Valerio Tazzari

Pharmacological profile of a novel H2S-releasing aspirin

The pharmacological profile of a new, safe, and effective hydrogen sulfide (H(2)S)-releasing derivative of aspirin (ACS14) is described. We report the synthesis of ACS14, and of its deacetylated metabolite (ACS21), the preliminary pharmacokinetics, and its in vivo metabolism, with the H(2)S plasma levels after intravenous administration in the rat. ACS14 maintains the thromboxane-suppressing activity of the parent compound, but seems to spare the gastric mucosa, by affecting redox imbalance through increased H(2)S/glutathione formation, heme oxygenase-1 promoter activity, and isoprostane suppression.

The hydrogen sulphide-releasing derivative of diclofenac protects against ischaemia–reperfusion injury in the isolated rabbit heart

Hydrogen sulphide (H2S) is an endogenous gaseous mediator active in the multilevel regulation of pathophysiological functions in mammalian cardiovascular tissues.

Effect of hydrogen sulphide-donating sildenafil (ACS6) on erectile function and oxidative stress in rabbit isolated corpus cavernosum and in hypertensive rats

To study the effect of the H2S‐donating derivative of sildenafil (ACS6) compared to sildenafil citrate and sodium hydrosulphide (NaHS) on relaxation, superoxide formation and NADPH oxidase and type 5 phosphodiesterase (PDE5) expression in isolated rabbit cavernosal tissue and smooth muscle cells (CSMCs), and in vivo on indices of oxidative stress induced with buthionine sulphoximine (BSO).

Effects of hydrogen sulfide-releasing L-DOPA derivatives on glial activation: potential for treating Parkinson disease

The main lesion in Parkinson disease (PD) is loss of substantia nigra dopaminergic neurons. Levodopa (l-DOPA) is the most widely used therapy, but it does not arrest disease progression. Some possible contributing factors to the continuing neuronal loss are oxidative stress, including oxidation of l-DOPA, and neurotoxins generated by locally activated microglia and astrocytes. A possible method of reducing these factors is to produce l-DOPA hybrid compounds that have antioxidant and antiinflammatory properties. Here we demonstrate the properties of four such l-DOPA hybrids based on coupling l-DOPA to four different hydrogen sulfide-donating compounds. The donors themselves were shown to be capable of conversion by isolated mitochondria to H2S or equivalent SH− ions. This capability was confirmed by in vivo results, showing a large increase in intracerebral dopamine and glutathione after iv administration in rats. When human microglia, astrocytes, and SH-SY5Y neuroblastoma cells were treated with these donating agents, they all accumulated H2S intracellularly as did their derivatives coupled to l-DOPA. The donating agents and the l-DOPA hybrids reduced the release of tumor necrosis factor-α, interleukin-6, and nitric oxide from stimulated microglia, astrocytes as well as the THP-1 and U373 cell lines. They also demonstrated a neuroprotective effect by reducing the toxicity of supernatants from these stimulated cells to SH-SY5Y cells. l-DOPA itself was without effect in any of these assays. The H2S-releasing l-DOPA hybrid molecules also inhibited MAO B activity. They may be useful for the treatment of PD because of their significant antiinflammatory, antioxidant, and neuroprotective properties.

Activity of a new hydrogen sulfide-releasing aspirin (ACS14) on pathological cardiovascular alterations induced by glutathione depletion in rats.

We investigated the effects of the hydrogen sulfide (H₂S)-releasing derivatives of aspirin (ACS14) and salicylic acid (ACS21) in a rat model of metabolic syndrome induced by glutathione (GSH) depletion, causing hypertension and other pathological cardiovascular alterations. GSH depletion was induced in normal rats by the GSH-synthase inhibitor buthionine sulfoximine (BSO, 30 mmol/L day for seven days in the drinking water). Systolic blood pressure and heart rate were measured daily by the tail-cuff method, and plasma thromboxane B₂, 6-keto-prostaglandin F(2α), 8-isoprostane, GSH, insulin and glucose were determined at the end of the seven-day BSO schedule. In addition, ischemia/reperfusion-induced myocardial dysfunction and endothelial dysfunction were assayed on isolated heart and aortic rings, respectively. Unlike aspirin and salicylic acid, ACS14 and ACS21 reduced BSO-induced hypertension, also lowering plasma levels of thromboxane B₂, 8-isoprostane and insulin, while GSH remained in the control range. Neither ACS14 nor ACS21 caused gastric lesions. Both restored the endothelial dysfunction observed in aortic rings from BSO-treated rats, and in ischemia/reperfusion experiments they lowered left ventricular end-diastolic pressure, consequently improving the developed pressure and the maximum rise and fall of left ventricular pressure. Together with this improvement of heart mechanics there were reductions in the activity of creatine kinase and lactate dehydrogenase in the cardiac perfusate. This implies that H₂S released by both ACS14 and ACS21 was involved in protecting the heart from ischemia/reperfusion, and significantly limited vascular endothelial dysfunction in aortic tissue and the related hypertension.

Therapeutic Effect of Hydrogen Sulfide-Releasing L-Dopa Derivative ACS84 on 6-OHDA-Induced Parkinson’s Disease Rat Model

Parkinson’s disease (PD), characterized by loss of dopaminergic neurons in the substantia nigra, is a neurodegenerative disorder of central nervous system. The present study was designed to investigate the therapeutic effect of ACS84, a hydrogen sulfide-releasing-L-Dopa derivative compound, in a 6-hydroxydopamine (6-OHDA)-induced PD model. ACS84 protected the SH-SY5Y cells against 6-OHDA-induced cell injury and oxidative stress. The protective effect resulted from stimulation of Nrf-2 nuclear translocation and promotion of anti-oxidant enzymes expression. In the 6-OHDA-induced PD rat model, intragastric administration of ACS84 relieved the movement dysfunction of the model animals. Immunofluorescence staining and High-performance liquid chromatography analysis showed that ACS84 alleviated the loss of tyrosine-hydroxylase positive neurons in the substantia nigra and the declined dopamine concentration in the injured striatums of the 6-OHDA-induced PD model. Moreover, ACS84 reversed the elevated malondialdehyde level and the decreased glutathione level in vivo. In conclusion, ACS84 may prevent neurodegeneration via the anti-oxidative mechanism and has potential therapeutic values for Parkinson’s disease.

New sulfurated derivatives of valproic acid with enhanced histone deacetylase inhibitory activity.

One dithiolthione and two new methanethiosulfonate derivatives of valproic acid (VPA) were synthesized and tested in vitro as histone deacetylase (HDAC) inhibitors. The new molecules, as well as their sulfurated moieties, exhibited a much stronger inhibition of HDAC enzymatic and antiproliferative activities and histone hyperacetylation than VPA. ACS 2 is the most interesting compound among the new VPA derivatives and its sulfurated moiety, 5-(4-hydroxyphenyl)-3H-1,2-dithiole-3-thione, also known to be a metabolite of anethole trithione, seems to contribute significantly to its activity. This is the first time that HDAC inhibitory activity is described for dithiolethiones and thiosulfonates.

Dithiolethiones Inhibit NF-κB Activity via Covalent Modification in Human Estrogen Receptor–Negative Breast Cancer

The NF-κB transcription factor family influences breast cancer outcomes by regulating genes involved in tumor progression, angiogenesis, and metastasis. Dithiolethiones, a class of naturally occurring compounds with cancer chemoprevention effects that have become clinically available, have been found to inhibit NF-κB activity. However, the mechanism of this inhibition has not been identified, and the influence of dithiolethines on NF-κB pathway in breast cancer cells has not been examined. Here, we investigated the chemical and biochemical effects of dithiolethione on NF-κB and downstream effector molecules in estrogen receptor-negative breast cancer cells and murine tumor xenografts. The dithiolethiones ACS-1 and ACS-2 inhibited NF-κB transcriptional activity. Interestingly, this inhibition was not due to H(2)S release or protein phosphatase 2A activation, which are key properties of dithiolethiones, but occurred via a covalent reaction with the NF-κB p50 and p65 subunits to inhibit DNA binding. Dithiolethione-mediated inhibition of NF-κB-regulated genes resulted in the inhibition of interleukin (IL)-6, IL-8, urokinase-type plasminogen activator, and VEGF production. ACS-1 also inhibited matrix metalloproteinase-9 activity, cellular migration, and invasion, and ACS-2 reduced tumor burden and resulted in increased tumor host interactions. Together, our findings suggest that dithiolethiones show potential clinical use for estrogen negative breast cancer as a chemotherapeutic or adjuvant therapy.

New aryldithiolethione derivatives as potent histone deacetylase inhibitors.

A series of dithiolethione derivatives was synthesized and the in vitro HDAC inhibitory activity was tested. The most active compounds, 1 and 2, exhibited an IC(50) in nM range with a strong hyperacetylation of histone H4 in A549 cells. The HDAC inhibitory activity comparable to that of SAHA and the inhibition of A549 cell proliferation suggest that these compounds are worthy of further studies as potential anticancer agents.

HPLC determination of novel dithiolethione containing drugs and its application for in vivo studies in rats

A panel of new drugs obtained by grafting a sulfurated moiety, i.e. 5-(4-hydroxyphenyl)-3H-1,2-dithiole-3-thione (ADTOH) onto existing drugs have been synthesized and their in vivo action is under preclinical evaluation. In the present paper we describe rapid HPLC methods to detect ADTOH derivatives of valproic acid (ACS2), sildenafil (ACS6), aspirin (ACS14) and diclofenac (ACS15) in plasma. These methods allow the simultaneous detection of the potential drugs and of ADTOH moiety. In the case of ACS14 the de-acetylated metabolite (ACS21) can also be concomitantly measured. The chromatographic separation was performed on a C18 column, applying a mobile phase consisting of a mixture of trifluoroacetic acid and acetonitrile. ADTOH, ACS6, ACS14, ACS21 were separated isocratically whereas ACS2 and ACS15 were separated applying gradient elution. The methods are precise and accurate, with a low quantification limit of 200 nM for ACS2, ACS15 and ACS21 or 100 nM for ADTOH, ACS6 and ACS14. The mean absolute recovery for all tested molecules was always found to be close to 100%. The methods are shown to be selective and linear in the range 0.2-50 microM and thus appear suitable for pharmacokinetic studies with ADTOH containing compounds, as indicated by exemplificative experiments performed with intravenous administration of the drugs to rats.