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Interaction of diclofenac and indomethacin with lysozyme was studied using several spectroscopic and molecular docking methods. Difference UV-visible spectra showed that the absorption profile of lysozyme changed when both diclofenac and indomethacin were mixed with the former. The sequential addition of both drugs to the lysozyme solution caused the decrease of the intrinsic fluorescence of the latter, however, when the data were corrected for inner filter effect, an enhancement in the fluorescence of lysozyme was detected. Accordingly, the fluorescence enhancement data were analyzed using Benesi-Hildebrand equation. Both, diclofenac and indomethacin showed good interaction with lysozyme, although, the association constants of indomethacin were nearly two-fold higher as compared to that of diclofenac. The binding was slightly more spontaneous in case of indomethacin and the major forces involved in the binding of both drugs with lysozyme were hydrogen bonding and hydrophobic interactions. Secondary structural analysis revealed that both drugs partially unfolded lysozyme. Results obtained through molecular docking were also in good agreement with the experimental outcomes. Both, diclofenac and indomethacin, are bounded at the same site inside lysozyme which is located in the big hydrophobic cavity of the protein., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Mitochondrion has appeared as one of the important targets for anti-cancer therapy. Subsequently, small molecule anti-cancer drugs are directed to the mitochondria for improved therapeutic efficacy. However, simultaneous imaging and impairing mitochondria by a single probe remained a major challenge. To address this, herein Chimeric Small Molecules (CSMs) encompassing drugs, fluorophore and mitochondria homing moiety were designed and synthesized through a concise strategy. Screening of the CSMs in a panel of cancer cell lines (HeLa, MCF7, A549, and HCT-116) revealed that one of the CSMs comprising Indomethacin V exhibited remarkable cervical cancer cell (HeLa) killing (IC 50 =0.97 μM). This lead CSM homed into the mitochondria of HeLa cells within 1 h followed by mitochondrial damage and reactive oxygen species (ROS) generation. This novel Indomethacin V-based CSM-mediated mitochondrial damage induced programmed cell death (apoptosis). We anticipate these CSMs can be used as tools to understand the drug effects in organelle chemical biology in diseased states., (© 2023 Wiley-VCH GmbH.)

Background: There are patents available related to fermented food and beverages which enhance to human health. Citrus limetta (Mosambi) has a high content of flavonoids and exhibits antioxidant activity, which could stimulate the digestive system and be useful for gastroprotective activity. It supports digestion by neutralizing the acidic digestive juices and reducing gastric acidity., Objective: This study explored the potential of using waste peel extract from Citrus limetta to prevent ulcers. The study specifically sought to assess the anti-ulcer properties of fermented and non-fermented extracts and compare them. Further, the study looked at the potential benefits of treating or preventing ulcers with Citrus limetta waste peels and whether fermentation affected the efficacy of the treatment., Methods: Thirty female Wistar albino rats were equally distributed into five different groups. Group 1 received distilled water (20 ml/kg/b.w); Group 2 received indomethacin (mg/kg/b.w); Group 3 received omeprazole (20 mg/kg/b.w); Group 4 received aqueous extract of Mosambi peel (400 mg/kg/b.w) and Group 5 received fermented product of extract of Mosambi peel (400 mg/kg/b.w)., Results: Findings explored that, compared to non-fermented citrus fruit juice, biofermented exhibited less gastric volume (1.58 ± 0.10 ml vs. 1.8 ± 0.14 ml), reduced MDA levels (355.23 ± 100.70 μmol/mg protein vs. 454.49 ± 155.88 μmol/mg protein), and low ulcer index (0.49 ± 0.07 vs. 0.72 ± 0.14)., Conclusion: The results suggest that the bio-fermented product of Citrus limetta peel has better anti-ulcer potential against peptic ulcer induced by indomethacin in Wistar albino rats compared to non-fermented., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The potentiation of pharmacological effects can be achieved through several strategies, such as the association of substances and delivery in nanostructured systems. In practice, potentiation can be measured by the law of mass action and joint evaluation of the combination index (CI) and dose-response curves. In this context, this study aimed to evaluate the anti-inflammatory effect of the association of β-caryophyllene and indomethacin in the free form and delivered in nanoemulsions using the in vitro model of LPS-stimulated murine macrophage. The results indicated potentiation of the anti-inflammatory effect of nanoemulsified substances compared to free substances, as well as synergistic action between the sesquiterpene and the selected NSAID. In comparison, the association of β-caryophyllene and indomethacin in the free form inhibited the production of nitric oxide by 50% at 48.60 µg/mL (CI = 0.21), while the nanoemulsified association of these substances resulted in an IC 50 of 1.45 µg/mL (CI = 0.14). In parallel, cytotoxicity assays on HaCaT and MRC-5 cell lines demonstrated the safety of IC 50 -equivalent concentrations of the anti-inflammatory action, and no irritating effects on the chorioallantoic membrane of embryonated eggs were observed (HET-CAM assay). The results suggest that β-caryophyllene may be an alternative to replace an inert oily core in nanoemulsion systems when anti-inflammatory effects are desirable., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Adolescent intermittent ethanol (AIE) exposure causes long-term changes in the brain and behavior of adult male rodents, including persistent induction of innate immune pathways, reductions in hippocampal neurogenic and forebrain cholinergic neuronal markers, and reversal learning deficits. The current study tests the hypothesis that proinflammatory induction mediates AIE-induced (1) loss of adult neurogenesis (i.e., doublecortin (DCX) expressing immature neurons), (2) reductions in forebrain and hippocampal cholinergic markers, and (3) reversal learning deficits., Methods: Male and female rats underwent AIE (5.0 g/kg/day ethanol or water, i.g., 2 day-on/2 day-off from postnatal day (PND) 25-54), followed by a 2-week regimen of the anti-inflammatory compound indomethacin (4.0 g/kg/day, PND 56-69) or vehicle, after which one cohort was euthanized for immunohistochemical markers (PND 70) and the second underwent the Morris water maze to assess reversal learning., Results: AIE reduced adult (PND 70) DCX+ immunoreactivity (IR) and increased hippocampal expression of the innate immune signal's high-mobility group box protein 1 (HMGB1 + IR) and cyclooxygenase-2 (COX-2 + IR) in adult male and female rats. AIE also reduced choline acetyltransferase (ChAT+IR) in the basal forebrain and co-labeling of hippocampal vesicular acetylcholine transporter (VAChT+) cholinergic terminals on DCX + IR neurons. Indomethacin treatment after AIE restored molecular endpoints to control levels and rescued AIE-induced reversal learning deficits in the Morris water maze in both sexes. Of note, indomethacin produced several adverse effects selectively in control conditions, highlighting the uniquely beneficial effect of indomethacin in AIE rats., Conclusions: These data suggest that in males and females, (1) AIE persistent neuroimmune induction mediates both the loss of adult hippocampal DCX and loss of basal forebrain cholinergic neurons and their innervation to hippocampal targets, and (2) anti-inflammatory indomethacin treatment following AIE that restores these persistent molecular pathologies also restores spatial reversal learning deficits., (© 2023 The Authors. Alcohol: Clinical and Experimental Research published by Wiley Periodicals LLC on behalf of Research Society on Alcohol.)

Nonsteroidal anti-inflammatory drugs (NSAIDs) increase risks of severe small intestinal injuries. Development of effective therapeutic strategies to overcome this issue remains challenging. Nitric oxide (NO) as a gaseous mediator plays a protective role in small intestinal injuries. However, small intestine-specific delivery systems for NO have not been reported yet. In this study, we reported a small intestine-targeted polymeric NO donor (CS-NO) which was synthesized by covalent grafting of α-glucosidase-activated NO donor onto chitosan. In vitro and in vivo experiments demonstrated that CS-NO could be activated by intestinal α-glucosidase to release NO in the small intestine. Pre-treatment of mice with CS-NO significantly alleviated small intestinal damage induced by indomethacin, as demonstrated by down-regulation of the levels of pro-inflammatory cytokines and chemokines CXCL1/KC. Moreover, CS-NO also attenuated indomethacin-induced gut barrier dysfunction as evidenced by up-regulation of the levels of tight junction proteins and restoration of the levels of goblet cells and MUC2 production. Meanwhile, CS-NO effectively restored the defense function of Paneth cells against pathogens in small intestine. Our present study paves the way to develop NO-based therapeutic strategy for NSAIDs-induced small intestinal injuries., Competing Interests: Declaration of competing interest There are no conflicts to declare., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The intracellular signaling pathways that regulate myometrial contractions can be targeted by drugs for tocolysis. The agents, 2-APB, glycyl-H-1152, and HC-067047, have been identified as inhibitors of uterine contractility and may have tocolytic potential. However, the contraction-blocking potency of these novel tocolytics was yet to be comprehensively assessed and compared to agents that have seen greater scrutiny, such as the phosphodiesterase inhibitors, aminophylline and rolipram, or the clinically used tocolytics, nifedipine and indomethacin. We determined the IC 50 concentrations (inhibit 50% of baseline contractility) for 2-APB, glycyl-H-1152, HC-067047, aminophylline, rolipram, nifedipine, and indomethacin against spontaneous ex vivo contractions in pregnant human myometrium, and then compared their tocolytic potency. Myometrial strips obtained from term, not-in-labor women, were treated with cumulative concentrations of the contraction-blocking agents. Comprehensive dose-response curves were generated. The IC 50 concentrations were 53 µM for 2-APB, 18.2 µM for glycyl-H-1152, 48 µM for HC-067047, 318.5 µM for aminophylline, 4.3 µM for rolipram, 10 nM for nifedipine, and 59.5 µM for indomethacin. A single treatment with each drug at the determined IC 50 concentration was confirmed to reduce contraction performance (AUC) by approximately 50%. Of the three novel tocolytics examined, glycyl-H-1152 was the most potent inhibitor. However, of all the drugs examined, the overall order of contraction-blocking potency in decreasing order was nifedipine > rolipram > glycyl-H-1152 > HC-067047 > 2-APB > indomethacin > aminophylline. These data provide greater insight into the contraction-blocking properties of some novel tocolytics, with glycyl-H-1152, in particular, emerging as a potential novel tocolytic for preventing preterm birth., (© 2022. The Author(s).)

Hyaluronic acid (HA), as a hygroscopic and biocompatible molecule, has displayed unique permeation enhancement in transdermal delivery systems. Hence, indomethacin (IND) was encapsulated in HA-modified transfersomes (IND-HTs) to enhance transdermal IND delivery to reduce adverse effects in this study. The physiochemical properties of IND-HTs were characterized. Results showed that the prepared IND-HTs were spherical and revealed good entrapment efficiency (87.88 ± 2.03%), with a nanometric particle size (221.8 ± 93.34 nm). Then, IND-HTs were further incorporated into a carbopol 940 hydrogel (IND-HTs/Gel) to prolong retention capacity on the skin. The in vitro release and skin permeation experiments of IND-HTs/Gel were carried out with the Franz diffusion cells. It was found that IND-HTs/Gel exhibited sustained drug release, as well as superior drug permeation and flux across the skin. Confocal laser scanning microscopy showed improved penetration of HTs/Gel with a wider distribution and higher fluorescence intensity. The hematoxylin-eosin stained showed that HA improved the transdermal effect by changing the microstructure of skin layers and decreasing skin barrier function. In addition, IND-HTs/Gel showed significant analgesic activity in hot plate test and no potentially hazardous skin irritation. This study indicated that the developed IND-HTs/Gel could be a promising alternative to conventional oral delivery of IND by topical administration.

Antitumor immune polarization is a key predictor of clinical outcomes to cancer therapy. An emerging concept influencing clinical outcome involves the spatial location of CD8 + T cells, within the tumor. Our earlier work demonstrated immunosuppressive effects of NOS2 and COX2 tumor expression. Here, we show that NOS2/COX2 levels influence both the polarization and spatial location of lymphoid cells including CD8 + T cells. Importantly, elevated tumor NOS2/COX2 correlated with exclusion of CD8 + T cells from the tumor epithelium. In contrast, tumors expressing low NOS2/COX2 had increased CD8 + T cell penetration into the tumor epithelium. Consistent with a causative relationship between these observations, pharmacological inhibition of COX2 with indomethacin dramatically reduced tumor growth of the 4T1 model of TNBC in both WT and Nos2 - mice. This regimen led to complete tumor regression in ∼20-25% of tumor-bearing Nos2 - mice, and these animals were resistant to tumor rechallenge. Th1 cytokines were elevated in the blood of treated mice and intratumoral CD4 + and CD8 + T cells were higher in mice that received indomethacin when compared to control untreated mice. Multiplex immunofluorescence imaging confirmed our phenotyping results and demonstrated that targeted Nos2/Cox2 blockade improved CD8 + T cell penetration into the 4T1 tumor core. These findings are consistent with our observations in low NOS2/COX2 expressing breast tumors proving that COX2 activity is responsible for limiting the spatial distribution of effector T cells in TNBC. Together these results suggest that clinically available NSAID's may provide a cost-effective, novel immunotherapeutic approach for treatment of aggressive tumors including triple negative breast cancer., Competing Interests: Declaration of competing interest The authors have no conflict of interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Challenges associated with topical analgesics and anti-inflammatory drugs include poor drug penetration and retention at the desired lesion site. Therefore, improving these challenges would help to reduce the toxic and side effects caused by drug absorption into the systemic circulation and improve the therapeutic efficacy of topical therapeutic drugs. Pentapeptide (KTTKS) is a signal peptide in skin tissue, it can be recognized and bound by signal recognition particles. In the current study, we successfully prepared novel indomethacin (IMC) loaded KTTKS-modified ethosomes (IMC-KTTKS-Es), and the physicochemical properties and topical efficacy were investigated. Results showed that the prepared IMC-KTTKS-Es displayed a particle size of about 244 nm, a negative charge, good deformability, and encapsulation efficiency (EE) exceeding 80% for IMC, with a sustained release pattern. In vitro percutaneous permeation studies revealed that the skin retention was increased after the drug was loaded in the IMC-KTTKS-Es. Confocal laser scanning microscopy also showed improved skin retention of IMC-KTTKS-Es. In addition, IMC-KTTKS-Es showed improved topical analgesic and anti-inflammatory activity with no potentially hazardous skin irritation. This study suggested that the IMC-KTTKS-Es might be an effective drug carrier for topical skin therapy with a good safety profile.

Src tyrosine kinases and spleen tyrosine kinase (Syk) have recently been shown to contribute to sustained platelet aggregation on collagen under arterial shear. In the present study, we have investigated whether Src and Syk are required for aggregation under minimal shear following activation of glycoprotein VI (GPVI) and have extended this to C-type lectin-like receptor-2 (CLEC-2) which signals through the same pathway. Aggregation was induced by the GPVI ligand collagen-related peptide (CRP) and the CLEC-2 ligand rhodocytin and monitored by light transmission aggregometry (LTA). Aggregation and tyrosine phosphorylation by both receptors were sustained for up to 50 min. The addition of inhibitors of Src, Syk or Bruton's tyrosine kinase (Btk) at 150 sec, by which time aggregation was maximal, induced rapid loss of tyrosine phosphorylation of their downstream proteins, but only Src kinase inhibition caused a weak (~10%) reversal in light transmission. A similar effect was observed when the inhibitors were combined with apyrase and indomethacin or glycoprotein IIb-IIIa (GPIIb-IIIa) antagonist, eptifibatide. On the other hand, activation of GPIIb-IIIa by GPVI in a diluted platelet suspension, as measured by binding of fluorescein isothiocyanate-labeled antibody specific for the activated GPIIb-IIIa (FITC-PAC1), was reversed on the addition of Src and Syk inhibitors showing that integrin activation is rapidly reversible in the absence of outside-in signals. The results demonstrate that Src but not Syk and Btk contribute to sustained aggregation as monitored by LTA, possibly as a result of inhibition of outside-in signaling from GPIIb-IIIa to the cytoskeleton through a Syk-independent pathway. This is in contrast to the role of Syk in supporting sustained aggregation on collagen under arterial shear.

Mitochondrion emerged as an important therapeutic target for anti-cancer strategy due to its involvement in cancer progression and development. However, progress of novel small molecules for selective targeting of mitochondria in cancer cells remained a major challenge. To address this, herein, through a concise synthetic strategy, we have synthesized a small molecule library of indomethacin and ibuprofen (non-steroidal anti-inflammatory drugs, NSAIDs) derivatives having triarylphosphonium moiety for mitochondria localization. Two of the library members were identified to induce mitochondrial damage through outer membrane permeabilization (MOMP) followed by generation of reactive oxygen species (ROS) leading to the remarkable MCF7 breast cancer cell death through apoptosis. These novel mitochondria targeted NSAID derivatives could open a new direction in understanding mitochondrial biology towards anti-cancer therapeutics in future., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Vanadium toxicity is a globally recognized threat to the reproductive health of man and animal. However the mechanism of vanadium-induced damage to the testicular and adrenocortical tissues is not fully characterized. It was hypothesized that prostaglandins may partially mediate the inflammatory response to vanadate damage. In this study prostaglandin (PG) mediated effects of vanadate on testicular and adrenocortical functions was substantiated by using indomethacin to block prostaglandin synthesis. Significant inhibition of spermatogenesis, decreased serum level of testosterone and gonadotropins in the vanadium-exposed group of rats indicated the damaging effects of vanadium-induced reactive oxygen species. This was also reflected in the appreciable increase in testicular lipid peroxidation (LPO) level and decline in the activities of steroidogenic and antioxidant enzymes. Histopathological studies revealed regressive and degenerative changes in testis. However, inhibition of cyclooxygenase activity by indomethacin increased steroid hormone production, gonadotropin level, elevated the specific activities of enzymes and decreased LPO level in rat testis exposed to vanadium. Vanadium also caused prostaglandin mediated adrenocortical hyperactivity, as inhibition of PG synthesis abolished these adrenal responses to vanadium. The studies showed that vanadium toxicity is directly linked to stimulation of prostaglandin synthesis. Therefore, indomethacin can be a good prospect to alleviate vanadium induced male infertility.

Carboxylesterase (CES) plays an important role in the hydrolysis metabolism of ester-type drugs and prodrugs. In this study, we investigated the change in the hydrolysis rate of hCE1 by focusing on the steric hindrance of the ester structure and the electron density. For 26 kinds of synthesized indomethacin prodrugs, the hydrolytic rate was measured in the presence of human liver microsomes (HLM), human small intestine microsomes (HIM), hCE1 and hCE2. The synthesized prodrugs were classified into three types: an alkyl ester type that is specifically metabolized by hCE1, a phenyl ester type that is more easily metabolized by hCE1 than by hCE2, and a carbonate ester type that is easily metabolized by both hCE1 and hCE2. The hydrolytic rate of 1-methylpentyl (hexan-2-yl) ester was 10-times lower than that of 4-methylpentyl ester in hCE1 solution. hCE2 was susceptible to electron density of the substrate, and there was a difference in the hydrolysis rate of up to 3.5-times between p-bromophenyl ester and p-acetylphenyl ester. By changing the steric hindrance and electron density of the alkoxy group, the factors that change the hydrolysis rate by CES were elucidated., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2021 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.)

A new double-layer, pH-sensitive, composite hydrogel sustained-release system based on polysaccharides and synthetic polymers with combined functions of different inner/outer hydrogels was prepared. The polysaccharides inner core based on sodium alginate (SA) and carboxymethyl cellulose (CMC), was formed by physical crosslinking with pH-sensitive property. The synthetic polymer out-layer with enhanced stability was introduced by chemical crosslinking to eliminate the expansion of inner core and the diffusion of inner content. The physicochemical structure of the double-layer hydrogels was characterized. The drug-release results demonstrated that the sustained-release effect of the hydrogels for different model drugs could be regulated by changing the composition or thickness of the hydrogel layer. The significant sustained-release effect for BSA and indomethacin indicated that the bilayer hydrogel can be developed into a novel sustained delivery system for bioactive substance or drugs with potential applications in drugs and functional foods.

Peptic ulcer is a major health challenge with high morbidity and mortality all over the world. This study investigated the involvement of oxidative stress in the healing and protective potentials of aqueous leave extract of Telfairia occidentalis (TO) on indomethacin induced gastric ulcers in adult Sprague Dawley male rats. The rats were divided into 6 groups (A-F) of 5 rats each, with A as normal control, B received single oral administration of 40mg/kg indomethacin without treatment for 4 hours; C received 40mg/kg indomethacin without treatment for 4 hours and scarified after 72 hours; D received 100mg/kg aqueous leave extract of TO for 7 days without ulcer induction; E (pre-treated test group) received 40mg/kg indomethacin after being pre-treated with 100mg/kg aqueous leave extract of TO daily for 7 days. Group F (Posttreated test) received 40mg/kg of indomethacin and treated four hours later with 100mg/kg aqueous leave extract of TO daily for 7 days. The results revealed changes in gastric macroscopic architecture of the mucosa, and changes in ulcer indices and oxidative stress markers levels in group B-F. These changes comparatively suggested that the leave-extract of Telfairia occidentalis has gastro-protective with minimal healing potentials mediated through reduced oxidative stress.

The contribution of metabotropic glutamate receptors (mGlu receptors) in depression is well known and tested worldwide. Our previous study showed the involvement of the cyclooxygenase-2 (COX-2) pathway in behavioral changes mediated by an antagonist of metabotropic glutamate receptor subtype 5 (mGlu5 receptor) 3-[(2-methyl-1,3-tiazol-4-yl)ethynyl]-pyridine (MTEP). Among others, we have found that chronic concomitant administration of a COX-2 inhibitor and sub-effective dose of MTEP accelerates antidepressant-like activity of MTEP. This paper seeks to explore whether the same effect would be observed with the use of a non-selective COX inhibitor 2-[1-(4-chlorobenzoyl)-5-methoxy-2-methylindol-3-yl]acetic acid (indomethacin). To that end, we have employed experimental procedure implemented in the earlier research. MTEP and indomethacin or MTEP + indomethacin were used chronically for 7 or 14 days. Then, the Porsolt test, tail suspension test and locomotor activity test were performed. Imipramine was used as a reference compound, as its action is connected with mGlu5 receptor. We found that, in contrast to COX-2 inhibition, indomethacin - acting both through COX-1 and COX-2 - did not release antidepressant-like potential of MTEP. The opposite effect was shown when imipramine was used., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Nanoparticles (NPs) produced from amphiphilic derivatives of poly- N -vinylpyrrolidone (Amph-PVP), composed of various molecular weight polymeric hydrophilic fragments linked into hydrophobic n -alkyl chains of varying lengths, were previously shown to exert excellent biocompatibility. Although routes of administration can be different, finally, most nanosystems enter the blood circulation or lymphatic vessels, and by this, they establish direct contact with endothelial cells. In this study, Amph-PVP NPs and fluorescently labeled Amph-PVP-based NPs, namely "PVP" NPs (Amph-PVP-NPs (6000 Da) unloaded) and "F"-NPs (Amph-PVP-NPs (6000 Da) loaded with fluorescent FITC), were synthesized to study Amph-PVP NPs interactions with HMEC-1 endothelial cells. PVP NPs were readily uptaken by HMEC-1 cells in a concentration-dependent manner, as demonstrated by immunofluorescence imaging. Upon uptake, the FITC dye was localized to the perinuclear region and cytoplasm of treated cells. The generation of lipopolysaccharide (LPS)-induced activated endothelium model revealed an increased uptake of PVPNPs, as shown by confocal microscopy. Both unloaded PVP NPs and F-NPs did not affect EC viability in the 0.01 to 0.066 mg/mL range. Furthermore, we focused on the potential immunological activation of HMEC-1 endothelial cells upon PVPNPs treatment by assessing the expression of their E-Selectin, ICAM-1, and VCAM-1 adhesion receptors. None of the adhesion molecules were affected by NP treatments of both activated by LPS and nonactivated HMEC-1 cells, at the utilized concentrations (p = NS). In this study, PVP (6000 Da) NPs were used to encapsulate indomethacin, a widely used anti-inflammatory drug. The synthesized drug carrier complex did not affect HMEC-1 cell growth and expression of E-selectin, ICAM-1, and VCAM-1 adhesion receptors. In summary, PVP-based NPs are safe for use on both basal and activated endothelium, which more accurately mimics pathological conditions. Amph-PVP NPs are a promising drug delivery system.

Incomplete drug release from mesoporous silica systems has been observed in several studies. This work aims to increase the understanding of this phenomenon by investigating the mechanism of drug-silica interactions and adsorption behavior from supersaturated aqueous solutions of two similar drug molecules with different hydrogen bonding capabilities. Drug-silica interactions between indomethacin or its methyl ester and SBA-15 were investigated using spectroscopic techniques (infrared, fluorescence and X-ray photoelectron) and adsorption experiments. The results demonstrate that the predominant mechanism of interaction of both drugs with silica is hydrogen bonding between drug acceptor carbonyl groups with donor groups on the silica surface. The presence of a drug hydrogen bond donor group did not enhance drug adsorption. No evidence was obtained for drug adsorption through nonspecific hydrophobic interactions. Drug adsorption onto the silica surface was investigated under supersaturating conditions through the generation of adsorption isotherms. Similar adsorption isotherms were observed for each compound when the concentration scale was normalized to the drug amorphous solubility. In other words, the equilibrium between the drug adsorbed on the silica surface and free drug in solution was related to the drug activity in solution. The high tendency of the drug to adsorb when the solution is supersaturated was, in turn, found to limit the extent of drug release during dissolution under nonsink conditions. Thus, adsorption provides an explanation for incomplete drug release.

Osthol, a pharmacologically active ingredient in various traditional Chinese medicines, is predominantly metabolized by CYP2C9. It may be co-administered with other drugs which are metabolized by CYP2C9 in clinical medicine. However, CYP2C9*1/*2/*3 genotype on the pharmacokinetics of osthole and its metabolic diversity between rat and human are unclear.In this study, we investigated the effects of osthole on enzyme activity of CYP2C11/CYP2C9 in rat liver microsomes (RLMs) and human liver microsomes (HLMs), to distinguish metabolic manner of osthole in different species. Interestingly, we found that osthole inhibits the activity of CYP2C11 in a non-competitive manner in RLMs, while inhibits CYP2C9 activity in a competitive manner in pooled HLMs. Then, the effects of CYP2C9*1/*2/*3 allele on the pharmacokinetics of osthole were identified. In human CYP2C9 isoform, the Ki value of 21.93 μM (CYP2C9*1), 18.10 μM (CYP2C9*2), 13.12 μM (CYP2C9*3) indicate that there are individual differences in the inhibition of osthole on CYP2C9 activity.We investigated how the indomethacin pharmacokinetics was affected by osthole in SD rat. To estimate the area under the curve (AUC), maximum plasma concentration ( C max ) and apparent clearance (CL/F), indomethacin (10 mg/kg) was given orally combined with osthole (20 mg/kg) in adult SD rat. We found the value of PK on indomethacin, such as the AUC 0-∞ , was from 176.40 ± 17.29 to 173.74 ± 27.69 μg/ml h -1 , C max from 9.02 ± 1.24 to 9.89 ± 0.82 μg/ml and CL/F from 0.11 ± 0.01 to 0.12 ± 0.04 mg/kg/h which were unsignificantly changed compared with the control groups. However, the T max was prolonged from 2.00 ± 0.00 h to 7.33 ± 1.15 h, and T 1/2 increased from 8.38 ± 2.30 h to 11.37 ± 2.11 h. These results indicate that osthole could potentially affect the metabolism of indomethacin in vivo .

The effect of the degree of supersaturation (DS) on absorption of the model drugs indomethacin and tadalafil was elucidated in a single-pass intestinal perfusion (SPIP) model in rats. In addition, the performance of the precipitation inhibitor (PI) hydroxypropylmethylcellulose (HPMC) was evaluated when added at a concentration of 0.1% (w/v) to fasted state simulated intestinal fluid (FaSSIF and FaSSIF HPMC ) used as perfusion medium. A supersaturated state was created by a solvent shift method where indomethacin or tadalafil dissolved in dimethyl sulfoxide (DMSO) were administered to a segment of the small intestine, which subsequently was perfused with FaSSIF or FaSSIF HPMC . The perfusate was collected for 60 min, and for one group of rats dosed with 30 mg tadalafil, for 120 min. Blood samples were drawn every 15 min. The solubility of indomethacin and tadalafil in the perfusate was determined. The DS of each drug in the perfusate was calculated by dividing the concentration in the perfusate at selected time points with the solubility. The DS was above one for all timepoints for both drugs, thus showing supersaturation during the time of perfusion. For indomethacin, no improvement of the DS was seen when perfusing with FaSSIF HPMC , compared to FaSSIF. For tadalafil, a higher DS was achieved when perfusing with FaSSIF HPMC compared to FaSSIF. Perfusing the drugs with FaSSIF HPMC resulted in a significantly lower area under the curve (AUC 0-60 min ) for plasma concentrations of indomethacin, and no increase in the AUC 0-60 min of plasma concentrations of tadalafil compared to perfusion with FaSSIF. The importance of simultaneously estimating the intraluminal DS and absorption of a drug was demonstrated by the SPIP model in the present study. Further, the study highlights the discrepancy between optimal in vitro supersaturation, intraluminal supersaturation and in vivo performance of two poorly soluble drugs, and further emphasizes the importance of optimization of in vitro methods in order to predict in vivo supersaturation and precipitation of drugs., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

(1) Background: Compounds with multitarget activity are of interest in basic research to explore molecular foundations of promiscuous binding and in drug discovery as agents eliciting polypharmacological effects. Our study has aimed to systematically identify compounds that form complexes with proteins from distinct classes and compare their bioactive conformations and molecular properties. (2) Methods: A large-scale computational investigation was carried out that combined the analysis of complex X-ray structures, ligand binding modes, compound activity data, and various molecular properties. (3) Results: A total of 515 ligands with multitarget activity were identified that included 70 organic compounds binding to proteins from different classes. These multiclass ligands (MCLs) were often flexible and surprisingly hydrophilic. Moreover, they displayed a wide spectrum of binding modes. In different target structure environments, binding shapes of MCLs were often similar, but also distinct. (4) Conclusions: Combined structural and activity data analysis identified compounds with activity against proteins with distinct structures and functions. MCLs were found to have greatly varying shape similarity when binding to different protein classes. Hence, there were no apparent canonical binding shapes indicating multitarget activity. Rather, conformational versatility characterized MCL binding., Competing Interests: The authors declare no conflict of interest.

Human carboxylesterase 1 (hCES1) is an enzyme that plays an important role in hydrolysis of pharmaceuticals in the human liver. In this study, elucidation of the chiral recognition ability of hCES1 was attempted using indomethacin esters in which various chiral alcohols were introduced. Indomethacin was condensed with various chiral alcohols to synthesize indomethacin esters. The synthesized esters were hydrolyzed with a human liver microsome (HLM) solution and a human intestine microsome (HIM) solution. High hydrolytic rate and high stereoselectivity were confirmed in the hydrolysis reaction in the HLM solution but not in the HIM solution, and these indomethacin esters were thought to be hydrolyzed by hCES1. Next, these indomethacin esters were hydrolyzed in recombinant hCES1 solution and the hydrolysis rates of the esters were calculated. The stereoselectivity confirmed in HLM solution was also confirmed in the hCES1 solution. In the hydrolysis reaction of esters in which a phenyl group is bonded next to the ester, the V max value of the (R) form was 10 times larger than that of the (S) form., (© 2019 Wiley Periodicals, Inc.)

The laccase (Lac), manganese peroxidases (MnP), and lignin peroxidase enzymes produced by basidiomycete have been studied due to their potential in bioremediation, therefore, in this study, degradation of diclofenac (DCF), sulfamethoxazole (SMX), indomethacin (IND), gemfibrozil (GFB), and bezafibrate (BZF) by enzymes produced by Trametes maxima , Pleurotus sp., and Pycnosporus sanguineus grown in culture was evaluated. The degradation of drugs can mainly be attributed to MnP because a correlation between the activity of this enzyme and the degree of removal was found. The specific activity of Lac did not show correlation with drug removal, while lignin peroxidase was not expressed. Trametes maxima showed the highest specific activity of MnP (387.6 ± 67.4 U/mg) and efficiency removal 90.2% of DCF, 72.62% of SMX, 60.76% of IND, 43.39% of GFB, and 32.59% of BZF) followed by Pleurotus sp . with specific activity of MnP of 55.9 ± 8.5 U/mg and 89.47% of DCF, 47.61% of GFB and 73% of IND were removed, P. sanguineus had the lowest specific activity of 18 ± 1.3 U/mg and was able to remove only 42% of SMX and 10.59% of IND. In order to prove that MnP remove drugs instead of Lac, the pure Lac was tested and only degraded DCF.

The present work attempts to develop and optimize the formula of a lipidic nanoemulsion (NE) containing sodium hyaluronate (HNa) and indomethacin (Ind) as HNa-Ind for enhanced transdermal antiarthritic activity. NEs were prepared by the spontaneous emulsification method and characterized by Fourier-transform infrared (FTIR) spectroscopy. The composition of the optimal formulation was statistically optimized using Box-Behnken experimental design method with three independent factors and was characterized for particle size, polydispersity index, and percent transmittance. The selected formula was tested for its in vitro antioxidant activity and in vivo anti-inflammatory activity. The optimized HNa-Ind NE formula was characterized and displayed a particle size of 12.87 ± 0.032 nm, polydispersity index of 0.606 ± 0.082, and 99.4 ± 0.1 percentage of transmittance. FTIR showed no interaction between HNa and Ind as a physical mixture. In addition, the optimized HNa-Ind NE was able to preserve the antioxidant ability of the two drugs, as evidenced through a 2,2-diphenyl-1-picrylhydrazyl (DPPH) inhibition assay used to assess free radical scavenging ability. The cell viability was increased while the free radical scavenging activity was decreased (94.28% inhibition at higher concentrations compared with vitamin C as a reference with an inhibition of 100%). Moreover, the pharmacological anti-inflammatory potential of the optimized HNa-Ind NE formulation was assessed using an in vivo model. Compared with reference drugs (ibuprofen gel 5%), the remarkable activity of the optimized formulation was established using xylene-induced ear edema in mice model, in which the inflamed region reduced by 92.5% upon treatment. The optimized HNa-Ind NE formulation showed considerably higher skin permeation and drug deposition capability compared with the HNa-Ind solution. HNa-Ind NE was demonstrated to be a successful carrier with enhanced antioxidant and anti-inflammatory potential while showing better skin penetration, thus being a promising vehicle for transdermal drug delivery.

Novel microparticles coated with poly-γ-glutamic acid (PGA) were developed to improve the oral absorption of indomethacin (IM), a poorly water-soluble drug. Microparticles containing γ-IM (IM bulk -PGA) or crystal polymorph α-IM (IM polymorph -PGA) were prepared. Additionally, microparticles were prepared containing α-IM without PGA (IM polymorph without PGA). IM bulk -PGA and IM polymorph -PGA exhibited better drug retention properties on mucin disks. Drug release rates from IM polymorph -PGA and IM polymorph without PGA were higher than from IM bulk powder, and drug release from IM bulk -PGA was also improved. Drug release from IM bulk -PGA could be improved with the use of Tween 80. In addition, PGA may influence the ionization of IM or affect specific molecular interactions. After the microparticles were administered orally to mice, IM bulk -PGA and IM polymorph -PGA increased the plasma drug concentration more rapidly compared with IM bulk powder, but IM polymorph without PGA did not increase the plasma drug concentration. It was considered that IM bulk -PGA and IM polymorph -PGA rapidly reached the intestinal membrane through the mucus layer and IM was absorbed quickly. Because IM bulk -PGA and IM polymorph -PGA showed a rapid increase in plasma drug concentration, IM bulk -PGA and IM polymorph -PGA could be useful preparations to improve the gastrointestinal absorption of IM. Furthermore, IM bulk -PGA may maintain higher plasma drug concentrations than IM polymorph -PGA.

LC-MS quantification of drug metabolites is sometimes impeded by the availability of internal standards that often requires customized synthesis and/or extensive purification. Although isotopically labeled internal standards are considered ideal for LC-MS/MS based quantification, de novo synthesis using costly isotope-enriched starting materials makes it impractical for early stage of drug discovery. Therefore, quick access to these isotope-enriched compounds without chemical derivatization and purification will greatly facilitate LC-MS/MS based quantification. Herein, we report a novel 18 O-labeling technique using metabolizing enzyme carboxylesterase (CES) and its potential application in metabolites quantification study. Substrates of CES typically undergo a two-step oxygen exchange with H 2 18 O in the presence of the enzyme, generating singly- and doubly- 18 O-labeled carboxylic acids; however, unexpected hydrolytic behavior was observed for three of the test compounds - indomethacin, piperacillin and clopidogrel. These unusual observations led to the discovery of several novel hydrolytic mechanisms. Finally, when used as internal standard for LC-MS/MS based quantification, these in situ labeled compounds generated accurate quantitation comparable to the conventional standard curve method. The preliminary results suggest that this method has potential to eliminate laborious chemical synthesis of isotope-labeled internal standards for carboxylic acid-containing compounds, and can be developed to facilitate quantitative analysis in early-stage drug discovery., (Copyright © 2019 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.)

Receptor tyrosine kinase (RTK)-mediated hyperactivation of the MAPK/Erk pathway is responsible for a large number of pathogenic outcomes including many cancers. Considerable effort has been directed at targeting this pathway with varying degrees of long term therapeutic success. Under non-stimulated conditions Erk is bound to the adaptor protein Shc preventing aberrant signalling by sequestering Erk from activation by Mek. Activated RTK recruits Shc, via its phosphotyrosine binding (PTB) domain (Shc PTB ), precipitating the release of Erk to engage in a signalling response. Here we describe a novel approach to inhibition of MAP kinase signal transduction through attempting to preserve the Shc-Erk complex under conditions of activated receptor. A library of existing drug molecules was computationally screened for hits that would bind to the Shc PTB and block its interaction with the RTKs EGFR and ErbB2. The primary hit from the screen was indomethacin, a non-steroidal anti-inflammatory drug. Validation of this molecule in vitro and in cellular efficacy studies in cancer cells provides proof of principle of the approach to pathway down-regulation and a potential optimizable lead compound., (Copyright © 2019 The Author(s). Published by Elsevier B.V. All rights reserved.)

The in vitro dissolution absorption system 2 (IDAS2), a recent invention comprised a conventional dissolution vessel containing 2 permeation chambers with Caco-2 cell monolayers mounted with their apical side facing the dissolution media, permits simultaneous measurement of dissolution and permeation of drugs from intact clinical dosage forms. The objectives of this study were (1) to assess the utility of IDAS2 in the determination of the effect of particle size on in vitro performance of indomethacin and (2) to find out whether the behavior in IDAS2 of 2 indomethacin products differing in particle size is correlated with their in vivo behavior. Indomethacin dissolution and permeation across Caco-2 cell monolayers were simultaneously measured in IDAS2; the dissolution and permeation profiles were simultaneously modeled using a simple two-compartment model. Compared to microsized indomethacin, the nanosized formulation increased the dissolution rate constant by fivefold, whereas moderately increasing the permeation rate constant and the kinetic solubility. As a result, the drug amount permeated across the Caco-2 cell monolayers doubled in the nanosized versus microsized formulation. The in vitro results showed a good correlation with in vivo human oral pharmacokinetic parameters, thus emphasizing the physiological relevance of IDAS2 data in predicting in vivo absorption., (Copyright © 2019 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)

We have synthesized a fluorinated analogue of indomethacin bearing a 3,3,3-trifluoroprop-1-enyl group at its 2-position and evaluated its inhibitory activity towards the COX-1 and COX-2 enzymes in vitro. The results revealed that this fluorinated analogue exhibited much greater inhibitory activity and selectivity towards COX-2 than indomethacin. The increased affinity between the fluorinated analogue and COX-2 was attributed to a significant increase in van der Waals contacts (i.e. van der Waals contributions in ΔG were -13.80 kcal/mol for COX-1 and -18.46 kcal/mol for COX-2), explaining an effect of the fluorine substituent in enzyme selectivity. This newly synthesized fluorinated analogue therefore represents a potent and selective COX-2 inhibitor., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Solubility is a physicochemical property highly dependent on the solid-state form of a compound. Thus, alteration of a compound's solid-state form can be undertaken to enhance the solubility of poorly soluble drug compounds. In the Biopharmaceutics Classification System (BCS), drugs are classified on the basis of their aqueous solubility and permeability. However, aqueous solubility does not always correlate best with in vivo solubility and consequently bioavailability. Therefore, the use of biorelevant media is a more suitable approach for mimicking in vivo conditions. Here, assessed with a novel image-based single-particle-analysis (SPA) method, we report a constant ratio of solubility increase of 3.3 ± 0.5 between the α and γ solid-state forms of indomethacin in biorelevant media. The ratio was independent of pH, ionic strength, and surfactant concentration, which all change as the drug passes through the gastrointestinal tract. On the basis of the solubility ratio, a free-energy difference between the two polymorphic forms of 2.9 kJ/mol was estimated. Lastly, the use of the SPA approach to assess solubility has proven to be simple, fast, and both solvent- and sample-sparing, making it an attractive tool for drug development.

We previously reported that oral formulations containing indomethacin nanoparticles (IND-NPs) showed high bioavailability, and, consequently, improved therapeutic effects and reduced injury to the small intestine. However, the pathway for the transintestinal penetration of nanoparticles remained unclear. Thus, in this study, we investigated whether endocytosis was related to the penetration of IND-NPs (72.1 nm) using a transcell set with Caco-2 cells or rat intestine. Four inhibitors of various endocytosis pathways were used [nystatin, caveolae-dependent endocytosis (CavME); dynasore, clathrin-dependent endocytosis (CME); rottlerin, macropinocytosis; and cytochalasin D, phagocytosis inhibitor], and all energy-dependent endocytosis was inhibited at temperatures under 4 °C in this study. Although IND-NPs showed high transintestinal penetration, no particles were detected in the basolateral side. IND-NPs penetration was strongly prevented at temperatures under 4 °C. In experiments using pharmacological inhibitors, only CME inhibited penetration in the jejunum, while in the ileum, both CavME and CME significantly attenuated penetration. In conclusion, we found a novel pathway for the transintestinal penetration of drug nanoparticles. Our hypothesis was that nanoparticles would be taken up into the intestinal epithelium by endocytosis (CME in jejunum, CavME and CME in ileum), and dissolved and diffused in the intestine. Our findings are likely to be of significant use for the development of nanomedicines., Competing Interests: The authors declare no conflict of interest.

Aim: Differential pulse polarography was used for the concurrent analysis of the coadministered dantrolene (DAN) and indomethacin (IND) in plasma., Materials & Methods: DAN and IND, Hanging mercury drop electrode and Britton-Robinson buffer at pH 5 were used. In plasma, cathodic reduction of DAN nitro group and its active metabolite at -0.2 V was done. IND was analyzed after carbonyl group reduction at -1.1 V., Results: Drugs determination in rat plasma with good recoveries and low limit of quantitation was done. Application to trace analysis of drugs in rat plasma was done with C max and T max determination., Conclusion: This technique shows high sensitivity, simplicity and low cost. The method is US FDA validated and it is applicable to human level.

Inhibitors selective towards the second isoform of prostaglandin synthase (cyclooxygenase, COX-2) are promising nonsteroidal anti-inflammatory drugs and antitumor medications. Methylation of the carboxylate group in the relatively nonselective COX inhibitor indomethacin confers significant COX-2 selectivity. Several other modifications converting indomethacin into a COX-2 selective inhibitor have been reported. Earlier experimental and computational studies on neutral indomethacin derivatives suggest that the methyl ester derivative likely binds to COX-2 with a similar binding mode as that observed for the parent indomethacin. However, docking studies followed by molecular dynamics simulations revealed two possible binding modes in COX-2 for indomethacin methyl ester, which differs from the experimental binding mode found for indomethacin. Both alternative binding modes might explain the observed COX-2 selectivity of indomethacin methyl ester. Graphical abstract Binding of indomethacin methyl ester to cyclooxygenase-2.

Phospholipase A 2 (PLA 2 ) catalyzes the hydrolysis of phospholipids into arachidonic acid and lysophospholipids. Arachidonic acid is modified by cyclooxygenases into active compounds called eicosanoids that act as signaling molecules in a number of physiological processes. Excessive production of eicosanoids leads to several pathological conditions such as inflammation. In order to block the inflammatory effect of these compounds, upstream enzymes such as PLA 2 are valid targets. In the present contribution, molecular dynamic analysis was performed to evaluate the binding of diclofenac, 9-hydroxy aristolochic acid (9-HAA) and indomethacin to PLA 2 . Obtained results revealed that 9-HAA could form a more stable complex with PLA 2 when compared to diclofenac and indomethacin. Furthermore, analysis of intermolecular binding energy components indicated that hydrophobic interactions were dominant in binding process. On the basis of obtained data, inhibitors bearing fused rings with hydrogen acceptor/donor substituent(s) interacted with His48 and Asp49 residues of the active site. More affinity toward PLA 2 might be envisaged through negatively charged moieties via interaction with Trp31, Lys34 and Lys69.

Purpose: To study and elucidate the effect of the intensity and duration of processing stresses on the possible solid-state changes during a hot melt extrusion granulation process., Methods: Blends of α-indomethacin and PEG 3350 (w/w 4:1) were granulated using various screw sizes/designs on the melt extruder under different temperature regimes. Differential Scanning Calorimetry and X-ray Powder Diffraction were employed for characterization. The dissolution behavior of the pure polymorphs and the resulting granules was determined using in-situ fiber optic UV testing system. An XRPD quantitation method using Excel full pattern fitting was developed to determine the concentration of each constituent (amorphous, α and γ indomethacin and PEG) in samples collected from each functioning zone and in granules., Results: Analysis of in-process samples and granules revealed that higher temperature (≥130°C) and shear stress accelerated the process induced phase transitions from amorphous and/or the α form to γ indomethacin during heating stage. However, rapid cooling resulted in an increased percentage of the α form allowing isolation of the meta-stable form., Conclusions: By determining the conditions that either prevent or facilitate process induced transformations of IMC polymorphs during melt granulation, a design space was developed to control the polymorph present in the resulting granules. This represents the conditions necessary to balance the thermodynamic relationships between the polymorphs of the IMC system and the kinetics of the possible transformations as a function of the processing stresses.

It is necessary to consider the affinity of prodrugs for metabolic enzymes for efficient activation of the prodrugs in the body. Although many prodrugs have been synthesized with consideration of these chemical properties, there has been little study on the design of a structure with consideration of biological properties such as substrate recognition ability of metabolic enzymes. In this report, chemical synthesis and evaluation of indomethacin prodrugs metabolically activated by human carboxylesterase 1 (hCES1) are described. The synthesized prodrugs were subjected to hydrolysis reactions in solutions of human liver microsomes (HLM), human intestine microsomes (HIM) and hCES1, and the hydrolytic parameters were investigated to evaluate the hydrolytic rates of these prodrugs and to elucidate the substrate recognition ability of hCES1. It was found that the hydrolytic rates greatly change depending on the steric hindrance and stereochemistry of the ester in HLM, HIM and hCES1 solutions. Furthermore, in a hydrolysis reaction catalyzed by hCES1, the V max value of n-butyl thioester with chemically high reactivity was significantly lower than that of n-butyl ester., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The objective of this study was to explore the feasibility of using alginate as a promising diphase solid dispersion carrier to enhance dissolution rate of BCS II drugs with improved stability. Taking lovastatin and indomethacin as model drugs, solvent evaporation method was used to prepare solid dispersions. The drug/polymer compatibility was predicted by Hansen solubility parameter and the drug/polymer ratio was screened based on dissolution study, drug existing state in solid dispersion was characterized by DSC and XRPD. Accelerated stability of the solid dispersion was assessed and compared with that of HPMCAS based system. Phase behavior of the solid dispersion before and after stability study was characterized using polar microscope and Raman mapping. It was found that the optimal drug/alginate ratio was drug dependent and drug existing state was related to drug/alginate miscibility. Stability studies revealed that alginate improved the stability of solid dispersions regardless of drug existing state and a better stability was obtained compared to HPMCAS based system. Raman mapping and SEM study revealed that micro phase separation of solid dispersion was the main reason for the slight decrease in drug dissolution after accelerating experiment. In conclusion, alginate can be used as a promising diphase solid dispersion carrier with significantly improved dissolution rate and storage stability., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Human serum albumin (HSA) has two major ligand-binding sites, sites I and II, and hydrolyzes compounds at both sites. Although the hydrolytic interaction of ester-type drugs with other drugs by HSA has been reported, there are only a few studies concerning the effect of pharmaceutical excipients on the hydrolysis of ester-type drugs by HSA. In the present study, we investigated the effect of ethanol (2 vol%; 345 mM) on the hydrolysis of aspirin, p-nitrophenyl acetate, and olmesartan medoxomil, which are ester-type drugs, with 4 different lots of HSA preparations. The hydrolysis activities of HSA toward aspirin, p-nitrophenyl acetate, and olmesartan medoxomil were measured from the pseudo-first-order degradation rate constant (k obs ) of salicylic acid, p-nitrophenol, and olmesartan, respectively, which are the HSA-hydrolyzed products. Ethanol inhibited hydrolysis of aspirin by HSA containing low levels of fatty acids, but not by fatty acid-free HSA. Ethanol inhibited hydrolysis of p-nitrophenyl acetate by both fatty acid-free HSA and HSA containing low levels of fatty acids. In contrast, the hydrolysis of olmesartan medoxomil by HSA was insignificantly inhibited by ethanol, but inhibited not only by warfarin and indomethacin but also by naproxen, which are site I binding drugs and a site II binding drug, respectively. These results suggest that the inhibitory action of ethanol on the hydrolysis of ester-type drugs by HSA differs between site I binding drugs and site II binding drugs.

The aim of this study was to investigate the influence of factors such as carrier type, drug/carrier ratio, binary carriers, and preparation method on the dissolution of an insoluble drug, indomethacin (IM), under supersaturation conditions. Using a solvent evaporation (SE) method, poloxamer 188 and PVP K30 showed better dissolution among the selected carriers. Furthermore, as the ratio of carriers increased (drug/carrier ratio from 1:0.5 to 1:2), the dissolution rate increased especially in almost two times poloxamer 188 solid dispersions (SDs), while the reverse results were observed for PVP K30 SDs. For the binary carrier SD, a lower dissolution was found. Under hot melt extrusion (HME), the dissolution of poloxamer 188 SD and PVP K30 SD was 0.83- and 0.94-folds lower than that using SE, respectively, while the binary carrier SD showed the best dissolution. For poloxamer 188 SDs, the drug's crystal form changed when using SE, while no crystal form change was observed using HME. IM was amorphous in PVP K30 SDs prepared by both methods. For binary carrier systems, amorphous and crystalline drugs coexisted in SD using SE, and negligible amorphous IM was in SD using HME. This study indicated that a higher amorphous proportion in SD did not correlate with higher dissolution rate, and other factors, such as carrier type, particle size, and density, were also critical.

Amorphization of active pharmaceutical ingredients (APIs) and the preparation of solid dispersions are strategies that can be synergized to improve the solubility of oral drugs. Immiscibility between an API and a carrier in the molten state that could be perceived as a problem in the preparation of solid dispersions, may actually introduce an advantage. In the present work, a two-phase amorphous-amorphous solid dispersion (AASD) was prepared by ultrasonicating a molten immiscible mixture of indomethacin (IND) and glucose (GLU) prior quenching. By introducing this novel ultrasound assisted method, the immiscible API particles were uniformly dispersed as microscopic glassy clusters of the drug in the solid amorphous GLU matrix; particle sizes of IND in the AASD range from 600nm to 1.4μm. As a result of the amorphization and particle size reduction of IND, its aqueous solubility increased to reach almost 40ppm (8 times more soluble compared to indomethacin in its crystalline state). In addition, the oral bioavailability and its resistance against crystallization were also enhanced; AASD samples have remained amorphous for more than two years of storage., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Noncovalent binding of biopharmaceuticals to human serum albumin protects against enzymatic degradation and renal clearance. Herein, we investigated the effect of mono- or divalent small-molecule albumin binders for half-life extension of peptides. For proof-of-principle, the clinically relevant glucagon-like peptide 1 (GLP-1) was functionalized with diflunisal, indomethacin, or both. In vitro, all GLP-1 analogues had subnanomolar GLP-1 receptor potency. Surface plasmon resonance revealed that both small molecules were able to confer albumin affinity to GLP-1 and indicated that affinity is increased for divalent analogues. In lean mice, the divalent GLP-1 analogues were superior to monovalent analogues with respect to control of glucose homeostasis and suppression of food intake. Importantly, divalent GLP-1 analogues showed efficacy comparable to liraglutide, an antidiabetic GLP-1 analogue that carries a long-chain fatty acid. Finally, pharmacokinetic investigations of a divalent GLP-1 analogue demonstrated a promising gain in circulatory half-life and absorption time compared to its monovalent equivalent.

Molecular modeling studies were performed in order to gain insight into the binding mode and interaction of carborane-containing derivatives of indomethacin methyl ester with the cyclooxygenase-2 (COX-2) isoform, and to assess the predictive capability of the computational tools available for studying carboranes, a unique class of pharmacophores. Docking simulations were able to identify the correct binding mode and reproduced the experimental binding affinity trends with encouraging quality. Nevertheless, the docking results needed to be verified through extensive and resource-intensive quantum chemical calculations, and the interpretation of the theoretical results would not have been straightforward without the supporting experimental data. The inclusion of full receptor and ligand flexibility into the molecular modeling of carborane-containing drug molecules may yield more definitive results, but is currently hindered by the lack of appropriate carborane force field parameters.

Dual stimuli-responsive nanoparticles capable of fine-tuning drug release to augment therapeutic efficacy have become a promising tool for anticancer drug delivery. However, the rational design of these "smart" nanoparticles for a selective delivery and controlled release of multidrug combinations in cancer cells to achieve synergistic effects remain challenging. Here we report the pH/redox dual responsive nanoparticle FA-DOX-Ind-NP (FA=folic acid, DOX=doxorubicin, Ind=indomethacin, NP=nanoparticle) based on the novel tumor targeting and biodegradable poly(β-amino ester) polymer, and demonstrate its high ability to enter into cancer cells and release a combination of the anticancer drug doxorubicin and the non-steroidal anti-inflammatory drug indomethacin to achieve synergistic chemo-anti-inflammatory effects and overcome multidrug resistance. This study highlights the great potential of tumor targeting and dual stimuli-responsive nanoparticles for an efficient delivery of multidrug combination to improve the cancer therapeutic efficacy., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A simple and cost-effective way to prepare water-stable zein-based nanofibers for potential drug delivery was presented in this article. Corn protein zein was co-electrospun with hydrophobic ethyl cellulose. Indomethacin, as a model drug, was incorporated in situ into the composite nanofibers. Scanning electron microscopy and element mapping revealed the morphologies of drug-loaded nanofibers and drug distribution, respectively. Fourier transform infrared spectra confirmed the physical blending among the components. Differential scanning calorimetry and X-ray diffraction demonstrated the physical state of drug and polymers in the nanofiber matrix. The composite nanofibers showed a sustained diffusion-controlled release according to the results of in vitro dissolution tests., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The solubility of a drug is ultimately governed by its chemical potential as it is present in the undissolved solute. For a pharmaceutical amorphous solid dispersion (ASD), its solubility depends on the state and composition of the undissolved solute when the ASD is equilibrated with water. Concerning the undissolved solute phase that can contain up to 3 components (drug, polymer, and water), we developed a complete thermodynamic model to calculate the chemical potential of a drug in the multicomponent, amorphous system. This approach enables the estimation of the true solubility advantage of ASD from calorimetric measurements and moisture sorption isotherms. Both theoretical estimation and experimental studies, using indomethacin (IMC)/Eudragit E ASD systems, show that the solubility advantage of the amorphous IMC is significantly reduced through ASD formation and water partitioning. For the ASD with 70% drug loading, the solubility of IMC is lower than its crystalline counterpart. Our results show that stabilization through the ASD formation and water sorption can be manifested by the lowering of drug solubility; they demonstrate that the core property in ASD development is the drug chemical potential, which is essentially the thermodynamic driving force and can be quantitated using the model presented in this work., (Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)

Pharmaceutical degradation in conventional wastewater treatment plants (WWTP) represents a challenge since municipal wastewater and hospital effluents contain pharmaceuticals in low concentrations (recalcitrant and persistent in WWTP) and biodegradable organic matter (BOM) is the main pollutant. This work shows the feasibility of coupling electro-oxidation with a biological system for the simultaneous removal of recalcitrant drugs (bezafibrate, gemfibrozil, indomethacin and sulfamethoxazole (BGIS)) and BOM from wastewater. High removal efficiencies were attained without affecting the performance of activated sludge. BGIS degradation was performed by advanced electrochemical oxidation and the activated sludge process for BOM degradation in a continuous reactor. The selected electrochemical parameters from microelectrolysis tests (1.2 L s(-1) and 1.56 mA cm(-2)) were maintained to operate a filter press laboratory reactor FM01-LC using boron-doped diamond as the anode. The low current density was chosen in order to remove drugs without decreasing BOM and chlorine concentration control, so as to avoid bulking formation in the biological process. The wastewater previously treated by FM01-LC was fed directly (without chemical modification) to the activated sludge reactor to remove 100% of BGIS and 83% of BOM; conversely, the BGIS contained in wastewater without electrochemical pre-treatment were persistent in the biological process and promoted bulking formation.

Because of its weakly acidic nature (pKa of 4.5), indomethacin presents an aqueous solubility that significantly increases when changing from acidic to neutral/alkaline pH (1.5 μg/mL at pH 1.2 and 105.2 μg/mL at pH 7.4). We have therefore investigated the impact of the dissolution medium pH on the dissolution performance of indomethacin:Kollidon VA64 extrudates. The impact of the drug loading on the dissolution properties of these systems was also examined (5%, 15%, 30%, 50%, 70%, and 90% drug loading). Time-resolved Raman spectroscopy along with in-line UV-vis spectrophotometry was employed to directly relate changes in dissolution behavior to physicochemical changes that occur to the extrudate during the test. The dissolution tests were performed in pH 2 HCl (to mimic the stomach conditions), and this was then switched during the experiment to pH 6.8 phosphate buffer (to simulate the poststomach conditions). The rotating disc dissolution rate test was also used to simultaneously measure the dissolution rate of both the drug and the polymer. We found that in pH 2 HCl buffer, for the 15% or higher drug-loaded extrudates, Kollidon VA64 preferentially dissolves from the exterior of the compact leaving an amorphous drug-rich hydrophobic shell, which, similarly to an enteric coating, inhibits the drug release. The in situ formation of an enteric coating has been previously hypothesized, and this has been the first time that is directly observed in a pH-variable dissolution test. The dissolution medium switch to pH 6.8 phosphate buffer, due to the large increase of the aqueous solubility of indomethacin at this pH, leads to rapid dissolution of the material forming the coating and therefore total drug release. In contrast, the 5% extrudate is fully hydrated and quickly dissolves at low pH pointing to a dissolution performance dependent on highly water-soluble Kollidon VA64.

Human serum albumin (HSA) has two major ligand-binding sites, sites I and II, and also hydrolyzes some compounds at both sites. In the present study, we investigated differences in esterase activity among HSA preparations, and also the effects of warfarin, indomethacin, and naproxen on the hydrolytic activities of HSA to aspirin and p-nitrophenyl acetate. The esterase activities of HSA to aspirin or p-nitrophenyl acetate were measured from the pseudo-first-order formation rate constant (kobs) of salicylic acid or p-nitrophenol by HSA. Inter-lot variations were observed in the esterase activities of HSA to aspirin and p-nitrophenyl acetate; however, the esterase activity of HSA to aspirin did not correlate with that to p-nitrophenyl acetate. The inhibitory effects of warfarin and indomethacin on the esterase activity of HSA to aspirin were stronger than that of naproxen. In contrast, the inhibitory effect of naproxen on the esterase activity of HSA to p-nitrophenyl acetate was stronger than those of warfarin and indomethacin. These results suggest that the administration of different commercial HSA preparations and the co-administration with site I or II high-affinity binding drugs may change the pharmacokinetic profiles of drugs that are hydrolyzed by HSA.

A new approach of transdermal drug delivery is the use of microneedles. This promising technique offers the potential to be broadly used for drug administration as it enables the dramatic increase in permeation of medicaments across the stratum corneum. The potential of microneedles has evolved to spawn a plethora of potential transdermal applications. In order to advance the microneedle capabilities and possibly revolutionize advanced drug delivery, this study introduces a novel transdermal electro-modulated hydrogel-microneedle array (EMH-MNA) device composed of a nano-porous, embeddable ceramic microneedle array as well as an optimized EMH for the electro-responsive delivery of indomethacin through the skin. The ex vivo permeation as well as drug release experiments were performed on porcine skin tissue to ascertain the electro-responsive capabilities of the device. In addition, the microbial permeation ability of the microneedles across the viable epidermis in both microneedle-punctured skin as well as hypodermic needle-punctured skin was determined. Ex vivo evaluation of the EMH-MNA device across porcine skin demonstrated that without electro-stimulation, significantly less drug release was obtained (±0.4540mg) as compared to electro-stimulation (±2.93mg)., (Copyright © 2015 Elsevier B.V. All rights reserved.)