tolfenamic-acid and Disease-Models--Animal

When Zika virus emerged as a public health emergency there were no drugs or vaccines approved for its prevention or treatment. We used a high-throughput screen for Zika virus protease inhibitors to identify several inhibitors of Zika virus infection. We expressed the NS2B-NS3 Zika virus protease and conducted a biochemical screen for small-molecule inhibitors. A quantitative structure-activity relationship model was employed to virtually screen ∼138,000 compounds, which increased the identification of active compounds, while decreasing screening time and resources. Candidate inhibitors were validated in several viral infection assays. Small molecules with favorable clinical profiles, especially the five-lipoxygenase-activating protein inhibitor, MK-591, inhibited the Zika virus protease and infection in neural stem cells. Members of the tetracycline family of antibiotics were more potent inhibitors of Zika virus infection than the protease, suggesting they may have multiple mechanisms of action. The most potent tetracycline, methacycline, reduced the amount of Zika virus present in the brain and the severity of Zika virus-induced motor deficits in an immunocompetent mouse model. As Food and Drug Administration-approved drugs, the tetracyclines could be quickly translated to the clinic. The compounds identified through our screening paradigm have the potential to be used as prophylactics for patients traveling to endemic regions or for the treatment of the neurological complications of Zika virus infection.

Topics: Animals; Antiviral Agents; Artificial Intelligence; Chlorocebus aethiops; Disease Models, Animal; Drug Evaluation, Preclinical; High-Throughput Screening Assays; Immunocompetence; Inhibitory Concentration 50; Methacycline; Mice, Inbred C57BL; Protease Inhibitors; Quantitative Structure-Activity Relationship; Small Molecule Libraries; Vero Cells; Zika Virus; Zika Virus Infection

Tolfenamic acid, a nonsteroidal anti-inflammatory drug, alleviated learning and memory deficits and decreased the expression of specificity protein 1 (SP1)-mediated cyclin-dependent kinase-5 (CDK5), a major protein kinase that regulates hyperphosphorylated tau, in Alzheimer's disease (AD) transgenic mice. However, whether tolfenamic acid can regulate the major tau protein kinase, glycogen synthase kinase-3β (GSK-3β), or tau protein phosphatase, protein phosphatase 2A (PP2A), further inhibiting hyperphosphorylation of tau, remains unknown. To this end, tolfenamic acid was administered i.p. in a GSK-3β overactivation postnatal rat model and orally in mice after intracerebroventricular (ICV) injection of okadaic acid (OA) to develop a PP2A inhibition model. We used four behavioural experiments to evaluate memory function in ICV-OA mice. In this study, tolfenamic acid attenuated memory dysfunction. Tolfenamic acid decreased the expression of hyperphosphorylated tau in the brain by inhibiting GSK-3β activity, decreasing phosphorylated PP2A (Tyr307), and enhancing PP2A activity. Tolfenamic acid also increased wortmannin (WT) and GF-109203X (GFX) induced phosphorylation of GSK-3β (Ser9) and prevented OA-induced downregulation of PP2A activity in PC12 cells. Altogether, these results show that tolfenamic acid not only decreased SP1/CDK5-mediated tau phosphorylation, but also inhibited GSK-3β and PP2A-mediated tau hyperphosphorylation in AD models.

Topics: Alzheimer Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Disease Models, Animal; Female; Glycogen Synthase Kinase 3 beta; Male; Mice; Mice, Inbred C57BL; ortho-Aminobenzoates; Phosphorylation; Protein Phosphatase 2; Rats; Rats, Wistar; tau Proteins

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Cell Line, Tumor; Cyclin-Dependent Kinase 5; Cyclooxygenase 2; Disease Models, Animal; Frontal Lobe; Humans; Memory; Mice, Knockout; ortho-Aminobenzoates; tau Proteins; Treatment Outcome

Tolfenamic acid is a nonsteroidal anti-inflammatory drug with neuroprotective properties, and it alleviates learning and memory deficits in the APP transgenic mouse model of Alzheimer's disease. However, whether tolfenamic acid can prevent motor and memory dysfunction in transgenic animal models of Huntington's disease (HD) remains unclear. To this end, tolfenamic acid was orally administered to transgenic R6/1 mice from 10 to 20 weeks of age, followed by several behavioral tests to evaluate motor and memory function. Tolfenamic acid improved motor coordination in R6/1 mice as tested by rotarod, grip strength, and locomotor behavior tests and attenuated memory dysfunction as analyzed using the novel object recognition test and passive avoidance test. Tolfenamic acid decreased the expression of mutant huntingtin in the striatum of 20-week-old R6/1 mice by inhibiting specificity protein 1 expression and enhancing autophagic function. Furthermore, tolfenamic acid exhibited antioxidant effects in both R6/1 mice and PC12 cell models. Collectively, these results suggest that tolfenamic acid has a good therapeutic effect on R6/1 mice, and may be a potentially useful agent in the treatment of HD.

Topics: Alzheimer Disease; Animals; Antioxidants; Behavior, Animal; Disease Models, Animal; Huntingtin Protein; Huntington Disease; Maze Learning; Memory Disorders; Mice; Mice, Transgenic; Mutation; ortho-Aminobenzoates; PC12 Cells; Psychomotor Performance; Rats

Tangles are deposits of hyperphosphorylated tau, which are found in multiple neurodegenerative disorders that are referred to as tauopathies, of which Alzheimer's disease (AD) is the most common. Tauopathies are clinically characterized by dementia and share common cortical lesions composed of aggregates of the protein tau.. In this study, we explored the therapeutic potential of tolfenamic acid (TA), in modifying disease processes in a transgenic animal model that carries the human tau gene (hTau).. Behavioral tests, Western blotting and Immunohistochemical analysis were used to demonstrate the efficacy of TA.. Treatment of TA improved improving spatial learning deficits and memory impairments in young and aged hTau mice. Western blot analysis of the hTau protein revealed reductions in total tau as well as in sitespecific hyperphosphorylation of tau in response to TA administration. Immunohistochemical analysis for phosphorylated tau protein revealed reduced staining in the frontal cortex, hippocampus, and striatum in animals treated with TA.. TA holds the potential as a disease-modifying agent for the treatment of tauopathies including AD.

Topics: Animals; Brain; Cognition; Disease Models, Animal; Female; Humans; Male; Maze Learning; Memory Disorders; Mice, Transgenic; Neuroprotective Agents; ortho-Aminobenzoates; tau Proteins; Tauopathies

Spinal bulbar muscular atrophy (SBMA) is a motor neuron disease caused by toxic gain of function of the androgen receptor (AR). Previously, we found that co-regulator binding through the activation function-2 (AF2) domain of AR is essential for pathogenesis, suggesting that AF2 may be a potential drug target for selective modulation of toxic AR activity. We screened previously identified AF2 modulators for their ability to rescue toxicity in a Drosophila model of SBMA. We identified two compounds, tolfenamic acid (TA) and 1-[2-(4-methylphenoxy)ethyl]-2-[(2-phenoxyethyl)sulfanyl]-1H-benzimidazole (MEPB), as top candidates for rescuing lethality, locomotor function and neuromuscular junction defects in SBMA flies. Pharmacokinetic analyses in mice revealed a more favorable bioavailability and tissue retention of MEPB compared with TA in muscle, brain and spinal cord. In a preclinical trial in a new mouse model of SBMA, MEPB treatment yielded a dose-dependent rescue from loss of body weight, rotarod activity and grip strength. In addition, MEPB ameliorated neuronal loss, neurogenic atrophy and testicular atrophy, validating AF2 modulation as a potent androgen-sparing strategy for SBMA therapy.

Topics: Animals; Benzimidazoles; Co-Repressor Proteins; Disease Models, Animal; Drosophila melanogaster; HEK293 Cells; Humans; Male; Mice, Transgenic; Muscular Atrophy, Spinal; Nerve Degeneration; ortho-Aminobenzoates; Phenotype; Pilot Projects; Protein Domains; Receptors, Androgen; Trinucleotide Repeat Expansion

Melioidosis is caused by the facultative intracellular bacterium Burkholderia pseudomallei and is potentially fatal. Despite a growing global burden and high fatality rate, little is known about the disease. Recent studies demonstrate that cyclooxygenase-2 (COX-2) inhibition is an effective post-exposure therapeutic for pulmonary melioidosis, which works by inhibiting the production of prostaglandin E2 (PGE2). This treatment, while effective, was conducted using an experimental COX-2 inhibitor that is not approved for human or animal use. Therefore, an alternative COX-2 inhibitor needs to be identified for further studies. Tolfenamic acid (TA) is a non-steroidal anti-inflammatory drug (NSAID) COX-2 inhibitor marketed outside of the United States for the treatment of migraines. While this drug was developed for COX-2 inhibition, it has been found to modulate other aspects of inflammation as well. In this study, we used RAW 264.7 cells infected with B pseudomallei to analyze the effect of TA on cell survival, PGE2 production and regulation of COX-2 and nuclear factor- kappaB (NF-ĸB) protein expression. To evaluate the effectiveness of post-exposure treatment with TA, results were compared to Ceftazidime (CZ) treatments alone and the co-treatment of TA with a sub-therapeutic treatment of CZ determined in a study of BALB/c mice. Results revealed an increase in cell viability in vitro with TA and were able to reduce both COX-2 expression and PGE2 production while also decreasing NF-ĸB activation during infection. Co-treatment of orally administered TA and a sub-therapeutic treatment of CZ significantly increased survival outcome and cleared the bacterial load within organ tissue. Additionally, we demonstrated that post-exposure TA treatment with sub-therapeutic CZ is effective to treat melioidosis in BALB/c mice.

Topics: Animals; Burkholderia pseudomallei; Ceftazidime; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Disease Models, Animal; Female; Humans; Melioidosis; Mice; Mice, Inbred BALB C; ortho-Aminobenzoates; Post-Exposure Prophylaxis

We have previously reported that tolfenamic acid treatment decreases the amyloidogenic proteins in C57BL/6 and in old hemizygous R1.40 transgenic mice via the degradation of the transcription factor specificity 1 protein (Sp1). The lowering of amyloid-β protein precursor (AβPP) and amyloid-β (Aβ) in hemizygous R1.40 transgenic mice was accompanied by reversal of the identified spatial reference and working memory deficits observed in the mouse model. In this study, we examined the ability of tolfenamic acid to reduce the amyloid plaque burden, as well as to ameliorate spatial learning and memory deficits in homozygous R1.40 mice. Results from immunohistochemical analysis indicated that tolfenamic acid treatment resulted in a profound decrease in cerebral Aβ plaque burden that was accompanied by improvements in spatial working memory assessed by spontaneous alternation ratio in the Y-maze. These results provide further evidence that tolfenamic acid could be utilized as a repurposed drug to modify Alzheimer's disease pathogenesis.

Topics: Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cognition Disorders; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Gene Expression Regulation; Humans; Male; Maze Learning; Memory Disorders; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutation; ortho-Aminobenzoates; Time Factors

Tau and its aggregates are linked to the pathology of Alzheimer's disease (AD) and other tauopathies and, therefore, are explored as therapeutic targets for such disorders. Tau belongs to a family of microtubule-associated proteins that promote microtubule assembly. When hyperphosphorylated, tau becomes prone to forming aggregates. Increased brain levels of hyperphosphorylated tau correlate with dementia. Specificity protein 1 (Sp1), a transcription factor elevated in AD, is responsible for the transcription of AD-related proteins including the amyloid precursor protein, tau, and its cyclin-dependent kinase-5 (CDK5) activators. Tolfenamic acid promotes the degradation of Sp1, our previous studies demonstrated its ability to down-regulate transcriptional targets of Sp1 like amyloid precursor protein and reduce amyloid beta (Aβ), the main component of AD plaques. In this study, we administered tolfenamic acid daily to hemizygous R1.40 transgenic mice for 34 days, and examined tau and CDK5 gene and protein expression within the brain. Our results demonstrate that tolfenamic acid lowers tau mRNA and protein, as well as the levels of its phosphorylated form and CDK5. Thus, we present a drug candidate that inhibits the transcription of multiple major intermediates in AD pathology, thereby helping uncover a new mechanism-based approach for targeting AD. A new approach for targeting Alzheimer's disease through a transcriptional based mechanism is presented. Tolfenamic acid lowers the levels of tau, which forms pathological aggregates in Alzheimer's disease and other tauopathies, by promoting the degradation of the transcription factor specificity protein 1 which regulates tau transcription.

Topics: Age Factors; Amyloid beta-Protein Precursor; Analysis of Variance; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cyclin-Dependent Kinase 5; Dementia; Disease Models, Animal; Gene Expression Regulation; Mice; Mice, Transgenic; ortho-Aminobenzoates; Phosphorylation; tau Proteins

Tolfenamic acid lowers the levels of the amyloid precursor protein (APP) and amyloid beta (Aβ) when administered to C57BL/6 mice by lowering their transcriptional regulator specificity protein 1 (SP1). To determine whether changes upstream in the amyloidogenic pathway that forms Aβ plaques would improve cognitive outcomes, we administered tolfenamic acid for 34 days to hemizygous R1.40 transgenic mice. After the characterization of cognitive deficits in these mice, assessment of spatial learning and memory functions revealed that treatment with tolfenamic acid attenuated long-term memory and working memory deficits, determined using Morris water maze and the Y-maze. These improvements occurred within a shorter period of exposure than that seen with clinically approved drugs. Cognitive enhancement was accompanied by reduction in the levels of the SP1 protein (but not messenger RNA [mRNA]), followed by lowering both the mRNA and the protein levels of APP and subsequent Aβ levels. These findings provide evidence that tolfenamic acid can disrupt the pathologic processes associated with Alzheimer's disease (AD) and are relevant to its scheduled biomarker study in AD patients.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Cognition; Disease Models, Animal; Down-Regulation; Maze Learning; Memory Disorders; Memory, Long-Term; Memory, Short-Term; Mice; Mice, Inbred C57BL; Mice, Transgenic; Nootropic Agents; ortho-Aminobenzoates; Sp1 Transcription Factor

The primary objective of this study is to identify small molecules that target critical transcription factors for potential application in the chemoprevention of esophageal cancer. Specificity proteins (Sp) play a critical role in the growth and metastasis of several malignancies including esophageal cancer. Researchers at the M. D. Anderson Cancer Center Orlando Cancer Research Institute have reported previously that tolfenamic acid (TA) inhibits cancer cell proliferation and tumor growth through the degradation of Sp1, Sp3, and Sp4. We evaluated the chemopreventive properties of TA against esophageal tumorigenesis in N-nitrosomethylbenzylamine (NMBA)-induced murine tumor model. Fischer-344 rats were treated with NMBA (0.5 mg/kg s.c. 3 times a week) for 5 weeks to initiate the tumor formation, and then treated with 50 mg/kg TA from week 6 through week 25. Tumor incidence, tumor multiplicity (number of papilloma per rat), and tumor volume were evaluated after 25 weeks. All rats in the control group that received only NMBA developed lesions (100% incidence), while the TA-treated group showed significantly lower (33%) tumor incidence and tumor multiplicity. Furthermore, the tumor volume was significantly diminished in the TA-treated group when compared with the control group. Using small molecules such as TA to target key transcription factors associated with tumorigenesis for the prevention of esophageal malignancies is a new and promising strategy. Results of the current study provide evidence that TA, when given orally after tumor initiation, can significantly suppress tumorigenesis induced by carcinogenic nitrosamines in rats. These appealing results demonstrate that TA may potentially serve as an effective chemopreventive agent in patient populations vulnerable to esophageal cancer.

Topics: Animals; Anticarcinogenic Agents; Carcinogens; Cell Line; Cell Survival; Dimethylnitrosamine; Disease Models, Animal; Esophageal Neoplasms; Humans; ortho-Aminobenzoates; Proto-Oncogene Proteins c-met; Rats; Rats, Inbred F344; Sp Transcription Factors; Tumor Burden

Evidence from our laboratory suggests that tolfenamic acid has a potential for slowing the progression of Alzheimer's disease (AD) through lowering cortical levels of the β-amyloid precursor protein (APP) and its pathogenic amyloid beta (Aβ) intermediates [1]. In this study, we examined the ability of tolfenamic acid to cross the blood brain barrier (BBB) by predicting its logBB and logPS values, the indexes of BBB permeability, using computational models. We also determined, via in vitro methods, the brain penetration capacity factor [(K(IAM)/MW(4))x10(10)] using phosphatidylcholine column chromatography. The obtained logBB, logPS and (K(IAM)/MW(4))x10(10) values predicted that tolfenamic acid can passively transfer into the central nervous system (CNS). These results were validated in vivo using LC-MS analysis after administration of tolfenamic acid intravenously to guinea pigs and mice. The present study provides the first evidence of the ability of tolfenamic acid to cross the BBB and offers a comparative analysis of approaches used to predict the ability of compounds to penetrate into the brain.

Topics: Alzheimer Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood-Brain Barrier; Brain; Cell Membrane Permeability; Disease Models, Animal; Drug Evaluation, Preclinical; Guinea Pigs; Mice; Mice, Inbred C57BL; ortho-Aminobenzoates

Injections of mild irritants intradermally (carrageenan, zymosan and dextran) and intracaveally (carrageenan) in a tissue cage model of inflammation were used in studies of the pharmacodynamics and pharmacokinetics of tolfenamic acid administered intramuscularly in calves. Inhibition of serum thromboxane (TX)B2 and inflammatory exudate prostaglandin (PG)E2 were used as indicators of the magnitude and time course of blockade of cyclo-oxygenase isoforms COX-1 and COX-2, respectively. Single doses of 2, 4 and 8 mgkg-1 tolfenamic acid partially inhibited irritant-induced rises in skin temperature (non-dose dependently) and skin oedema (dose-dependently). These doses also markedly inhibited serum TXB2 synthesis and the duration of inhibition was dose-related. A dose of 2 mgkg-1 tolfenamic acid also attenuated skin temperature rise over carrageenan-injected tissue cages, and markedly inhibited exudate PGE2 synthesis, even though drug penetration into both exudate and tissue cage transudate was limited. Tolfenamic acid pharmacokinetics were characterized by a relatively short tmax (0.94-2.04 h), a high estimated Vdarea (1.79-3.20 Lkg-1), an estimated t1/2 beta of 8.01-13.50 h and Cl beta of 0.142-0.175 Lkg-1h-1. The actions of tolfenamic acid in inhibiting PGE2 synthesis and in attenuating two of the cardinal signs of inflammation (heat and swelling) suggest that a dosage of 2 mgkg-1 administered intramuscularly should be effective clinically as an anti-inflammatory agent.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Cattle; Dinoprostone; Disease Models, Animal; Inflammation; Irritants; Male; ortho-Aminobenzoates; Skin Diseases; Skin Temperature; Thromboxane B2

An acute non-immune inflammation model was used to compare the action of two non-steroidal anti-inflammatory drugs, flunixin meglumine and tolfenamic acid, on prostaglandin E2 (PGE2) synthesis in bovine inflammatory exudate. The tissue cage model used involves subcutaneous implantation of polypropylene cages and subsequent stimulation by carrageenan injection of the granulation tissue which develops within the cage. Twelve calves were randomly assigned to three groups receiving placebo, flunixin meglumine and tolfenamic acid, respectively. Inflammatory exudate was sampled 30 min after carrageenan injection and at seven subsequent time points. PGE2 levels were determined by radioimmunoassay. At each time point post-carrageenan injection, flunixin meglumine inhibited PGE2 synthesis to a greater extent than tolfenamic acid. At 4, 8, 12 and 24 h these differences were statistically significant.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Carrageenan; Cattle; Clonixin; Dinoprostone; Disease Models, Animal; Exudates and Transudates; Inflammation; Male; ortho-Aminobenzoates; Random Allocation

Dithranol-induced skin irritation and the modulatory effects of different pharmacological agents were studied using the mouse ear model. A single topical application of dithranol caused a dose-dependent skin irritation which resulted in delayed swelling of the mouse ear with two separate peak responses, 1-2 and 6-10 days after application. The irritation was most effectively and persistently inhibited by topical treatment with corticosteroids, the free radical scavenger DL-alpha-tocopherol (DLAT) and the serotonin antagonist metergoline. The effect of corticosteroids, however, was slightly diminished during the second peak irritation. The lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA), the dual lipoxygenase and cyclo-oxygenase inhibitor tolfenamic acid and the cyclo-oxygenase inhibitor indomethacin as well as trifluoperazine retained their inhibitory activity. Of these compounds, indomethacin was active only during the first irritation peak, NDGA during both peaks and trifluoperazine principally during the second peak. Retinoic acid did not inhibit the ear swelling. The results confirm and extend the observations that the formation of free radicals is essential for dithranol inflammation. The inflammation can also be suppressed by inhibiting the formation of arachidonic acid or its pro-inflammatory metabolites.

Topics: Administration, Topical; Adrenal Cortex Hormones; Animals; Anthralin; Anti-Inflammatory Agents, Non-Steroidal; Disease Models, Animal; Dose-Response Relationship, Drug; Ear; Edema; Female; Hyperplasia; Indomethacin; Inflammation; Masoprocol; Mice; ortho-Aminobenzoates; Skin Diseases; Time Factors; Tretinoin; Trifluoperazine