Structural insight into the molecular mechanism of cilofexor binding to the farnesoid X receptor.

Farnesoid X receptor (FXR) is a bile acid-related nuclear receptor and is considered a promising target to treat several liver disorders. Cilofexor is a selective FXR agonist and has already entered phase III trials in primary sclerosing cholangitis (PSC) patients. Pruritis caused by cilofexor treatment is dose dependent. The binding characteristics of cilofexor with FXR and its pruritogenic mechanism remain unclear. In our research, the affinity of cilofexor bound to FXR was detected using an isothermal titration calorimetry (ITC) assay. The binding mechanism between cilofexor and FXR-LBD is explained by the cocrystal structure of the FXR/cilofexor complex. Structural models indicate the possibility that cilofexor activates Mas-related G protein-coupled receptor X4 (MRGPRX4) or G protein-coupled bile acid receptor 1 (GPBAR1), leading to pruritus. In summary, our analyses provide a molecular mechanism of cilofexor binding to FXR and provide a possible explanation for the dose-dependent pruritis of cilofexor.

Isonicotinylation is a histone mark induced by the anti-tuberculosis first-line drug isoniazid.

Isoniazid (INH) is a first-line anti-tuberculosis drug used for nearly 70 years. However, the mechanism underlying the side effects of INH has remained elusive. Here, we report that INH and its metabolites induce a post-translational modification (PTM) of histones, lysine isonicotinylation (Kinic), also called 4-picolinylation, in cells and mice. INH promotes the biosynthesis of isonicotinyl-CoA (Inic-CoA), a co-factor of intracellular isonicotinylation. Mass spectrometry reveals 26 Kinic sites in histones in HepG2 cells. Acetyltransferases CREB-binding protein (CBP) and P300 catalyse histone Kinic, while histone deacetylase HDAC3 functions as a deisonicotinylase. Notably, MNase sensitivity assay and RNA-seq analysis show that histone Kinic relaxes chromatin structure and promotes gene transcription. INH-mediated histone Kinic upregulates PIK3R1 gene expression and activates the PI3K/Akt/mTOR signalling pathway in liver cancer cells, linking INH to tumourigenicity in the liver. We demonstrate that Kinic is a histone acylation mark with a pyridine ring, which may have broad biological effects. Therefore, INH-induced isonicotinylation potentially accounts for the side effects in patients taking INH long-term for anti-tuberculosis therapy, and this modification may increase the risk of cancer in humans.

Crystal structures of two inhibitors in complex with histone lysine demethylase 4D (KDM4D) provide new insights for rational drug design.

Histone lysine demethylase 4D (KDM4D), also known as JMJD2D, plays an important role in cell proliferation and survival and has been associated with several tumor types. KDM4D has emerged as a potential target for the treatment of human cancer. Here, we reported crystal complex structures for two KDM4D inhibitors, OWS [2-(1H-pyrazol-3-yl)isonicotinic acid] and 10r (5-hydroxy-2-methylpyrazolo[1,5-a]pyrido[3,2-e]pyrimidine-3-carbonitrile), which were both determined to 2.0 Å. OWS is a newly discovered KDM4D inhibitor (IC50 = 4.28 µM) and the critical pharmacophores of this compound are confirmed by the complex structure. Compound 10r is a KDM4D inhibitor reported by us previously. To clarify the binding mode in more detail, the crystal structure was determined and the comparison analysis revealed unique interactions that had never been observed before. Overall, our data provide new structural insights for rational design and offer an opportunity for optimization of KDM4D inhibitors.

Synthesis of Highly Potent Anti-Inflammatory Compounds (ROS Inhibitors) from Isonicotinic Acid.

In search of anti-inflammatory compounds, novel scaffolds containing isonicotinoyl motif were synthesized via an efficient strategy. The compounds were screened for their in vitro anti-inflammatory activity. Remarkably high activities were observed for isonicotinates 5-6 and 8a-8b. The compound 5 exhibits an exceptional IC50 value (1.42 ± 0.1 µg/mL) with 95.9% inhibition at 25 µg/mL, which is eight folds better than the standard drug ibuprofen (11.2 ± 1.9 µg/mL). To gain an insight into the mode of action of anti-inflammatory compounds, molecular docking studies were also performed. Decisively, further development and fine tuning of these isonicotinates based scaffolds for the treatment of various aberrations is still a wide-open field of research.

Design, synthesis and evaluation of carbazole derivatives as potential antimicrobial agents.

Five series of novel carbazole derivatives containing an aminoguanidine, dihydrotriazine, thiosemicarbazide, semicarbazide or isonicotinic moiety were designed, synthesised and evaluated for their antimicrobial activities. Most of the compounds exhibited potent inhibitory activities towards different bacterial strains (including one multidrug-resistant clinical isolate) and one fungal strain with minimum inhibitory concentrations (MICs) between 0.5 and 16 µg/ml. Compounds 8f and 9d showed the most potent inhibitory activities (MICs of 0.5-2 µg/ml). Furthermore, compounds 8b, 8d, 8f, 8k, 9b and 9e with antimicrobial activities were not cytotoxic to human gastric cancer cell lines (SGC-7901 and AGS) or a normal human liver cell line (L-02). Structure-activity relationship analyses and docking studies implicated the dihydrotriazine group in increasing the antimicrobial potency and reducing the toxicity of the carbazole compounds. In vitro enzyme activity assays suggested that compound 8f binding to dihydrofolate reductase might account for the antimicrobial effect.

Novel isoniazid derivative as promising antituberculosis agent.

Background: A major focus of tuberculosis drug discovery is aimed at the development of novel antibiotics with activity against drug-resistant strains of Mycobacterium tuberculosis. Results: We have synthesized ten isoniazid derivatives and investigated for antibacterial activity toward M. tuberculosis H37Rv and isoniazid-resistant strain SRI 1369. It was revealed that only one compound, isonicotinic acid (1-methyl-1H-pyrrol-2-ylmethylene)-hydrazide (1), is active toward isoniazid-resistant strain with minimum inhibitory concentration value of 0.14 µM. This compound is not cytotoxic toward human liver cells (HepG2; IC50 >100 µM), demonstrates good permeability in Caco-2 cells. Accordingly to the results of plasma protein binding assay, unbound fraction of compound 1, which potentially exhibits pharmacologic effects, is 57.9%. Conclusion: Therefore, isonicotinic acid (1-methyl-1H-pyrrol-2-ylmethylene)-hydrazide is a promising compound for further preclinical studies.

Inhibitory effects of the phytohormone inhibitors fluridone and inabenfide against Babesia gibsoni in vitro.

Pharmacological options to treat canine babesiosis caused by Babesia gibsoni, are limited. To address this challenge, screening for novel drug candidates and drug targets against B. gibsoni is urgently needed. In this study, we explored the inhibitory effects of two phytohormone inhibitors, fluridone (FLU) and inabenfide (INA), against B. gibsoni in vitro. The half-maximal inhibitory concentration (IC50) values of FLU and INA against B. gibsoni were 60.6 ± 3.4 and 4.3 ± 0.3 µM, respectively. Parasitemia and viability at 24, 48, and 72 h after FLU and INA treatments were significantly lower than those in the control group. The cytotoxicity of FLU and INA was evaluated using the dog-derived Madin-Darby canine kidney (MDCK) cell line; both FLU and INA were less toxic to the MDCK cells than to the control cells. The selectivity index of FLU and INA were higher than 16.5 and 232.6, respectively. In summary, the present study demonstrated that FLU and INA were effective against B. gibsoni infection in vitro and that these compounds might have potential as candidate drugs for the treatment of B. gibsoni.

Pyridinecarboxylic Acid Derivative Stimulates Pro-Angiogenic Mediators by PI3K/AKT/mTOR and Inhibits Reactive Nitrogen and Oxygen Species and NF-κB Activation Through a PPARγ-Dependent Pathway in T. cruzi-Infected Macrophages.

Chagas disease is caused by Trypanosoma cruzi infection and represents an important public health concern in Latin America. Macrophages are one of the main infiltrating leukocytes in response to infection. Parasite persistence could trigger a sustained activation of these cells, contributing to the damage observed in this pathology, particularly in the heart. HP24, a pyridinecarboxylic acid derivative, is a new PPARγ ligand that exerts anti-inflammatory and pro-angiogenic effects. The aim of this work was to deepen the study of the mechanisms involved in the pro-angiogenic and anti-inflammatory effects of HP24 in T. cruzi-infected macrophages, which have not yet been elucidated. We show for the first time that HP24 increases expression of VEGF-A and eNOS through PI3K/AKT/mTOR and PPARγ pathways and that HP24 inhibits iNOS expression and NO release, a pro-inflammatory mediator, through PPARγ-dependent mechanisms. Furthermore, this study shows that HP24 modulates H2O2 production in a PPARγ-dependent manner. It is also demonstrated that this new PPARγ ligand inhibits the NF-κB pathway. HP24 inhibits IKK phosphorylation and IκB-α degradation, as well as p65 translocation to the nucleus in a PPARγ-dependent manner. In Chagas disease, both the sustained increment in pro-inflammatory mediators and microvascular abnormalities are crucial aspects for the generation of cardiac damage. Elucidating the mechanism of action of new PPARγ ligands is highly attractive, given the fact that it can be used as an adjuvant therapy, particularly in the case of Chagas disease in which inflammation and tissue remodeling play an important role in the pathophysiology of this disease.

Antiviral effect of a derivative of isonicotinic acid enisamium iodide (FAV00A) against influenza virus.

With only a single class of antiviral drugs existing for treatment of influenza (neuraminidase inhibitors), the search for novel effective compounds is urgently needed. We evaluated a low molecular mass compound, enisamium iodide (FAV00A), against influenza virus infections in primary differentiated normal human bronchial epithelial (NHBE) cells, and in ferrets. FAV00A (500 µg/ml) markedly inhibited influenza virus replication and reduced viral M-gene expression in NHBE cells. Treatment of ferrets with FAV00A (200 mg/kg once daily for 7 days) initiated 24 h after inoculation with 105 TCID50 of influenza A/Wisconsin/67/2005 (H3N2) virus resulted in a significant decrease in virus titers in the upper respiratory tract. Our data show that FAV00A exhibits an antiviral effect against influenza virus in NHBE cells and provides some benefits in a ferret model. Thus, further Keywords: antiviral agents; enisamium iodide; influenza virus; MDCK cells; NHBE cells; ferrets.

Synthetic nicotinic/isonicotinic thiosemicarbazides: In vitro urease inhibitory activities and molecular docking studies.

Nicotinic and isonicotinic thiosemicarbazide or hydrazine carbothioamides 3-27 were synthesized and the structures of synthetic compounds were elucidated by various spectroscopic techniques such as EI-MS, 1H-, and 13C NMR. Synthetic derivatives were evaluated for their urease inhibitory activity which revealed that except few all derivatives demonstrated excellent inhibition in the range of IC50 values of 1.21-51.42 µM as compared to the standard thiourea (IC50 = 21.25 ±â€¯0.13 µM). Among the twenty-five synthetic derivatives nineteen 1-5, 7, 8, 10, 12, 14-18, 20-22, 24-27 were found to be more active showing IC50 values between 1.13 and 19.74 µM showing superior activity than the standard. Limited structure-activity relationship demonstrated that the positions of substituent as well as position of nitrogen in pyridine ring are very important for inhibitory activity of this class of compound. To verify these interpretations, in silico study was also performed. A good correlation was obtained between the biological evaluation of active compounds and docking study.

New hydrazide-hydrazones of isonicotinic acid: synthesis, lipophilicity and in vitro antimicrobial screening.

This study describes the synthesis, lipophilicity and in vitro antimicrobial assays of 15 new hydrazide-hydrazones of isonicotinic acid. New derivatives were obtained on the basis of the condensation reaction of isonicotinic acid hydrazide with different aromatic aldehydes. The chemical structure of synthesized compounds was confirmed by spectral methods. Experimental lipophilicity of new isonicotinic acid derivatives was determined using reversed-phase thin-layer chromatography. All synthesized compounds were subjected to in vitro antimicrobial assays against reference strains of Gram-positive bacteria, Gram-negative bacteria and fungi belonging to Candida spp. Some of the synthesized hydrazide-hydrazones proved to be significant antibacterial compounds and more potent than commonly used chemotherapeutic agents.

Pharmacological Evaluation of Novel Isonicotinic Carboxamide Derivatives as Potential Anti-Hyperlipidemic and Antioxidant Agents.

Hyperlipidemia and oxidative stress have been implicated as contributing factors to the development of atherosclerosis and cardiovascular diseases (CVDs). Currently, a large number of antihyperlipidemic medications are conveniently available in the market. Nonetheless, the majority of antihyperlipidemics lack the desired safety and efficacy. Thus, the present study was undertaken to evaluate the potential effect of novel N-(benzoylphenyl)pyridine-4-carboxamide and N-(9,10-dioxo-9,10-dihydroanthracenyl)pyridine-4-carboxamide derivatives in controlling hyperlipidemia and oxidative stress using the Triton WR-1339-induced hyperlipidemic rat model for antihyperlipidemic activity and the DPPH radical scavenging assay for antioxidant activity. This study revealed the antihyperlipidemic activities of some of the newly synthesized, novel carboxamide derivatives, mainly C4 and C12 (p < 0.05). The majority of the compounds displayed a relatively low or no DPPH radical scavenging effect, with C20 possessing the best radical scavenging effect (22%) among all. This research opens the door for new potential antihyperlipidemic compounds derived from isonicotinic acid. N-(3-Benzoylphenyl)pyridine-4-carboxamide (C4) was found to have promising lipid-lowering and antioxidant effects, which may create a protective effect against CVDs, by reducing the LDL-C levels and diminishing the generation of reactive oxygen species.

Antimicrobial activity of organometallic isonicotinyl and pyrazinyl ferrocenyl-derived complexes.

Isonicotinyl and pyrazinyl ferrocenyl-derived complexes were prepared using various hydrazides and ferrocenyl aldehydes. Three heterobimetallic complexes were also synthesized from the Schiff base-derived isonicotinyl ferrocene complex using various platinum group metal dimers based on ruthenium, rhodium and iridium. All complexes were evaluated in vitro for antimycobacterial and antiparasitic activity. Against Mycobacterium tuberculosis H37Rv, the platinum group metal complexes showed glycerol-dependent antimycobacterial activity. The antiplasmodial activities against the NF54 chloroquine-sensitive strain of Plasmodium falciparum of some compounds were moderate, while some complexes also showed promising activity against Trichomonas vaginalis. Incorporation of the ferrocenyl-salicylaldimine moiety resulted in enhanced antimicrobial activity compared to the non-ferrocenyl compound in some cases. The bimetallic iridium-ferrocene isonicotinyl complex exhibited superior antitrichomonal activity relative to its organic counterpart, isoniazid. Furthermore, all these compounds, when screened on several normal flora bacteria of humans, showed no effect on the microbiome, emphasizing the selection of these compounds for these pathogens. The promising antimicrobial activities of the complexes thus supports incorporation of ferrocene as part of existing antimicrobial therapies in order to alter their biological activities favorably.

The hydrolytic susceptibility of prochelator BSIH in aqueous solutions.

The prochelator BSIH ((E)-N'-(2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzylidene)isonicotinohydrazide) contains a boronate group that prevents metal coordination until reaction with peroxide releases the iron chelator SIH ((E)-N'-(2-hydroxybenzylidene)isonicotinohydrazide). BSIH exists in aqueous buffer and cell culture media in equilibrium with its hydrolysis products isoniazid and (2-formylphenyl)boronic acid (FBA). The relative concentrations of these species limit the yield of intact SIH available for targeted iron chelation. While the hydrolysis fragments are nontoxic to retinal pigment epithelial cells, these results suggest that modifications to BSIH that improve its hydrolytic stability yet maintain its low inherent cytotoxicity are desirable for creating more efficient prochelators for protection against cellular oxidative damage.

QSAR, docking studies of 1,3-thiazinan-3-yl isonicotinamide derivatives for antitubercular activity.

The enzyme - enoyl acyl carrier protein reductase (enoyl ACP reductase) is a validated target for antitubercular activity. Inhibition of this enzyme interferes with mycolic acid synthesis which is crucial for Mycobacterium tuberculosis cell growth. In the present work 2D and 3D quantitative structure activity relationship (QSAR) studies were carried out on a series of thiazinan-Isoniazid pharmacophore to design newer analogues. For 2D QSAR, the best statistical model was generated using SA-MLR method (r2=0.958, q2=0.922) while 3D QSAR model was derived using the SA KNN method (q2=0.8498). These studies could guide the topological, electrostatic, steric, hydrophobic substitutions around the nucleus based on which the NCEs were designed. Furthermore, molecular docking was performed to gauze the binding affinity of the designed analogues for enoyl ACP reductase enzyme. Amongst all the designed analogues the binding energies of SKS 01 and SKS 05 were found to be -5.267kcal/mol and -5.237kcal/mol respectively which was comparable with the binding energy of the standard Isoniazid (-6.254kcal/mol).

Why Can Unnatural Electron Acceptors Protect Photosynthesizing Organisms but Kill the Others?

The polychlorinated compounds captafol (CPL) and 2,6-dichloroisonicotinic acid (INA) are able to protect plants acting as a fungicide or an inductor of plant resistance, respectively. At the same time, CPL and INA are dangerous for the respiratory organisms, i.e. mammalians, bacteria, and fungi. The high electron-withdrawing ability of these compounds enables them to serve as unnatural electron acceptors in the cellular ambient near to electron transport pathways located in the thylakoid membrane of chloroplasts or in the mitochondrial respiratory chain. Low-energy electron attachment to CPL and INA in vacuo leads to formation of many fragment species mainly at thermal electron energy as it is shown using dissociative electron attachment spectroscopy. On the basis of the experimental findings, assigned with the support of density functional theory calculations it is suggested that the different bioactivity of CPL and INA in respiratory and photosynthetic organisms is due to the interplay between the dissociative electron attachment process and the energies of electrons leaked from the electron transport pathways.

Discovery of N-(3-((1-Isonicotinoylpiperidin-4-yl)oxy)-4-methylphenyl)-3-(trifluoromethyl)benzamide (CHMFL-KIT-110) as a Selective, Potent, and Orally Available Type II c-KIT Kinase Inhibitor for Gastrointestinal Stromal Tumors (GISTs).

c-KIT kinase is a validated drug discovery target for gastrointestinal stromal tumors (GISTs). Clinically used c-KIT kinase inhibitors, i.e., Imatinib and Sunitinib, bear other important targets such as ABL or FLT3 kinases. Here we report our discovery of a more selective c-KIT inhibitor, compound 13 (CHMFL-KIT-110), which completely abolished ABL and FLT3 kinase activity. KinomeScan selectivity profiling (468 kinases) of 13 exhibited a high selectivity (S score (1) = 0.01). 13 displayed great antiproliferative efficacy against GISTs cell lines GIST-T1 and GIST-882 (GI50: 0.021 and 0.043 µM, respectively). In the cellular context, it effectively affected c-KIT-mediated signaling pathways and induced apoptosis as well as cell cycle arrest. In addition, 13 possessed acceptable bioavailability (36%) and effectively suppressed the tumor growth in GIST-T1 cell inoculated xenograft model without apparent toxicity. 13 currently is undergoing extensive preclinical evaluation and might be a potential drug candidate for GISTs.

Substituted 2-(2-aminopyrimidin-4-yl)pyridine-4-carboxylates as potent inhibitors of JumonjiC domain-containing histone demethylases.

BACKGROUND: Aberrant expression of iron(II)- and 2-oxoglutarate-dependent JumonjiC histone demethylases has been linked to cancer. Potent demethylase inhibitors are drug candidates and biochemical tools to elucidate the functional impact of demethylase inhibition. METHODS & RESULTS: Virtual screening identified a novel lead scaffold against JMJD2A with low-micromolar potency in vitro. Analogs were acquired from commercial sources respectively synthesized in feedback with biological testing. Optimized compounds were transformed into cell-permeable prodrugs. A cocrystal x-ray structure revealed the mode of binding of these compounds as competitive to 2-oxoglutarate and confirmed kinetic experiments. Selectivity studies revealed a preference for JMJD2A and JARID1A over JMJD3. CONCLUSION: Virtual screening and rational structural optimization led to a novel scaffold for highly potent and selective JMJD2A inhibitors.

Synthesis and evaluation of antibacterial activity of (R)-2-methylheptyl isonicotinate, a putative naturally occurring bioactive agent.

A putative antibacterial and antifungal compound, (R)-2-methylheptyl isonicotinate, was synthesized via reductive lactone alkylation of (R)-4-methyldihydrofuran-2(3H)-one. Structural characterisation data as well as bioassay results (with Bacillus subtilis or Escherichia coli) contradict those previously reported.

A multifunctional, light-activated prochelator inhibits UVA-induced oxidative stress.

UVA radiation can damage cells and tissues by direct photodamage of biomolecules as well as by initiating metal-catalyzed oxidative stress. In order to alleviate both concerns simultaneously, we synthesized a multifunctional prochelator PC-HAPI (2-((E)-1-(2-isonicotinoylhydrazono)ethyl)phenyl (trans)-3-(2,4-dihydroxyphenyl)acrylate) that contains a trans-(o-hydroxy)cinnamate ester photocleavable protecting group that is cleaved upon UVA exposure to release a coumarin, umbelliferone, and an aroylhydrazone metal chelator, HAPI (N'-[1-(2-hydroxyphenyl)ethyliden]isonicotinoylhydrazide). While the prochelator PC-HAPI exhibits negligible affinity for iron, it responds rapidly to UVA irradiation and converts to an iron-binding chelator that inhibits iron-catalyzed formation of reactive oxygen species and protects cells from UVA damage.