Phloridzin derivatives inhibiting pro-inflammatory cytokine expression in human cystic fibrosis IB3-1 cells.

Cystic Fibrosis (CF) is the most diffuse autosomal recessive genetic disease affecting Caucasians. A persistent recruitment of neutrophils in the bronchi of CF patients contributes to exacerbate the airway tissue damage, suggesting that modulation of chemokine expression may be an important target for the patient's well being thus the identification of innovative anti-inflammatory drugs is considered a longterm goal to prevent progressive tissue deterioration. Phloridzin, isolated from Malus domestica by a selective molecular imprinting extraction, and its structural analogues, Phloridzin heptapropionate (F1) and Phloridzin tetrapropionate (F2), were initially investigated because of their ability to reduce IL-6 and IL-8 expression in human CF bronchial epithelial cells (IB3-1) stimulated with TNF-α. Release of these cytokines by CF cells was shown to be controlled by the Transcription Factor (TF) NF-kB. The results of the present investigation show that of all the derivatives tested, Phloridzin tetrapropionate (F2) is the most interesting and has greatest potential as it demonstrates inhibitory effects on the expression and production of different cytokines involved in CF inflammation processes, including RANTES, VEGF, GM-CSF, IL-12, G-CSF, MIP-1b, IL-17, IL-10 and IP-10, without any correlated anti-proliferative and pro-apoptotic effects.

Targeting transcription factor activity as a strategy to inhibit pro-inflammatory genes involved in cystic fibrosis: decoy oligonucleotides and low-molecular weight compounds.

The development of drugs able to inhibit the expression of pro-inflammatory genes is of great interest in the treatment of cystic fibrosis (CF). Chronic pulmonary inflammation in the lungs of patients affected by CF is characterized by massive intra-bronchial infiltrates of neutrophils. This process is initiated upon interaction of pathogens (including Pseudomonas aeruginosa) with surface bronchial cells. Consequently, they release cytokines, the most represented being the potent neutrophilic chemokine Interleukin (IL)-8 and the pro-inflammatory cytokine IL-6. The chronic inflammatory process is crucial, since it leads to progressive tissue damage and severe respiratory insufficiency. In order to reduce the adverse effects of the excessive inflammatory response, one of the approaches leading to inhibition of IL-8 and IL-6 gene expression is the transcription factor (TF) decoy approach, based on intracellular delivery of double stranded oligodeoxynucleotides (ODNs) mimicking the binding sites of TFs and causing inhibition of binding of TF-related proteins to regulatory sequences identified in the promoters of specific genes. Since the promoters of IL-8 and IL-6 contain consensus sequences for NF-κ B and Sp1, double stranded TF "decoy" ODNs targeting NF-κB and Sp1 can be used. Alternatively, screening of drugs targeting relevant TFs can be performed using drug cocktails constituted by extracts from medicinal plants inhibiting TF/DNA interactions. Finally, virtual screening might lead to identification of putative bioactive molecules to be validated using molecular and cellular approaches. By these means, low-molecular drugs targeting NF-κB and inhibiting IL-8 gene expression are available for pre-clinical testing using experimental systems recapitulating chronic pulmonary inflammation of patients affected by CF.

Bangladeshi medicinal plant extracts inhibiting molecular interactions between nuclear factors and target DNA sequences mimicking NF-kappaB binding sites.

Several medicinal plants can be employed to produce extracts exhibiting biological effects. The aim of this work was to verify the ability of extracts derived from different medicinal plants of Bangladesh in interfering with specific DNA-protein interactions. The rationale for this study is based on the observation that alteration of gene transcription represents a very promising approach to control the expression of selected genes and could be obtained using different molecules acting on the interactions between DNA and transcription factors (TFs). We have analysed the antiproliferative activity of extracts from the medicinal plants Hemidesmus indicus, Polyalthia longifolia, Aphanamixis polystachya, Moringa oleifera, Lagerstroemia speciosa, Paederia foetida, Cassia sophera, Hygrophila auriculata and Ocimum sanctum. Antiproliferative activity was assayed on different human cell lines, including erythroleukemia K562, B-lymphoid Raji, T-lymphoid Jurkat and erythroleukemia HEL cell lines. We employed the electrophoretic mobility shift assay (EMSA) as a suitable technique for the identification of plant extracts altering the binding between transcription factors and the specific DNA elements. We found that low concentrations of Hemidesmus indicus, Polyalthia longifolia, Moringa oleifera and Lagerstroemia speciosa, and very low concentrations of Aphanamixis polystachya extracts inhibit the interactions between nuclear factors and target DNA elements mimicking sequences recognized by the nuclear factor kappaB (NF-kappaB). On the contrary, high amount of extracts from Paederia foetida, Cassia sophera, Hygrophila auriculata or Ocimum sanctum were unable to inhibit NF-kappaB/DNA interactions. Extracts inhibiting both NF-kappaB binding activity and tumor cell growth might be a source for anti-tumor compounds, while extracts inhibiting NF-kappaB/DNA interactions with lower effects on cell growth, could be of interest in the search of compounds active in inflammatory diseases, for which inhibition of NF-kappaB binding activity without toxic effects should be obtained.

Peptide nucleic acids (PNA)-DNA chimeras targeting transcription factors as a tool to modify gene expression.

Peptide nucleic acids (PNAs)-DNA chimeras have been recently described as DNA mimics constituted of a part of PNA and of a part of DNA. We have demonstrated that double stranded molecules based on PNA-DNA chimeras bind to transcription factors in a sequence-dependent manner. Accordingly, these molecules can be used for transcription factor decoy (TFD) pharmacotherapy. Effects of double stranded PNA-DNA chimeras targeting NF-kappaB and Sp1 were determined on in vitro cultured human cells and were found to be comparable to those observed using double-stranded DNA decoys. The TFD molecules based on PNA-DNA chimeras can be further engineered by addition of short peptides facilitating cell penetration and nuclear localization. Therefore, these engineered molecules could be of great interest for in vivo experiments for non-viral gene therapy of a variety of diseases, including neoplastic and viral diseases, for which the TFD approach has been already demonstrated as a very useful strategy.

In vitro antiproliferative effects on human tumor cell lines of extracts from the Bangladeshi medicinal plant Aegle marmelos Correa.

In the present paper we show that extracts from Aegle marmelos Correa are able to inhibit the in vitro proliferation of human tumor cell lines, including the leukemic K562, T-lymphoid Jurkat, B-lymphoid Raji, erythroleukemic HEL, melanoma Colo38, and breast cancer MCF7 and MDA-MB-231 cell lines. Molecules present within the studied Aegle marmelos C. extracts were identified by gas-chromatography/mass-spectrometry analysis; three derivatives (butyl p-tolyl sulfide, 6-methyl-4-chromanone and butylated hydroxyanisole) were found to exhibit strong activity in inhibiting in vitro cell growth of human K562 cells. The antiproliferative activity of these compounds was found to be comparable to that of known antitumor agents, including cisplatin, chromomycin, cytosine arabinoside and 5-fluorouracil. In addition, the antiproliferative activity of butyl-p-tolyl sulfide, 6-methyl-4-chromanone and 5-methoxypsolaren was associated to activation of the differentiation pattern of K562 cells.

Synthesis and biological effects of a new series of 2-amino-3-benzoylthiophenes as allosteric enhancers of A1-adenosine receptor.

New derivatives of PD 81,723, an allosteric enhancer of agonist binding to the A1-adenosine receptor, have been synthesized and evaluated in an intact cell assay. Compounds 3a, 3o and 3p appeared to be more potent than PD 81,723 and at a concentration of 0.1 microM caused significant reductions of cAMP content of CHO cells expressing the human A1-adenosine receptor. Compounds 4e and 4o appeared to be allosteric enhancers at a low concentration and antagonists at a higher concentration, whereas compounds 3c, 3g, 3s and 4l appeared to be weak antagonists that are also allosteric enhancers at the higher concentration of 10 microM.

Design, synthesis and binding at cloned muscarinic receptors of N-[5-(1'-substituted-acetoxymethyl)-3-oxadiazolyl] and N-[4-(1'-substituted-acetoxymethyl)-2-dioxolanyl] dialkyl amines.

Few muscarinic antagonists differentiate between the M4 and M2 muscarinic receptors. In a structure activity study, aimed at discovering leads for the development of a M4 muscarinic receptor-selective antagonist, we have synthesized and tested at cloned muscarinic receptors the binding of a group of dioxolane- or oxadiazole-dialkyl amines, and compared them to our compound 1, which contains the furan nucleus. Although none of these agents were particularly potent at M4 receptors (Kd values were typically 30-70 nM), furan derivatives (-)1 and (+)1 were significantly more potent at M4 receptors than at M2 receptors (approximately 3- and 4-fold, respectively). The dioxolane derivatives 12b and 12c were more than 10-fold selective for the M4 versus the M2 receptors, while the dioxolane derivative 12e was 15-fold more potent at M4 receptors than for M2 receptors. However, these agents bound to M3 receptors with potencies like that for the M4 receptor, so they are not M4-selective. The M4/M2 relative selectivities of some of our compounds are similar to the better hexahydrosiladifenidol derivatives, and may provide some important structural clues for the development of potent and selective M4 antagonists.

3,6-Disubstituted 1,2,4-triazolo[3,4-b][1,3,4]thiadiazoles: synthesis and evaluation for antimicrobial and antiviral activity. III.

A series of novel 3,6-disubstituted 1,2,4-triazolo[3,4-b][1,3,4]thiadiazole derivatives was prepared and tested to evaluate their antimycotic, antibacterial and anti-HIV-1 activities. The reaction of thiocarbohydrazide with carboxylic acids at the melting temperature allows an improved preparation of the 5-substituted 4-amino-3-mercapto-1,2,4-triazole heterocycles which in turn allows an easier preparation of the 1,2,4-triazolo[3,4-b] [1,3,4]thiadiazole ring system. All tested compounds didn't show any significant activity.