Xanthine oxidoreductase inhibition ameliorates high glucose-induced glomerular endothelial injury by activating AMPK through the purine salvage pathway.

Xanthine oxidoreductase (XOR) contributes to reactive oxygen species production. We investigated the cytoprotective mechanisms of XOR inhibition against high glucose (HG)-induced glomerular endothelial injury, which involves activation of the AMP-activated protein kinase (AMPK). Human glomerular endothelial cells (GECs) exposed to HG were subjected to febuxostat treatment for 48 h and the expressions of AMPK and its associated signaling pathways were evaluated. HG-treated GECs were increased xanthine oxidase/xanthine dehydrogenase levels and decreased intracellular AMP/ATP ratio, and these effects were reversed by febuxostat treatment. Febuxostat enhanced the phosphorylation of AMPK, the activation of peroxisome proliferator-activated receptor (PPAR)-gamma coactivator (PGC)-1α and PPAR-α and suppressed the phosphorylation of forkhead box O (FoxO)3a in HG-treated GECs. Febuxostat also decreased nicotinamide adenine dinucleotide phosphate oxidase (Nox)1, Nox2, and Nox4 expressions; enhanced superoxide dismutase activity; and decreased malondialdehyde levels in HG-treated GECs. The knockdown of AMPK inhibited PGC-1α-FoxO3a signaling and negated the antioxidant effects of febuxostat in HG-treated GECs. Despite febuxostat administration, the knockdown of hypoxanthine phosphoribosyl transferase 1 (HPRT1) also inhibited AMPK-PGC-1α-FoxO3a in HG-treated GECs. XOR inhibition alleviates oxidative stress by activating AMPK-PGC-1α-FoxO3a signaling through the HPRT1-dependent purine salvage pathway in GECs exposed to HG conditions.

Baricitinib protects ICIs-related myocarditis by targeting JAK1/STAT3 to regulate Macrophage polarization.

PURPOSE: The emergence of immune checkpoint inhibitors (ICIs) has revolutionized cancer treatment, but these drugs can also cause severe immune-related adverse effects (irAEs), including myocarditis. Researchers have become interested in exploring ways to mitigate this side effect, and one promising avenue is the use of baricitinib, a Janus kinase inhibitor known to have anti-inflammatory properties. This study aimed to examine the potential mechanism by which baricitinib in ICIs-related myocarditis. METHODS: To establish an ICIs-related myocarditis model, BALB/c mice were administered murine cardiac troponin I (cTnI) peptide and anti-mouse programmed death 1 (PD-1) antibodies. Subsequently, baricitinib was administered to the mice via intragastric administration. Echocardiography, HE staining, and Masson staining were performed to evaluate myocardial functions, inflammation, and fibrosis. Immunofluorescence was used to detect macrophages in the cardiac tissue of the mice.In vitro experiments utilized raw264.7 cells to induce macrophage polarization using anti-PD-1 antibodies. Different concentrations of baricitinib were applied to assess cell viability, and the release of pro-inflammatory cytokines was measured. The activation of the JAK1/STAT3 signaling pathway was evaluated through western blot analysis. RESULTS: Baricitinib demonstrated its ability to improve cardiac function and reduce cardiac inflammation, as well as fibrosis induced by ICIs. Mechanistically, baricitinib treatment promoted the polarization of macrophages towards the M2 phenotype. In vitro and in vivo experiments showed that anti-PD-1 promoted the release of inflammatory factors. However, treatment with baricitinib significantly inhibited the phosphorylation of JAK1 and STAT3. Additionally, the use of RO8191 reversed the effects of baricitinib, further confirming our findings. CONCLUSION: Baricitinib demonstrated its potential as a protective agent against ICIs-related myocarditis by modulating macrophage polarization. These findings provide a solid theoretical foundation for the development of future treatments for ICIs-related myocarditis.

One-carbon unit supplementation fuels purine synthesis in tumor-infiltrating T cells and augments checkpoint blockade.

Nucleotides perform important metabolic functions, carrying energy and feeding nucleic acid synthesis. Here, we use isotope tracing-mass spectrometry to quantitate contributions to purine nucleotides from salvage versus de novo synthesis. We further explore the impact of augmenting a key precursor for purine synthesis, one-carbon (1C) units. We show that tumors and tumor-infiltrating T cells (relative to splenic or lymph node T cells) synthesize purines de novo. Shortage of 1C units for T cell purine synthesis is accordingly a potential bottleneck for anti-tumor immunity. Supplementing 1C units by infusing formate drives formate assimilation into purines in tumor-infiltrating T cells. Orally administered methanol functions as a formate pro-drug, with deuteration enabling kinetic control of formate production. Safe doses of methanol raise formate levels and augment anti-PD-1 checkpoint blockade in MC38 tumors, tripling durable regressions. Thus, 1C deficiency can gate antitumor immunity and this metabolic checkpoint can be overcome with pharmacological 1C supplementation.

Molecular docking approach for the design and synthesis of new pyrazolopyrimidine analogs of roscovitine as potential CDK2 inhibitors endowed with pronounced anticancer activity.

Cyclin-dependent kinase 2 (CDK2) is a vital protein for controlling cell cycle progression that is critically associated with various malignancies and its inhibition could offer a convenient therapeutic approach in designing anticancer remedies. Consequently, this study aimed to design and synthesize new CDK2 inhibitors featuring roscovitine as a template model. The purine ring of roscovitine was bioisosterically replaced with the pyrazolo[3,4-d]pyrimidine scaffold, in addition to some modifications in the side chains. A preliminary molecular docking study for the target chemotypes in the CDK2 binding domain revealed their ability to accomplish similar binding patterns and interactions to that of the lead compound roscovitine. Afterwards, synthesis of the new derivatives was accomplished. Then, the initial anticancer screening at a single dose by the NCI revealed that compounds 7a, 9c, 11c, 17a and 17b achieved the highest GI% values reaching up to 150 % indicating their remarkable activity. These derivatives were subsequently selected to undertake five-dose testing, where compounds 7a, 9c, 11c and 17a unveiled the most pronounced activity against almost the full panel with GI50 ranges; 1.41-28.2, 0.116-2.39, 0.578-60.6 and 1.75-42.4 µM, respectively and full panel GI50 (MG-MID); 8.24, 0.6, 2.46 and 6.84 µM, respectively. CDK2 inhibition assay presented compounds 7a and 9c as the most potent inhibitors with IC50 values of 0.262 and 0.281 µM, respectively which are nearly 2.4 folds higher than the reference ligand roscovitine (IC50 = 0.641 µM). Besides, flow cytometric analysis on the most susceptible and safe cell lines depicted that 7a caused cell cycle arrest at G1/S phase in renal cancer cell line (RXF393) while 9c led to cell growth arrest at S phase in breast cancer cell line (T-47D) along with pronounced apoptotic induction in the mentioned cell lines. These findings afforded new anticancer pyrazolo[3,4-d]pyrimidine, roscovitine analogs, acting via CDK2 inhibition.

Model of Chronic Itch in Aged Mice: Beneficial Effects of Drugs Affecting Descending Modulatory Systems.

Pruritus in the elderly, particularly those cases without skin dryness or other identifiable causes, makes treatment challenging due to the lack of evidence regarding the therapeutic effects of antipruritics. This study proposes an age-related alloknesis mouse model for an evaluation system for such cases, and aimed to investigate the effectiveness and mechanisms of action of several drugs commonly used as antipruritics in Japan, utilizing this model. Mice 69-80 weeks old were used as aged mice, and the level of mechanical alloknesis was counted as the number of scratching behaviours in response to innocuous stimuli. Bepotastine, neurotropin, pregabalin, baricitinib, and abrocitinib were used as antipruritics, and yohimbine and methysergide as inhibitors of the descending inhibitory pathway. The findings suggest that mechanical alloknesis in aged mice is a suitable animal model for assessing pruritus in the elderly without xerosis, and pregabalin, neurotropin, baricitinib, and abrocitinib may be effective antipruritics in the elderly through activating both the noradrenergic and serotonergic descending inhibitory pathways. These findings may be useful for the selection of antipruritics for pruritus in the elderly without skin lesions or dryness.

Synthesis and cytotoxicity of novel 6,8,9-trisubstituted purine analogs against liver cancer cells.

A series of novel 6-(substituted phenyl piperazine)-8-(4-substituted phenyl)-9-cyclopentyl purines, 10-51, were synthesized by a four-step synthesis, achieving an overall yield of about 43 %. The reaction conditions were effectively optimized, and the final products were obtained with high purity and yield in all synthesis steps. The synthesized nucleobases were evaluated for their in vitro cytotoxic activities on selected human cancer cell lines (HUH7 (liver), HCT116 (colon), and MCF7 (breast)) using the Sulforhodamine B (SRB) assay. Among these analogs, compounds bearing 4-trifluoromethyl phenyl (19, 20 and 21), 4-methoxy phenyl (27) and 4-fluoro phenyl (34) substitutions at C-8 of purine were the most potent, and they were also analyzed in drug-resistance and drug-sensitive hepatocellular cancer cell (HCC) panels. Compound 19 displayed remarkable anticancer activities (IC50 = 2.9-9.3 µM) against Huh7, FOCUS, SNU475, SNU182, HepG2, and Hep3B cells compared to the positive control, Fludarabine. Additionally, the pharmacological properties and toxicity profiles of the molecules were investigated computationally by the Swiss-ADME and Pro-Tox II online tools, respectively. Results showed that our compounds have favorable physicochemical characteristics for oral bioavailability and do not reveal any toxicity endpoints such as carcinogenicity, immunotoxicity, mutagenicity, or cytotoxicity.

Discovery of N-(4-((6-(3,5- Dimethoxyphenyl)-9H-purine derivatives as irreversible covalent FGFR inhibitors.

Fibroblast growth factor receptor (FGFR) is an attractive target for cancer therapy, but existing FGFR inhibitors appear to hardly meet the demand for clinical application. Herein, a number of irreversible covalent FGFR inhibitors were designed and synthesized by selecting several five- and six-membered azaheterocycles as parent scaffold with different substituents to take over the hydrophobic region in the active pocket of FGFR proteins. Among the resulting target compounds, III-30 showed the most potent effect on enzyme activity inhibition and anti-proliferative activity against the tested cancer cell lines. Significantly, III-30 could inhibit the enzyme activity by achieving irreversible covalent binding with FGFR1 and FGFR4 proteins. It could also regulate FGFR-mediated signaling pathway and mitochondrial apoptotic pathway to promote cancer cell apoptosis and inhibit cancer cell invasion and metastasis. Moreover, III-30 had a good metabolic stability and showed relatively potent anti-tumor activity in the MDA-MB-231 xenograft tumor mice model.

Purine-Doped g-C3N4-Modified Fabrics for Personal Protective Masks with Rapid and Sustained Antibacterial Activity.

Protective masks are critical to impeding microorganism transmission but can propagate infection via pathogen buildup and face touching. To reduce this liability, we integrated electrospun photocatalytic graphitic carbon nitride (g-C3N4) nanoflakes into standard surgical masks to confer a self-sanitization capacity. By optimizing the purine/melamine precursor ratio during synthesis, we reduced the g-C3N4 band gap from 2.92 to 2.05 eV, eliciting a 4× increase in sterilizing hydrogen peroxide production under visible light. This narrower band gap enables robust photocatalytic generation of reactive oxygen species from environmental and breath humidity to swiftly eliminate accumulated microbes. Under ambient sunlight, the g-C3N4 nanocomposite mask layer achieved a 97% reduction in the bacterial viability during typical use. Because the optimized band gap also allows photocatalytic activity under shadowless lamp illumination, the self-cleaning functionality could mitigate infection risk from residual pathogens in routine hospital settings. Both g-C3N4 and polycaprolactone demonstrate favorable biocompatibility and biodegradability, making this approach preferable over current commercially available metal-based options. Given the abundance and low cost of these components, this scalable approach could expand global access to reusable self-sanitizing protective masks, serving as a sustainable public health preparedness measure against future pandemics, especially in resource-limited settings.

Novel JAK Inhibitors to Reduce Graft-Versus-Host Disease after Allogeneic Hematopoietic Cell Transplantation in a Preclinical Mouse Model.

Allogeneic hematopoietic cell transplantation (allo-HCT) is a highly effective, well-established treatment for patients with various hematologic malignancies and non-malignant diseases. The therapeutic benefits of allo-HCT are mediated by alloreactive T cells in donor grafts. However, there is a significant risk of graft-versus-host disease (GvHD), in which the donor T cells recognize recipient cells as foreign and attack healthy organs in addition to malignancies. We previously demonstrated that targeting JAK1/JAK2, mediators of interferon-gamma receptor (IFNGR) and IL-6 receptor signaling, in donor T cells using baricitinib and ruxolitinib results in a significant reduction in GvHD after allo-HCT. Furthermore, we showed that balanced inhibition of JAK1/JAK2 while sparing JAK3 is important for the optimal prevention of GvHD. Thus, we have generated novel JAK1/JAK2 inhibitors, termed WU derivatives, by modifying baricitinib. Our results show that WU derivatives have the potential to mitigate GvHD by upregulating regulatory T cells and immune reconstitution while reducing the frequencies of antigen-presenting cells (APCs) and CD80 expression on these APCs in our preclinical mouse model of allo-HCT. In addition, WU derivatives effectively downregulated CXCR3 and T-bet in primary murine T cells. In summary, we have generated novel JAK inhibitors that could serve as alternatives to baricitinib or ruxolitinib.

Purines enrich root-associated Pseudomonas and improve wild soybean growth under salt stress.

The root-associated microbiota plays an important role in the response to environmental stress. However, the underlying mechanisms controlling the interaction between salt-stressed plants and microbiota are poorly understood. Here, by focusing on a salt-tolerant plant wild soybean (Glycine soja), we demonstrate that highly conserved microbes dominated by Pseudomonas are enriched in the root and rhizosphere microbiota of salt-stressed plant. Two corresponding Pseudomonas isolates are confirmed to enhance the salt tolerance of wild soybean. Shotgun metagenomic and metatranscriptomic sequencing reveal that motility-associated genes, mainly chemotaxis and flagellar assembly, are significantly enriched and expressed in salt-treated samples. We further find that roots of salt stressed plants secreted purines, especially xanthine, which induce motility of the Pseudomonas isolates. Moreover, exogenous application for xanthine to non-stressed plants results in Pseudomonas enrichment, reproducing the microbiota shift in salt-stressed root. Finally, Pseudomonas mutant analysis shows that the motility related gene cheW is required for chemotaxis toward xanthine and for enhancing plant salt tolerance. Our study proposes that wild soybean recruits beneficial Pseudomonas species by exudating key metabolites (i.e., purine) against salt stress.

Serum starvation is as efficient as roscovitine on the cycle synchronization in G0/G1 of red-rumped agouti fibroblasts.

Fibroblast cycle synchronization in G0/G1 is an essential step for nuclear reprogramming by cloning or induced cells to pluripotency. Considering the diversity among rodents and the ecological and scientific importance of these animals, we compared the contact inhibition, serum starvation, and 10 µM of roscovitine as methods of synchronization of red-rumped agouti fibroblasts. The effects of each protocol were evaluated on the percentage of cycle phase, morphology, viability, and apoptosis levels. The results showed that culturing the cells to serum starvation for 24 h (75.9%), 48 h (81.6%), 72 h (86.2%), 96 h (84.0%), and 120 h (83.7%) yielded a significantly higher percentage of cells arrested in the G0/G1 (P < 0.05) phase than cells not subjected to any cell cycle synchronization method (31.4%). Also, this effect was not different between the times of 48 and 120 h (P > 0.05). A similar response was observed for cells cultured with roscovitine for 12 h (86.9%), 24 h (74.8%), and 48 h (81.7%), with a higher percentage of synchronized cells in G0/G1 compared to cells not submitted to any synchronization treatment (52.2%). Nevertheless, this effect was best evidenced at 12 h (P < 0.05). Also, the contact inhibition for 24-120 h could not synchronize cells in G0/G1, with values ranging from 70.9 to 77.9% (P > 0.05). Moreover, no difference was observed for morphology, viability, and apoptosis levels in any synchronization method (P > 0.05). Therefore, serum starvation is as efficient as roscovitine on cycle synchronization in G0/G1 of red-rumped agouti fibroblasts.

Spermidine attenuates monocrotaline-induced pulmonary arterial hypertension in rats by inhibiting purine metabolism and polyamine synthesis-associated vascular remodeling.

Ensuring the homeostatic integrity of pulmonary artery endothelial cells (PAECs) is essential for combatting pulmonary arterial hypertension (PAH), as it equips the cells to withstand microenvironmental challenges. Spermidine (SPD), a potent facilitator of autophagy, has been identified as a significant contributor to PAECs function and survival. Despite SPD's observed benefits, a comprehensive understanding of its protective mechanisms has remained elusive. Through an integrated approach combining metabolomics and molecular biology, this study uncovers the molecular pathways employed by SPD in mitigating PAH induced by monocrotaline (MCT) in a Sprague-Dawley rat model. The study demonstrates that SPD administration (5 mg/kg/day) significantly corrects right ventricular impairment and pathological changes in pulmonary tissues following MCT exposure (60 mg/kg). Metabolomic profiling identified a purine metabolism disorder in MCT-treated rats, which SPD effectively normalized, conferring a protective effect against PAH progression. Subsequent in vitro analysis showed that SPD (0.8 mM) reduces oxidative stress and apoptosis in PAECs challenged with Dehydromonocrotaline (MCTP, 50 µM), likely by downregulating purine nucleoside phosphorylase (PNP) and modulating polyamine biosynthesis through alterations in S-adenosylmethionine decarboxylase (AMD1) expression and the subsequent production of decarboxylated S-adenosylmethionine (dcSAM). These findings advocate SPD's dual inhibitory effect on PNP and AMD1 as a novel strategy to conserve cellular ATP and alleviate oxidative injuries, thus providing a foundation for SPD's potential therapeutic application in PAH treatment.

Differential effects of benzo[a]pyrene exposure on glutathione and purine metabolism in keratinocytes: Dose-dependent and UV co-exposure effects.

Polycyclic aromatic hydrocarbons with the key substance benzo[a]pyrene (B[a]P) are widespread pollutants in the environment and at working places. Nonetheless, the exact underlying mechanisms of toxicological effects caused by B[a]P especially in absence and presence of UV irradiation remain uncertain. This study examines variations in exposure conditions: low B[a]P (4 nM), low B[a]P + UV and high B[a]P (4 µM), selected based on pertinent cytotoxicity assessments. Following cell viability evaluations post-treatment with varied B[a]P concentrations and UV irradiation, the identified concentrations underwent detailed metabolomic analysis via gas chromatography-mass spectrometry. Subsequently, resulting changes in metabolic profiles across these distinct exposure groups are comprehensively compared. Chemometric analyses showed modest regulation of metabolites after low B[a]P exposure compared to control conditions. High B[a]P and low B[a]P + UV exposure significantly increased regulation of metabolic pathways, indicating that additional UV irradiation plus low B[a]P is as demanding for the cells as higher B[a]P treatment alone. Further analysis revealed exposure-dependent regulation of glutathione-important for oxidative defence-and purine metabolism-important for DNA base synthesis. Only after low B[a]P, oxidative defence appeared to be able to compensate for B[a]P-induced perturbations of the oxidative homeostasis. In contrast, purine metabolism already responded towards adversity at low B[a]P. The metabolomic results give an insight into the mechanisms leading to the toxic response and confirm the strong effects of co-exposure on oxidative defence and DNA repair in the model studied.

Cananga oil inhibits Salmonella infection by mediating the homeostasis of purine metabolism and the TCA cycle.

ETHNOPHARMACOLOGY RELEVANCE: Cananga oil (CO) is derived from the flowers of the traditional medicinal plant, the ylang-ylang tree. As a traditional antidepressant, CO is commonly utilized in the treatment of various mental disorders including depression, anxiety, and autism. It is also recognized as an efficient antibacterial insecticide, and has been traditionally utilized to combat malaria and acute inflammatory responses resulting from bacterial infections both in vitro and in vivo. AIM OF THE STUDY: The objective of this study is to comprehensively investigate the anti-Salmonella activity and mechanism of CO both in vitro and in vivo, with the expectation of providing feasible strategies for exploring new antimicrobial strategies and developing novel drugs. METHODS: The in vitro antibacterial activity of CO was comprehensively analyzed by measuring MIC, MBC, growth curve, time-killing curve, surface motility, biofilm, and Live/dead bacterial staining. The analysis of the chemistry and active ingredients of CO was conducted using GC-MS. To examine the influence of CO on the membrane homeostasis of Salmonella, we conducted utilizing diverse techniques, including ANS, PI, NPN, ONPG, BCECF-AM, DiSC3(5), and scanning electron microscopy (SEM) analysis. In addition, the antibacterial mechanism of CO was analyzed and validated through metabolomics analysis. Finally, a mouse infection model of Salmonella typhimurium was established to evaluate the toxic side effects and therapeutic effects of CO. RESULTS: The antibacterial effect of CO is the result of the combined action of the main chemical components within its six (palmitic acid, α-linolenic acid, stearic acid, benzyl benzoate, benzyl acetate, and myristic acid). Furthermore, CO disrupts the balance of purine metabolism and the tricarboxylic acid cycle (TCA cycle) in Salmonella, interfering with redox processes. This leads to energy metabolic disorders and oxidative stress damage within the bacteria, resulting in bacterial shock, enhanced membrane damage, and ultimately bacterial death. It is worth emphasizing that CO exerts an effective protective influence on Salmonella infection in vivo within a non-toxic concentration range. CONCLUSION: The outcomes indicate that CO displays remarkable anti-Salmonella activity both in vitro and in vivo. It triggers bacterial death by disrupting the balance of purine metabolism and the TCA cycle, interfering with the redox process, making it a promising anti-Salmonella medication.

Duvelisib: A comprehensive profile.

Duvelisib (DUV) is chemically named as (S)-3-(1-((9H-Purin-6-yl)amino)ethyl)-8-chloro-2-phenylisoquinolin-1(2H)-one. It is a novel drug with a small molecular weight and characterized by dual phosphoinositide-3-kinase (PI3K)- and PI3K-inhibitory activity. The Food and Drug Administration (FDA) recently approved DUV for the management of small lymphocytic lymphoma (SLL) and relapsed or refractory chronic lymphocytic leukemia (CLL) in adult patients. DUV is marketed under the brand name of Copiktra® (Verastem, Inc., Needham, MA, USA). This chapter provides a critical extensive review of the literature, the description of DUV in terms of its names, formulae, elemental composition, appearance, and use in the treatment of CLL, SLL, and follicular lymphoma. The chapter also describes the methods for preparation of DUV, its physical-chemical properties, analytical methods for its determination, pharmacological properties, and dosing information.

Local and Sustained Baricitinib Delivery to the Skin through Injectable Hydrogels Containing Reversible Thioimidate Adducts.

Janus kinase (JAK) inhibitors are approved for many dermatologic disorders, but their use is limited by systemic toxicities including serious cardiovascular events and malignancy. To overcome these limitations, injectable hydrogels are engineered for the local and sustained delivery of baricitinib, a representative JAK inhibitor. Hydrogels are formed via disulfide crosslinking of thiolated hyaluronic acid macromers. Dynamic thioimidate bonds are introduced between the thiolated hyaluronic acid and nitrile-containing baricitinib for drug tethering, which is confirmed with 1H and 13C nuclear magnetic resonance (NMR). Release of baricitinib is tunable over six weeks in vitro and active in inhibiting JAK signaling in a cell line containing a luciferase reporter reflecting interferon signaling. For in vivo activity, baricitinib hydrogels or controls are injected intradermally into an imiquimod-induced mouse model of psoriasis. Imiquimod increases epidermal thickness in mice, which is unaffected when treated with baricitinib or hydrogel alone. Treatment with baricitinib hydrogels suppresses the increased epidermal thickness in mice treated with imiquimod, suggesting that the sustained and local release of baricitinib is important for a therapeutic outcome. This study is the first to utilize a thioimidate chemistry to deliver JAK inhibitors to the skin through injectable hydrogels, which has translational potential for treating inflammatory disorders.

Optical Control of Adenosine A2A Receptor Using Istradefylline Photosensitivity.

In recent years, there has been growing interest in the potential therapeutic use of inhibitors of adenosine A2A receptors (A2AR) for the treatment of neurodegenerative diseases and cancer. Nevertheless, the widespread expression of A2AR throughout the body emphasizes the importance of temporally and spatially selective ligands. Photopharmacology is an emerging strategy that utilizes photosensitive ligands to attain high spatiotemporal precision and regulate the function of biomolecules using light. In this study, we combined photochemistry and cellular and in vivo photopharmacology to investigate the light sensitivity of the FDA-approved antagonist istradefylline and its potential use as an A2AR photopharmacological tool. Our findings reveal that istradefylline exhibits rapid trans-to-cis isomerization under near-UV light, and prolonged exposure results in the formation of photocycloaddition products. We demonstrate that exposure to UV light triggers a time-dependent decrease in the antagonistic activity of istradefylline in A2AR-expressing cells and enables real-time optical control of A2AR signaling in living cells and zebrafish. Together, these data demonstrate that istradefylline is a photoinactivatable A2AR antagonist and that this property can be utilized to perform photopharmacological experiments in living cells and animals.

Synthesis of 6,8-diaminopurines via acid-induced cascade cyclization of 5-aminoimidazole precursors and preliminary anticancer evaluation.

Inspired by the pharmacological interest generated by 6-substituted purine roscovitine for cancer treatment, 5-aminoimidazole-4-carboxamidine precursors containing a cyanamide unit were prepared by condensation of 5-amino-N-cyanoimidazole-4-carbimidoyl cyanides with a wide range of primary amines. When these amidine precursors were combined with acids, a fast cascade cyclization occurred at room temperature, affording new 6,8-diaminopurines with the N-3 and N-6 substituents changed relatively to the original positions they occupied in the amidine and imidazole moieties of precursors. The efficacy and wide scope of this method was well demonstrated by an easy and affordable synthesis of 22 6,8-diaminopurines decorated with a wide diversity of substituents at the N-3 and N-6 positions of the purine ring. Preliminary in silico and in vitro assessments of these 22 compounds were carried out and the results showed that 13 of these tested compounds not only exhibited IC50 values between 1.4 and 7.5 µM against the colorectal cancer cell line HCT116 but also showed better binding energies than known inhibitors in docking studies with different cancer-related target proteins. In addition, good harmonization observed between in silico and in vitro results strengthens and validates this preliminary evaluation, suggesting that these novel entities are good candidates for further studies as new anticancer agents.

L-arginine metabolism inhibits arthritis and inflammatory bone loss.

OBJECTIVES: To investigate the effect of the L-arginine metabolism on arthritis and inflammation-mediated bone loss. METHODS: L-arginine was applied to three arthritis models (collagen-induced arthritis, serum-induced arthritis and human TNF transgenic mice). Inflammation was assessed clinically and histologically, while bone changes were quantified by µCT and histomorphometry. In vitro, effects of L-arginine on osteoclast differentiation were analysed by RNA-seq and mass spectrometry (MS). Seahorse, Single Cell ENergetIc metabolism by profilIng Translation inHibition and transmission electron microscopy were used for detecting metabolic changes in osteoclasts. Moreover, arginine-associated metabolites were measured in the serum of rheumatoid arthritis (RA) and pre-RA patients. RESULTS: L-arginine inhibited arthritis and bone loss in all three models and directly blocked TNFα-induced murine and human osteoclastogenesis. RNA-seq and MS analyses indicated that L-arginine switched glycolysis to oxidative phosphorylation in inflammatory osteoclasts leading to increased ATP production, purine metabolism and elevated inosine and hypoxanthine levels. Adenosine deaminase inhibitors blocking inosine and hypoxanthine production abolished the inhibition of L-arginine on osteoclastogenesis in vitro and in vivo. Altered arginine levels were also found in RA and pre-RA patients. CONCLUSION: Our study demonstrated that L-arginine ameliorates arthritis and bone erosion through metabolic reprogramming and perturbation of purine metabolism in osteoclasts.

Anticancer effect of covalent purine-containing EGFR TKI, ZZC4 and its mechanism of action through network pharmacology.

AIMS: Epidermal growth factor receptor (EGFR) has been documented in many malignancies as participating in the progression of cancer cells. Here, we present a novel EGFR tyrosine kinase inhibitor, ZZC4, and examine its effect on cancer cell proliferation, migration, and tumor-bearing xenograft models. MAIN METHODS: The antiproliferative effect of ZZC4 was assessed in vitro by MTT assay, colony formation, and wound healing assay and in vivo with tumor-bearing xenograft nude mice. Further, Western blotting analysis and computational network pharmacology were used to explore and understand the mechanism of ZZC4. KEY FINDINGS: The results showed that ZZC4 potently inhibited the proliferation of lung, breast, and melanoma cells, and was more sensitive to lung cancer cells HCC827, H1975, and breast cancer cell T47D. In vitro findings were corroborated in vivo as results showed the suppressive effect of ZZC4 on HCC827 and H1975 tumor growth. Western blotting analysis confirmed that ZZC4 is an effective inhibitor of the EGFR pathways as it down-regulated p-EGFR, p-Akt, and p-MAPK. Computational molecular docking confirmed the strong binding affinity between ZZC4 and EGFR. Moreover, network pharmacology suggested that ZZC4 might play a suppressive role in the progression of malignancies with EGFR/PI-3K/Akt axis dysregulation or in cancer-related drug resistance. SIGNIFICANCE: Our study showed that ZZC4 is an anticancer drug candidate.