Synthesis and activity of the salicylic acid ester of bakuchiol in psoriasis-surrogate keratinocytes and skin substitutes.

BACKGROUND: Topical retinoids are effective in retarding skin ageing and restoring homeostasis in skin conditions such as psoriasis. However their adverse effects (AEs), which include irritation (retinoid dermatitis), photosensitivity and teratogenicity, limit their use and patient compliance. Development of retinoid analogues with minimal AEs would allow a broader and more compliant use. AIM: To synthesise a novel molecule, bakuchiol salicylate (bakusylan), with a modulatory gene expression profile similar to retinoids, using as reference three prescription retinoids: tretinoin, tazarotene and adapalene. METHODS: We hypothesized that because bakuchiol salicylate has a structure entirely different from existing retinoids, there would be at least a partial uncoupling of AEs from the skin-normalizing activity of this retinoid. This hypothesis was tested at the transcriptional level in psoriatic cytokine-treated cultures of keratinocytes and organotypic skin substitutes, using DNA microarrays and custom PCR arrays. RESULTS: Evaluation of the gene expression profile of bakuchiol salicylate revealed elimination of several components of the retinoid-like proinflammatory response and teratogenic signature, without a substantial loss of normalizing potential. A possible mechanism of action, consisting of keratinocyte desensitization to psoriatic cytokine signalling through inhibition of the signal transducer and regulator of transcription (STAT)1/3/interferon inflammatory signal transduction axis was also identified. CONCLUSION: Bipartite materials obtained by merging two skin-active entities with specific, complementary bioactivities, such as bakuchiol and salicylic acid, may yield a new class of functional retinoids.

2,4-Disubstituted pyridine derivatives are effective against intracellular and biofilm-forming tubercle bacilli.

It was recently reported that 4-substituted picolinohydrazonamides carrying hydrophilic cyclic amines, such as morpholine and pyrrolidine, at the end of their thiosemicarbazide chain have potent antimycobacterial activity in vitro at concentrations below 1 µg/ml. Here, two selected compounds, 2,4-disubstituted pyridine derivatives 11 and 15, revealed significant bactericidal activity against Mycobacterium tuberculosis localized intracellularly within human macrophages, as well as against biofilm-forming tubercle bacilli. Mutants were selected that were resistant to the investigated compounds at an efficiency similar to that identified in the presence of the first line antituberculosis drug rifampicin. The resistant mutants were viable in the presence of the tested compounds exclusively on solid media. Genome-wide sequencing of the mutants selected in the presence of compound 11 revealed the accumulation of nonsynonymous mutations in the mmpR5 gene encoding a transcriptional repressor of the MmpS5-MmpL5 efflux pump, whose upregulation has been associated with bedaquiline resistance. The depletion of MmpR5 in wild-type M. tuberculosis using CRISPR-Cas9 technology increased the resistance of this strain to compound 11. Mass spectrometry-based proteomics (LC-MS/MS) of wild-type tubercle bacilli growing in subinhibitory concentrations of compounds 11 or 15 revealed 15 overproduced proteins not detectable in the control cells, including virulence-related proteins.

Anthrapyridones, a novel group of antitumour non-cross resistant anthraquinone analogues. Synthesis and molecular basis of the cytotoxic activity towards K562/DOX cells.

1. Multidrug resistance (MDR) to antitumour agents, structurally dissimilar and having different intracellular targets, is the major problem in cancer therapy. MDR phenomenon is associated with the presence of membrane proteins which belong to the ATP-binding cassette family transporters responsible for the active drug efflux leading to the decreased intracellular accumulation. 2. The search of new compounds able to overcome MDR is of prime importance. 3. Recently we have synthesized a new family of anthrapyridone compounds. The series contained derivatives modified with appropriate hydrophobic or hydrophylic substituents at the side chain. 4. The interaction of these derivatives with erythroleukemia K562 sensitive and K562/DOX resistant (overexpressing P-glycoprotein) cell lines has been examined. The study was performed using a spectrofluorometric method which allows to continuously follow the uptake and efflux of fluorescent molecules by living cells. 5. It was demonstrated that the increase in the lipophilicity of anthrapyridones favoured the very fast cellular uptake exceeding the rate of P-gp dependent efflux out of the cell. For these derivatives, very high accumulation (the same for sensitive and resistant cells) was observed and the in vitro biological data confirmed that these compounds exhibited comparable cytotoxic activity towards sensitive and P-gp resistant cell line. In contrast, anthrapyridones modified with hydrophylic substituents exhibited relatively low kinetics of cellular uptake. 6. For these derivatives decreased accumulation in resistant cells was observed and the in vitro biological data demonstrated that they were much less active against P-gp resistant cells in comparison to sensitive cells. 7. We also studied, using confocal microscopy, the intracellular distribution of anthrapyridones in NIH-3T3 cells. Our data showed that these compounds were strongly accumulated in the nucleus and lysosomes.

Transport of new non-cross-resistant antitumor compounds of the benzoperimidine family in multidrug resistant cells.

Multidrug resistance (MDR) phenotype in mammalian cells is often correlated with overexpression of P-glycoprotein or multidrug resistance-associated protein (MRP1). Both proteins are energy-dependent drug efflux pumps that efficiently reduce the intracellular accumulation and hence the cytotoxicity of many natural cytotoxins. The influx and efflux of drugs across the cell membrane are in large part responsible for their intracellular concentrations, and in the search for new compounds able to overcome MDR, it is of prime importance to determine the molecular parameters whose modification would lead to an increase in the kinetics of uptake and/or to a decrease in the pump-mediated efflux. Here, we studied three members of a new family of benzoperimidine antitumor compounds which exhibit comparable cytotoxicity towards resistant cells expressing P-glycoprotein, or MRP1, and sensitive cells. We used spectrofluorometric methods to determine the kinetics of the uptake and release of these three drugs in different cell lines: the erythroleukemia cell line K562 and the resistant K562/Adr expressing P-glycoprotein, the small-cell lung cancer cell line GLC4 and resistant GLC4/Adr expressing MRP1. We also studied, using confocal microscopy, the intracellular distribution of these drugs in NIH/3T3 cells. Our data show that (i) the kinetics for the uptake of these drugs is very rapid, higher than 2 x 10(-17) mole cell(-1) s(-1), (ii) the drugs are strongly accumulated in the nucleus and lysosomes, (iii) the three drugs are recognized and pumped out by both transporters, as shown by the inhibition of P-glycoprotein- and MRP1-mediated efflux of pirarubicin by benzoperimidine, with inhibitory constants of 1.5 and 2.1 microM for P-glycoprotein and MRP1, respectively, suggesting that benzoperimidine is transported by the two transporters with K(m) approximately 2 microM. In conclusion, the fast uptake kinetics of the benzoperimidines counterbalance their efflux by P-glycoprotein and MRP1.

The role of structural factors of anthraquinone compounds and their quinone-modified analogues in NADH dehydrogenase-catalysed oxygen radical formation.

Anthraquinone compounds belong to the most important class of clinical antitumour agents. However, their use is limited by their peroxidating activity, being the consequence of free radical formation initiated by three oxyreductases. This activity is considered to be the main cause of cardiotoxic effects. The affinity of anthraquinone compounds to these enzymes is an essential factor governing the rate of one-electron transfer and the generation of oxygen radicals. A series of novel derivatives and analogues of natural and synthetic anthraquinones has been examined with the aim of identifying the structural factors essential for the ability to stimulate oxygen radical formation catalysed by NADH dehydrogenase. Functional groups and moieties favouring or disfavouring the interaction of the compounds with the enzyme have been determined. The quinonoid moiety as well as at least two phenolic groups in peri positions favoured the affinity of these compounds for NADH dehydrogenase. The modification of the quinonoid structure to iminoquinonoid or carboquinonoid forms dramatically decreased interaction with the enzyme. The O'-substitution by a bulky group in the sugar moiety of daunorubicin decreased the ability of the derivatives to stimulate oxygen radical formation. It has also been shown that the presence of an ionizable amino group on the sugar moiety of daunorubicin favours interaction with the NADH dehydrogenase. However, its location is not essential for this effect.