Sabrina Dallavalle

Development and therapeutic impact of HDAC6-selective inhibitors

Histone deacetylases (HDAC) play a key role in regulating gene expression by deacetylating histones. Some HDAC isoforms can also modulate the function of nonhistone proteins implicated in regulatory processes, and therefore HDACs are recognized as useful targets for therapeutic purposes. HDAC inhibitors have generated substantial interest as antitumor agents, because they induce various cellular effects, including apoptosis, cell cycle arrest and inhibition of angiogenesis. The nature of cellular response likely depends on the biological context and on the pattern of HDAC isoform inhibition. Various HDAC inhibitors belonging to different structural classes have been developed. Many inhibitors are characterized by a pan-HDAC inhibitory profile. The potential advantages of isoform-selective inhibitors over pan-HDAC inhibitors in terms of efficacy or toxicity remain to be defined. The emerging interest for HDAC6-selective inhibitors is related to the modulation of acetylation of nonhistone regulatory proteins implicated in cancer-relevant processes, including cell migration, metastasis, angiogenesis and stress-response pathways. This review is focused on the recent development of HDAC inhibitors, with particular reference to HDAC6-selective inhibitors, and the efforts and perspectives in optimization of their therapeutic applications.

Current Status and Perspectives in the Development of Camptothecins

Camptothecins are cytotoxic agents with a wide spectrum of antitumor activity. The unique mechanism of action, the impressive preclinical efficacy and the clinical success of irinotecan and topotecan have stimulated intensive efforts to identify novel analogues. The development of novel camptothecins was recently rationalized on the basis of the detailed knowledge of mechanism of drug-target interaction and was aimed to overcome the major limitations of these drugs (i.e. lactone ring instability and reversibility of topoisomerase I-DNA cleavage complexes). The development of novel series of analogues (7-substituted camptothecins, silatecans and homocamptothecins) resulted in identification of promising compounds, which are currently in clinical development. Considering the lack of precise correlations between preclinical activity and clinical efficacy of camptothecins, the potential advantages of novel analogs in clinical therapy remains to be documented. However, a rational basis for drug selection and development is now provided by the recognition of major limitations of these agents and by a detailed knowledge of multiple interactions between drug, cellular target and serum albumin. Inhibition of the nuclear enzyme DNA topoisomerase I has proven to be a promising strategy in the design of antitumor agents, in spite of a limited cellular basis of selectivity in cytotoxic action of camptothecins (i.e., overexpression of the target enzyme in tumor cells, and increased sensitivity of proliferating cells). The interest in topoisomerase I as a therapeutic target promoted various efforts to identify other chemotypes effective as topoisomerase inhibitors and chemical/modelling efforts to rationally design specific analogs among known inhibitors. Additional approaches, including drug delivery/formulation, optimization of dose/schedule and route of administration, are expected to improve the therapy with camptothecins and other inhibitors.

Design, synthesis, and evaluation of biphenyl-4-yl-acrylohydroxamic acid derivatives as histone deacetylase (HDAC) inhibitors

A series of hydroxamic acid-based histone deacetylase (HDAC) inhibitors were designed on the basis of a model of the HDAC2 binding site and synthesized. They are characterized by a cinnamic spacer, capped with a substituted phenyl group. Modifications of the spacer are also reported. In an in vitro assay with the isoenzyme HDAC2, a good correlation of the activity with the docking energy was found. In human ovarian carcinoma IGROV-1 cells, selected compounds produced significant acetylation of p53 and alpha-tubulin. Most compounds showed an antiproliferative activity comparable to that of SAHA. At equitoxic concentrations, the tested compounds were more effective than SAHA in inducing apoptotic cell death. Compounds selected for in vivo evaluation exhibited a significant antitumor activity on three tumor models at well tolerated doses, thus suggesting a good therapeutic index.

Novel tumor-targeted RGD peptide-camptothecin conjugates: synthesis and biological evaluation.

Five RGD peptide-camptothecin (CPT) conjugates were designed and synthesized with the purpose to improve the therapeutic index of this antitumoral drug family. New RGD cyclopeptides were selected on the basis of their high affinity to alpha(v) integrin receptors overexpressed by tumor cells and their metabolic stability. The conjugates can be divided in two groups: in the first the peptide was attached to the drug through an amide bond, in the second through a hydrazone bond. The main difference between the two spacers lies in their acid stability. Affinity to the receptors was maintained for all conjugates and their internalization into tumor cells was demonstrated. The first group conjugates showed lower in vitro and in vivo activity than the parent drug, probably due to the excessive stability of the amide bond, even inside the tumor cells. Conversely, the hydrazone conjugates exhibited in vitro tumor cell inhibition similar to the parent drug, indicating high conversion in the culture medium and/or inside the cells, but their poor solubility hampered in vivo experiments. On the basis of these results, information was acquired for additional development of derivatives with different linkers and better solubility for in vivo evaluation.

Enol Carbamates as Inhibitors of Fatty Acid Amide Hydrolase (FAAH) Endowed with High Selectivity for FAAH over the Other Targets of the Endocannabinoid System

Enol Carbamates as Inhibitors of Fatty Acid Amide Hydrolase (FAAH) Endowed with High Selectivity for FAAH over the Other Targets of the Endocannabinoid System Sonia Gattinoni, Chiara De Simone, c] Sabrina Dallavalle, Filomena Fezza, c] Raffaella Nannei, Daniele Amadio, c] Patrizia Minetti, Gianandrea Quattrociocchi, Antonio Caprioli, Franco Borsini, Walter Cabri, Sergio Penco, Lucio Merlini,* and Mauro Maccarrone*

A new synthesis of the cytotoxic alkaloid Luotonine A

A convenient synthesis of the cytotoxic alkaloid Luotonine A has been achieved using cheap and readily accessible reagents. The key intermediate in the synthesis is the tricyclic compound 2,3-dihydro[1H]-pyrrolo[3,4-b]quinolin-3-one (5).

Antitumor activity of the retinoid-related molecules (E)-3-(4'-hydroxy-3'-adamantylbiphenyl-4-yl)acrylic acid (ST1926) and 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) in F9 teratocarcinoma : Role of retinoic acid receptor gamma and retinoid-independent pathways

The retinoid-related molecules (RRMs) ST1926 [(E)-3-(4′-hydroxy-3′-adamantylbiphenyl-4-yl)acrylic acid] and CD437 (6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid) are promising anticancer agents. We compared the retinoic acid receptor (RAR) trans-activating properties of the two RRMs and all-trans-retinoic acid (ATRA). ST1926 and CD437 are better RARγ agonists than ATRA. We used three teratocarcinoma cell lines to evaluate the significance of RARγ in the activity of RRMs: F9-wild type (WT); F9γ-/-, lacking the RARγ gene; F9γ51, aF9γ-/-derivative, complemented for the RARγ deficit. Similar to ATRA, ST1926 and CD437 activate cytodifferentiation only in F9-WT cells. Unlike ATRA, ST1926 and CD437 arrest cells in the G2/M phase of the cell cycle and induce apoptosis in all F9 cell lines. Our data indicate that RARγ and the classic retinoid pathway are not relevant for the antiproliferative and apoptotic activities of RRMs in vitro. Increases in cytosolic calcium are fundamental for apoptosis, in that intracellular calcium chelators abrogate the process. Comparison of the gene expression profiles associated with ST1926 and ATRA in F9-WT and F9γ-/-indicates that the RRM activates a conspicuous nonretinoid response in addition to the classic and RAR-dependent pathway. The pattern of genes regulated by ST1926 selectively, in a RARγ-independent manner, provides novel insights into the possible molecular determinants underlying the activity of RRMs in vitro. Furthermore, it suggests that RARγ-dependent responses are relevant to the activity of RRMs in vivo. Indeed, the receptor hinders the antitumor activity in vivo, in that both syngeneic and immunosuppressed SCID mice bearing F9γ-/- tumors have increased life spans after treatment with ST1926 and CD437 relative to their F9-WT counterparts.

Preclinical profile of antitumor activity of a novel hydrophilic camptothecin, ST1968

ST1968 is a novel hydrophilic camptothecin (CPT) derivative of the 7-oxyiminomethyl series. Because ST1968 retained ability to form remarkably stable cleavable complexes, this study was done to investigate its preclinical profile of antitumor activity in a large panel of human tumor models, including irinotecan-resistant tumors. Although less potent than SN38 in vitro, i.v. administered ST1968 caused a marked tumor inhibition, superior to that of irinotecan, in most tested models. ST1968 exhibited an impressive activity against several tumors including models of ovarian and colon carcinoma in which a high rate of cures was observed. In the most responsive tumors, complete and persistent tumor regressions were achieved even with low suboptimal doses. Even tumors derived from intrinsically resistant cells exhibited a significant responsiveness. Histologic analysis of treated tumors supports a contribution of both proapoptotic and antiangiogenic effects to ST1968 antitumor efficacy. A study done in yeast cells transformed with CPT-resistant mutant forms of topoisomerase I documented that, in contrast to other tested CPT, ST1968 was active against yeasts expressing the mutant K720E enzyme. Based on its outstanding efficacy superior to that of irinotecan and of its good therapeutic index, ST1968 has been selected for clinical development. [Mol Cancer Ther 2008;7(7):2051–9]

Synthesis, modeling, and RET protein kinase inhibitory activity of 3- and 4-substituted β-carbolin-1-ones

A series of beta-carbolin-2-ones and 3,10-dihydro-2H-azepino[3,4-b]indol-1-ones have been designed, synthesized, and evaluated as RET protein kinase inhibitors on the basis of their structural similarity with the prototype indolin-2-one RPI-1. Some beta-carbolin-2-ones (structure 2) showed an ability to inhibit RET enzymatic activity in vitro and proliferation of RETC634R oncogene-transformed NIH3T3 cells comparable to that of the reference compound. The docking analysis of the interaction of these compounds with the crystallographic structure of RET tyrosine kinase domain suggested a new binding interaction scheme different from the one proposed during their design. The rigid structure of the compounds of this series represents a new scaffold with potential advantages in the design of RET protein kinase inhibitors.

Novel cytotoxic 7-iminomethyl and 7-aminomethyl derivatives of camptothecin

A series of new 7-iminomethyl derivatives of camptothecin were obtained from camptothecin-7-aldehyde and aromatic, alicyclic and aliphatic amines. Their hydrogenation led to the corresponding amines. All the imines and the less polar amines showed a marked increase of the cytotoxic activity against H460 non-small lung carcinoma cell line, with respect to topotecan. The lipophilicity of the substituent in position 7 of camptothecin seems to play an important role for cytotoxic potency. The 7-phenyliminomethyl derivative showed efficacy comparable to topotecan in vivo against NSCLC H460 xenografted in athymic nude mice.