The Importance of the Pyrazole Scaffold in the Design of Protein Kinases Inhibitors as Targeted Anticancer Therapies.

The altered activation or overexpression of protein kinases (PKs) is a major subject of research in oncology and their inhibition using small molecules, protein kinases inhibitors (PKI) is the best available option for the cure of cancer. The pyrazole ring is extensively employed in the field of medicinal chemistry and drug development strategies, playing a vital role as a fundamental framework in the structure of various PKIs. This scaffold holds major importance and is considered a privileged structure based on its synthetic accessibility, drug-like properties, and its versatile bioisosteric replacement function. It has proven to play a key role in many PKI, such as the inhibitors of Akt, Aurora kinases, MAPK, B-raf, JAK, Bcr-Abl, c-Met, PDGFR, FGFRT, and RET. Of the 74 small molecule PKI approved by the US FDA, 8 contain a pyrazole ring: Avapritinib, Asciminib, Crizotinib, Encorafenib, Erdafitinib, Pralsetinib, Pirtobrutinib, and Ruxolitinib. The focus of this review is on the importance of the unfused pyrazole ring within the clinically tested PKI and on the additional required elements of their chemical structures. Related important pyrazole fused scaffolds like indazole, pyrrolo[1,2-b]pyrazole, pyrazolo[4,3-b]pyridine, pyrazolo[1,5-a]pyrimidine, or pyrazolo[3,4-d]pyrimidine are beyond the subject of this work.

Effects of Venlafaxine, Risperidone and Febuxostat on Cuprizone-Induced Demyelination, Behavioral Deficits and Oxidative Stress.

Multiple sclerosis (MS) is a demyelinating, autoimmune disease that affects a large number of young adults. Novel therapies for MS are needed considering the efficiency and safety limitations of current treatments. In our study, we investigated the effects of venlafaxine (antidepressant, serotonin-norepinephrine reuptake inhibitor), risperidone (atypical antipsychotic) and febuxostat (gout medication, xanthine oxidase inhibitor) in the cuprizone mouse model of acute demyelination, hypothesizing an antagonistic effect on TRPA1 calcium channels. Cuprizone and drugs were administered to C57BL6/J mice for five weeks and locomotor activity, motor performance and cold sensitivity were assessed. Mice brains were harvested for histological staining and assessment of oxidative stress markers. Febuxostat and metabolites of venlafaxine (desvenlafaxine) and risperidone (paliperidone) were tested for TRPA1 antagonistic activity. Following treatment, venlafaxine and risperidone significantly improved motor performance and sensitivity to a cold stimulus. All administered drugs ameliorated the cuprizone-induced deficit of superoxide dismutase activity. Desvenlafaxine and paliperidone showed no activity on TRPA1, while febuxostat exhibited agonistic activity at high concentrations. Our findings indicated that all three drugs offered some protection against the effects of cuprizone-induced demyelination. The agonistic activity of febuxostat can be of potential use for discovering novel TRPA1 ligands.

New 2-(4-(4-bromophenylsulfonyl)phenyl)-4-arylidene-oxazol-5(4H)-ones: analgesic activity and histopathological assessment.

This paper reports the synthesis, analgesic activity, acute toxicity and histopathological (HP) assessment of four new compounds from oxazol-5(4H)-ones class that contain in their molecule a diarylsulfone moiety. The new 2-(4-(4-bromophenylsulfonyl)phenyl)-4-arylidene-oxazol-5(4H)-ones were obtained by reaction of 2-(4-(4-bromophenyl-sulfonyl)benzamido)acetic acid intermediate with aromatic aldehydes (benzaldehyde, 4-methoxy, 4-nitro or 4-bromobenzaldehyde), in acetic anhydride and in the presence of anhydrous sodium acetate. The new compounds have been characterized by spectral techniques, such as: Fourier-transform infrared spectroscopy (FT-IR), mass spectrometry (MS), proton nuclear magnetic resonance (1H-NMR) and by elemental analysis. The acute toxicity of the new oxazol-5(4H)-ones in mice was assessed through "acute toxic class" method, according to Organization for Economic Co-operation and Development (OECD) Guidelines. The HP assessment of some preserved organs collected from mice has been performed. The analgesic activity of all new synthesized compounds was carried out with two pharmacological tests: the writhing test and the hot plate test. In order to predict the binding affinities of the synthesized oxazol-5(4H)-ones derivatives against molecular targets involved in pain and inflammation, molecular docking simulations were performed. The results of the writhing test indicated that the most active compound was the oxazolone that contains in the molecule a methoxy group. The acute oral toxicity study revealed no lethal effect of new compounds. The HP assessment of the preserved organs collected from mice did not indicate any cytohistopathological aspects that can be linked to any inflammatory, neoplastic or cytotoxic process, demonstrating the low toxicity of new compounds.

Predictive power of the Triticum root elongation test for the assessment of novel anti­proliferative therapies.

The use of alternative techniques to reduce the number of animals used in anticancer research is an issue of current interest. The aim of this study was to validate the use of a simple and efficient alternative tool for the assessment of the potential of novel anti­proliferative agents. A set of 20 compounds with various mechanisms were tested in the Triticum aestivum root elongation assay, using aminophylline as negative control. Hierarchical cluster analyses were performed using the furthest neighbor method based on Euclidean distance measure, and the compounds were statistically analyzed in reference to their anti­proliferative pattern registered in the NCI60 human tumor cell line anticancer drug screen. A correlation between the Triticum test results and the NCI60 anti­proliferative profile was made for a number of human cells that we defined as the Triticum cell panel. Linear equations were computed that can be used to transform the inhibitory effect measured in any future Triticum assay in order to predict the effect on particular human cells. Of the tested anti­proliferative agents, methotrexate, colchicine, cantharidin, cisplatin and verapamil produced a growth inhibition over 50%. On the whole, the findings of this study suggest that the Triticum test can be used to detect several types of anti­proliferative mechanisms, particularly those targeting tubulin, rendering it a useful tool with which to identify novel mitotic spindle inhibitors.

Toxicity of plant extracts containing pyrrolizidine alkaloids using alternative invertebrate models.

Pyrrolizidine alkaloids (PAs) are a widespread class of hepatotoxic heterocyclic organic compounds found in approximately 3% of world flora. Some PAs have been shown to have genotoxic and carcinogenic effects. The present study focuses on the toxicity effects of four dry extracts obtained from medicinal plants (Senecio vernalis, Symphytum officinale, Petasites hybridus and Tussilago farfara), on two aquatic organisms, Artemia salina and Daphnia magna, and the correlation with their PAs content. A new GC­MS method, using a retention time (TR)­5MS type capillary column was developed. PAs Kovats retention indices, for this type of column were computed for the first time. The lethal dose 50% (LC50) values for the two invertebrate models were correlated (Pearson 's coefficient, >0.9) and the toxicity was PA concentration-dependent, for three of the four extracts. All tested extracts were found to be toxic in both aquatic organism models. The results can be used to develop a GC­MS validated method for the assay of PAs in medicinal plants with a further potential application in the risk assessment study of PAs toxicity in humans.

Therapeutic potential of certain drug combinations on paclitaxel-induced peripheral neuropathy in rats.

BACKGROUND AND AIMS: Experimental research and clinical data support the potential combination therapy for the treatment of neuropathic pain. We aimed to investigate the analgesic effect of the following associations: gabapentin + etifoxine; tramadol + etifoxine; gabapentin + tramadol, in an experimental model of peripheral neuropathy induced by paclitaxel. MATERIALS AND METHODS: Neuropathy was induced in male Wistar rats by the daily administration of 2 mg÷kg body weight (bw) paclitaxel intraperitoneally, four days in a row. Analgesics were given concomitantly with paclitaxel, in the following doses: tramadol 15 mg÷kg bw, etifoxine 100 mg÷kg bw, gabapentin 300 mg÷kg bw. Tactile allodynia and mechanical hyperalgesia were assessed using the Dynamic Plantar Aesthesiometer apparatus (Ugo Basile). After 18 days of treatment, the brain and liver tissue susceptibility to lipid peroxidation was evaluated and the sciatic nerve histological examination of the effect on myelin fibers was performed. RESULTS AND CONCLUSIONS: Experimental data have shown a strong analgesic effect of these three tested combinations expressed mainly by the statistically significant increased maximum response time, both in the assessment of allodynia and hyperalgesia. The gabapentin + tramadol combination lead to the maximum analgesic effect, immediately after the discontinuation of paclitaxel (44.94%, p<0.0001) and throughout the study. The treatment associated with tramadol caused a reduction in lipid peroxidation in the brain as compared to paclitaxel group. Combination therapy showed reduced damage to myelinated fiber density in the sciatic nerve. The drug combinations used in the experiment showed therapeutic potential in the fight against neuropathic pain induced by the administration of taxanes.

Pharmacotoxicological screening on new derivatives of beta-phenylethylamine, potential agonists of beta3-adrenergic receptors.

BACKGROUND AND AIMS: Beta3-adrenergic receptors (beta3-ARs) have been initially characterized in 1989. Afterwards, their tissue distribution was established: white and brown adipose tissue, central nervous system, myocardium (atrial and ventricular), blood vessels, smooth gastrointestinal muscles (stomach, small intestine, colon), gallbladder, urinary bladder, prostate, skeletal muscles. Non-clinical trials have demonstrated the major implication of beta3-ARs in glucose metabolism, implicitly, in insulin release, and also in obesity. Therefore, new compounds were synthesized starting from beta-phenylethylamine nucleus and substituted in various positions, for possible antidiabetic and÷or antiobesity action. MATERIALS AND METHODS: In the present research, the antidiabetic action of newly synthesized compounds was investigated on an experimental model of alloxan-induced diabetes, administered in dose of 130 mg÷kg body weight (bw), intraperitoneally (i.p.). After 14 days of treatment, glycemia and enzymes involved in homeostasis of glucose metabolism, glucose-6-phosphate dehydrogenase (G6PD), glucose-6-phosphatase (G6Pase) and hexokinase were determined. Animals were then euthanized and histopathology examinations were performed on harvested liver, kidney, spleen and brain in order to document pathological changes induced by alloxan-induced diabetes and÷or by tested compounds. RESULTS AND CONCLUSIONS: Glycemia in animals treated with the tested compounds decreased statistically significant for groups C2 and C3 (-42.13% and -37.2%, respectively), compared to diabetic control group. C2 was also the compound to favorably modify the dynamics of determined enzymes, together with the display of very good safety profile supported by minor, non-significant, histopathological changes.

Oral toxicity study of certain plant extracts containing pyrrolizidine alkaloids.

Pyrrolizidine alkaloids (PAs) are a class of toxic compounds which are found in plants. Poisoning caused by these toxins is associated with acute and chronic liver damage. Tussilago farfara (coltsfoot), Petasites hybridus (common butterbur), Senecio vernalis (eastern groundsel) and Symphytum officinale (comfrey) are traditional phytotherapic species, which beside the therapeutic bioactive compounds contain PAs. The aim of the paper was to assess the safety of some dry extracts obtained from these species. For the determination of acute toxicity, Organization for Economic Cooperation and Development (OECD) Guideline No. 423 was used. For the determination of repeated dose oral toxicity, Senecionis vernalis herba and Symphyti radix extracts (250 mg÷kg) were administrated, by gavage, for 28 days, and their effects on animal weight, liver and biliary functions, hepatic tissue and oxidative stress were investigated. After the acute toxicity testing, the dry extracts were placed in the GHS Category V (LD50>5000 mg÷kg, p.o.). For the subacute toxicity testing, no death or any signs of toxicity were observed. Also, no significant differences in biochemical parameters were observed between control and treated groups. The observed histopathological lesions were non-specific and were not consistent with the data reported in the literature for PAs exposure. In conclusion, the administration for 28 days, of the tested extracts, in a dose which correspond to a PAs concentration over the limits imposed in some countries, produced no hepatic and biliary toxic effects. Further studies, extended over a longer period of time, are needed in order to determine the safety of plant extracts containing PAs.

Influence of hyperforin on the morphology of internal organs and biochemical parameters, in experimental model in mice.

BACKGROUND AND AIMS: Hyperforin (HY) is used to treat depression and skin irritation and has been shown a number of pharmacological activities. The literature does no cite data on changes that may occur in the body after HY intake (ethylene diammonium salt - EDS) in long-term administration. From this point of view, the present work is a key to determining the pharmacotoxicological profile of the HY-EDS, in long-term administration. MATERIALS AND METHODS: In present research, the influence of toxic doses of HY-EDS was investigated on the biochemical serum parameters and the histopathological changes in internal organs on the experimental mice model. For acute toxicity determination, the HY-EDS was tested in doses of 2000-5000 mg÷kg, administered once per day orally. For subacute toxicity, the HY-EDS was tested in three groups of mice, in doses of 50, 75 and 100 mg÷kg÷day, administered once daily, for 28 consecutive days. RESULTS AND CONCLUSIONS: As concern acute toxicity, a lethal effect has not occurred at any of the two tested doses and HY-EDS was classified as Class V toxic: median lethal dose (LD50) >5000 mg÷kg, p.o. After 14 days of follow-up in acute toxicity, the experimental results showed a statistically significant increase of aspartate transaminase (AST) and alanine transaminase (ALT), compared to the control group. There were no changes in creatinine and serum glucose compared to the control group. After the administration of repeated doses, it was observed an increase of serum transaminases and alkaline phosphatase. Histological examination revealed that the liver injuries were in an initial stage, making them reversible in case of HY-EDS treatment discontinuation. There was no evidence of kidney damage to any of the doses of HY-EDS.