of approved DHPs into account, it is noteworthy that C-4 position is the least modified part of the ring system. Therefore, in the present study, we focused on this location and carried out various molecular modifications to obtain twelve potential calcium channel blockers with a DHP-based hexahydroquinoline scaffold (DA1-DA12). The whole-cell patch clamp technique applied to analyze the blocking ability of the synthesized compounds on both L- (Cav1.2) and T- (Cav3.2) type calcium channels revealed five blockers with different selectivity profiles. Introducing naphthyl moiety onto the C-4 position of the main scaffold led to the identification of a selective blocker of Cav1.2 (DA8). The benzodioxole-substituted derivative (DA1) was the most potent and selective Cav3.2 inhibitor, therefore, its enantiomers were separated using HPLC on a chiral stationary phase. Retesting single isomers on Cav3.2 revealed that S-enantiomer was mainly responsible for the block. Finally, DA compounds were docked into two generated homology models of L- and T-type calcium channels. Molecular dynamics (MD) simulations and 3D pharmacophore modeling provided further insights into the detailed binding mechanism of DHPs to Cav1.2 as well as to Cav3.2.

Discovery of pimozide derivatives as novel T-type calcium channel inhibitors with little binding affinity to dopamine D2 receptors for treatment of somatic and visceral pain.

T-type Ca2+ channels (T-channels), particularly Cav3.2 and Cav3.1 isoforms, are promising targets for treating various diseases including intractable pain. Given the potent inhibitory activity of pimozide, an antipsychotic, against T-channels, we conducted structure-activity relationship studies of pimozide derivatives, and identified several compounds including 3a, 3s, and 4 that had potency comparable to that of pimozide in inhibiting T-channels, but little binding affinity to dopamine D2 receptors. The introduction of a phenylbutyl group on the benzoimidazole nuclei of pimozide was considered a key structural modification to reduce the binding affinity to D2 receptors. Those pimozide derivatives potently suppressed T-channel-dependent somatic and visceral pain in mice, without causing any motor dysfunctions attributable to D2 receptor blockade, including catalepsy. The present study thus provides an avenue to develop novel selective T-channel inhibitors available for pain management via the structural modification of existing medicines.