Melatonin is widely present in Nature. It has pleiotropic activities, in part mediated by interactions with high-affinity G-protein-coupled melatonin type 1 and 2 (MT1 and MT2) receptors or under extreme conditions, e.g., ischemia/reperfusion. In pharmacological concentrations, it is given to counteract the massive damage caused by MT1- and MT2-independent mechanisms. The aryl hydrocarbon receptor (AhR) is a perfect candidate for mediating the latter effects because melatonin has structural similarity to its natural ligands, including tryptophan metabolites and indolic compounds. Using a cell-based Human AhR Reporter Assay System, we demonstrated that melatonin and its indolic and kynuric metabolites act as agonists on the AhR with EC₅₀’s between 10⁻⁴ and 10⁻⁶ M. This was further validated via the stimulation of the transcriptional activation of the CYP1A1 promoter. Furthermore, melatonin and its metabolites stimulated AhR translocation from the cytoplasm to the nucleus in human keratinocytes, as demonstrated by ImageStream II cytometry and Western blot (WB) analyses of cytoplasmic and nuclear fractions of human keratinocytes. These functional analyses are supported by in silico analyses. We also investigated the peroxisome proliferator-activated receptor (PPAR)γ as a potential target for melatonin and metabolites bioregulation. The binding studies using a TR-TFRET kit to assay the interaction of the ligand with the ligand-binding domain (LBD) of the PPARγ showed agonistic activities of melatonin, 6-hydroxymelatonin and N-acetyl-N-formyl-5-methoxykynuramine with EC₅₀’s in the 10⁻⁴ M range showing significantly lower affinities that those of rosiglitazone, e.g., a 10⁻⁸ M range. These interactions were substantiated by stimulation of the luciferase activity of the construct containing PPARE by melatonin and its metabolites at 10⁻⁴ M. As confirmed by the functional assays, binding mode predictions using a homology model of the AhR and a crystal structure of the PPARγ suggest that melatonin and its metabolites, including 6-hydroxymelatonin, 5-methoxytryptamine and N-acetyl-N-formyl-5-methoxykynuramine, are excellent candidates to act on the AhR and PPARγ with docking scores comparable to their corresponding natural ligands. Melatonin and its metabolites were modeled into the same ligand-binding pockets (LBDs) as their natural ligands. Thus, functional assays supported by molecular modeling have shown that melatonin and its indolic and kynuric metabolites can act as agonists on the AhR and they can interact with the PPARγ at high concentrations. This provides a mechanistic explanation for previously reported cytoprotective actions of melatonin and its metabolites that require high local concentrations of the ligands to reduce cellular damage under elevated oxidative stress conditions. It also identifies these compounds as therapeutic agents to be used at pharmacological doses in the prevention or therapy of skin diseases.

Markers of bladder cancer cells remain elusive, which is a major cause of the low recognition of this malignant neoplasm and its recurrence. This implies an urgent need for additional diagnostic tools which are based on the identification of the chemism of bladder cancer. In this study, we employed label-free techniques of molecular imaging—Fourier Transform Infrared and Raman spectroscopic imaging—to investigate bladder cancer cell lines of various invasiveness (T24a, T24p, HT-1376, and J82). The urothelial HCV-29 cell line was the healthy control. Specific biomolecules discriminated spatial distribution of the nucleus and cytoplasm and indicated the presence of lipid bodies and graininess in some cell lines. The most prominent discriminators are the total content of lipids and sugar moieties as well as the presence of glycogen and other carbohydrates, un/saturated lipids, cytochromes, and a level of S-S bridges in proteins. The combination of the obtained hyperspectral database and chemometric methods showed a clear differentiation of each cell line at the level of the nuclei and cytoplasm and pointed out spectral signals which differentiated bladder cancer cells. Registered spectral markers correlated with biochemical composition changes can be associated with pathogenesis and potentially used for the diagnosis of bladder cancer and response to experimental therapies.

Cannabinoids (CBs) are used to treat chronic pain, chemotherapy-induced nausea and vomiting, and multiple sclerosis spasticity. Recently, the medicinal use of CBs has attracted increasing interest as a new therapeutic in many diseases. Data indicate a correlation between CBs and PPARs via diverse mechanisms. Both the endocannabinoid system (ECS) and peroxisome proliferator-activated receptors (PPARs) may play a significant role in PCOS and PCOS related disorders, especially in disturbances of glucose-lipid metabolism as well as in obesity and fertility. Taking into consideration the ubiquity of PCOS in the human population, it seems indispensable to search for new potential therapeutic targets for this condition. The aim of this review is to examine the relationship between metabolic disturbances and obesity in PCOS pathology. We discuss current and future therapeutic interventions for PCOS and related disorders, with emphasis on the metabolic pathways related to PCOS pathophysiology. The link between the ECS and PPARs is a promising new target for PCOS, and we examine this relationship in depth.

The balance between apoptosis and autophagy plays a key role in cancer biology and treatment strategies. The aim of this study was to assess the effect of the mTOR kinase inhibitor everolimus and chloroquine on the regulation of proliferation, caspase-3 activation, and apoptosis in melanoma cells. We studied the activity of caspase-3 and the levels of caspase-3 and -9 using the Western blot technique. Cellular apoptosis was examined using a DNA fragmentation assay, and changes in the cell nucleus and cytoskeleton were examined using fluorescence microscopy DAPI, OA/IP. We also studied the rearrangement of lipid structures using fluorescent dyes: Nile Red and Nile Blue. A low nanomolar concentration of the mTOR kinase inhibitor everolimus in combination with chloroquine activated the apoptosis process and decreased cell proliferation. These changes were accompanied by an obvious change in cell morphology and rearrangement of lipid structures. Alterations in lipid redistribution accompanying the process of apoptosis and autophagy are among the first to occur in the cell and can be easily monitored in in vitro studies. The combination of mTOR inhibitors and chloroquine represents a promising area of research in cancer therapy. It has the potential to enhance treatment efficacy through complementary mechanisms.

Despite the development of modern drugs, drug resistance in oncology remains the main factor limiting the curability of patients. This paper shows the use of a group of hydrophobic statins to inhibit drug resistance (Pgp protein). In a chemoresistance melanoma cell model, viability, necroptosis with DNA damage, the absorption of the applied pharmaceuticals, and the functional activity of the ABCB1 drug transporter after administration of docetaxel or docetaxel with a selected hydrophobic statin were studied. Taxol-resistant human melanoma cells from three stages of development were used as a model: both A375P and WM239A metastatic lines and radial growth phase WM35 cells. An animal model (Mus musculus SCID) was developed for the A375P cell line. The results show that hydrophobic statins administered with docetaxel increase the accumulation of the drug in the tumor cell a.o. by blocking the ABCB1 channel. They reduce taxol-induced drug resistance. The tumor size reduction was observed after the drug combination was administrated. It was shown that the structural similarity of statins is of secondary importance, e.g., pravastatin and simvastatin. Using cytostatics in the presence of hydrophobic statins increases their effectiveness while reducing their overall toxicity.

The presented paper describes a new isolation method of recovery and analysis of selected drugs developed for preclinical research. The method uses the RP-HPLC technique (in a single chromatographic separation) and serves the recovery and analysis of selected drugs from neoplastic cells. It enables the determination of cytostatics statins, fibrates, and pioglitazone. Chromatographic separations of the tested compounds were carried out on a Gemini-NX 5 µ C18 (4.6 × 150 mm i.d.) column, in a gradient system with a mobile phase consisting of ACN (0.1% TFA) and water (0.1% TFA) at ambient temperature. The separations were carried out at a flow of 1 ml/min and UV detection of 220 nm. The inter-day and intra-day precision and accuracy of the method were determined. Extending the extraction time at reduced temperature resulted in a significant increase in the recovery of the pharmaceuticals in comparison with traditional extraction methods. The presence of the tested pharmaceuticals at defined retention times was confirmed by mass spectrometry. A recovery procedure for the tested compounds from biological material (medium, cell pellets) was developed at a level ranging between 93 and 99%. The utility of the new HPLC method has been confirmed in drug absorption studies as screening tests for the analysis of the new therapeutic compositions on melanoma cell lines.