Elena Cini

Design and synthesis of cyclopeptide analogues of the potent histone deacetylase inhibitor FR235222.

Various structurally modified analogues of FR235222 (1), a natural tetrapeptide inhibitor of mammalian histone deacetylases, were prepared in a convergent approach. The design of the compounds was aimed to investigate the effect of structural modifications of the tetrapeptide core involved in enzyme binding in order to overcome some synthetic difficulties connected with the natural product 1. The modifications introduced could also help identify key structural features involved in the mechanism of action of these compounds. The prepared molecules were subjected to in vitro pharmacological tests, and their potency was tested on cultured cells. Two of the components of the array were found to be more potent than the parent compound 1 and almost as efficient as trichostatin A (TSA). These results demonstrate that it is possible to synthesize highly active cyclic tetrapeptides using commercially available amino acids (with the exception of 2‐amino‐8‐oxodecanoic acid, Ahoda). The nature of the residue in the second position of the cyclic peptide and the stereochemistry of the Ahoda tail are important for the inhibitory activity of this class of cyclic tetrapeptide analogues.

Lactam based 7-amino suberoylamide hydroxamic acids as potent HDAC inhibitors

A series of SAHA-like molecules were prepared introducing different lactam-carboxyamides in position 7 of the suberoylanilide skeleton. The activity against different HDAC isoforms was tested and the data compared with the corresponding linear products, without substituent in position 7. In general, this modification provided an effective reinforcement of in vitro activity. While the lactam size or the CO/NH group orientation did not strongly influence the inhibition, the contemporary modification of the suberoylamide fragment gave vary active variants in the lactam series, with compound 28 (ST8078AA1) that showed IC50 values between 2 and 10nM against all Class I HDAC isoforms, demonstrating it to be a large spectrum pan-inhibitor. This strong affinity with HDAC was also confirmed by the value of IC50=0.5μM against H460 cells, ranking 28 as one of the most potent HDAC inhibitors described so far.

Downregulation of Hypoxia-related Responses by Novel Antitumor Histone Deacetylase Inhibitors in MDAMB231 Breast Cancer Cells

The tumor microenvironment is characterized by a poor circulation which results in the selection of neoplastic cells that can grow or survive under hypoxic conditions. The relationship between hypoxia and histone deacetylase (HDAC) inhibitors has been previously established. In this work we evaluated the effects of novel HDAC inhibitors (the natural peptide FR235222 and three tetrapeptide analogs) in the human breast cancer cell line MDAMB231, cultured under hypoxia (2% O2 ≉ 14 mmHg) or normoxia (20% O2 ≉ 140 mmHg). First, we found that the novel HDAC inhibitors reduced cell proliferation in MDAMB231 cells at an extent which was similar or even higher than that exerted by the classic HDAC inhibitors trichostatin-A and suberoylanilide hydroxamic acid. More interestingly, the antiproliferative effects of the novel HDAC inhibitors were, in general, significantly higher in hypoxic cells than in normoxic controls. Hypoxic MDAMB231 cells expressed high levels of the hypoxia-inducible factor (HIF)-1α and HIF-1α-related genes, such as vascular endothelial growth factor, Bcl-2/E1B 19 kDa interacting protein-3, glucose transporter-1, carbonic anhydrase IX, as determined by Western blot analysis and qRT-PCR. Finally, we found that HIF-1α and HIF-1α-related genes were significantly downregulated by FR235222 and analogs. In conclusion, the identification of novel effects exerted by the HDAC inhibitors, characterized by a strong efficacy in inhibiting the expression of HIF-1α and its related genes, may have important implications in the pharmacological control of several tumors, including breast cancer, characterized by the presence of hypoxia, angiogenesis and metabolic derangements.

Investigating the Neuroprotective Effects of Turmeric Extract: Structural Interactions of β-Amyloid Peptide with Single Curcuminoids

A broad biophysical analysis was performed to investigate the molecular basis of the neuroprotective action of Curcuma longa extracts in Alzheimer’s disease. By combining circular dichroism and electron paramagnetic resonance experiments with molecular modeling calculations, the minor components of Curcuma longa extracts, such as demethoxycurcumin (2, DMC), bisdemethoxycurcumin (3, BDMC) and cyclocurcumin (4, CYC), were analyzed in a membrane environment mimicking the phospholipid bilayer. Our study provides the first evidence on the relative role of single curcuminoids interacting with Aβ-peptide. When the CYC and curcumin metabolite tetrahydrocurcumin (5, THC) were inserted into an anionic lipid solution, a significant modification of the Aβ CD curves was detected. These data were implemented by EPR experiments, demonstrating that CYC reaches the inner part of the bilayer, while the other curcuminoids are localized close to the membrane interface. Computational studies provided a model for the curcuminoid-Aβ interaction, highlighting the importance of a constrained “semi-folded” conformation to interact with Aβ analogously to the pattern observed in α-helical coiled-coil peptide structures. This combined approach led to a better understanding of the intriguing in vitro and in vivo activity of curcuminoids as anti-Alzheimer agents, paving a new path for the rational design of optimized druggable analogues.

Ruthenium-catalysed C-alkylation of 1,3-dicarbonyl compounds with primary alcohols and synthesis of 3-keto-quinolines

The mono-alkylation of 1,3-diketones using alcohols is possible in the presence of catalytic amounts of Ru(CO)(PPh3)3HCl and 10% mol of the Hantzsch ester. The borrowing hydrogen process between the catalyst and the dihydropyridine/pyridine couple prevents the common double alkylation of the Knoevenagel adduct without the need of stoichiometric reducing agents or sacrificial nucleophiles. The reaction was applied to the synthesis of a lactone intermediate for the preparation of the anti-obesity drug orlistat. Moreover, under the same Ru catalysis, a Friedlander reaction occurred with o-amino benzyl alcohols giving access to different 3-keto-substituted quinolines.

Benzodiazepine Scaffold as Drug-like Molecular Simplification of FR235222: A Chemical Tool for Exploring HDAC Inhibition

BACKGROUND Synthesis, computational study and biological evaluation of peptidomimetic analogues of FR235222 (3), a natural immunosuppressant and HDAC inhibitor, have been reported. These new compounds, bearing α-hydroxyketone moiety, as more stable zinc binding group (ZBG), were evaluated in vitro as HDAC inhibitors against the human HDACs isoforms 1-9 and in cellular antiproliferative assays on U937 human leukemia cell line. The 1,4-benzodiazepin-2,5-dione (BDZ), capping group and the natural ZBG, (S,R)-2-amino-9-hydroxy-8-oxodecanoic acid (Ahoda), were evaluated in order to probe HDAC inhibition and/or paralogue selectivity. Some of the new derivatives showed an interesting activity against a number of HDAC isozymes. The observed activity profile was rationalized by a computational assisted SAR study, in order to understand how the BDZ classes interact with the enzyme into the catalytic pocket. Despite its poor solubility, compound 17b showed significant antiproliferative profile and HDAC inhibition activity. RESULT In order to assess how the solubility issue could have affected the biological outcome, bioassay conditions were reproduced and quantification of precipitated particulate material was evaluated by turbidimetric and NMR studies together with physicochemical descriptors prediction. Thus, BDZ 17b has been chosen to be promising lead compounds for further optimization, in order to elucidate molecule- enzyme surface recognition.

A highly stereo-controlled protocol to prepare pipecolic acids based on Heck and cyclohydrocarbonylation reactions

A novel synthetic protocol based on an indium-mediated glyoxylate allylation, Heck coupling and Rh-catalysed cyclohydrocarbonylation (CHC) was established to access enantiomerically pure polysubstituted pipecolic acids. The key steps are the Heck reaction, which is performed exclusively using a phenone-oxime derived palladacycle and the domino hydroformylation-cyclisation of a styryl derivative obtained from the Heck coupling. The reaction scheme, proceeding with good stereoselectivity, is also suitable for the preparation of substituted piperidine derivatives.