Search

Sintesi di due nuovi composti a struttura tetrazepinonica, analoghi della Temozolomide. I due nuovi composti hanno mostrato una interessante attività antiproliferativa nei confronti di linee cellulari tumorali esprimenti l'enzima MGMT

Epigenetic therapy is a new promising area in cancer research that is based on the use of a series of molecules capable of affecting tumor cell growth, differentiation and death by modifying the cellular mechanisms underlying the control of gene expression. Significant enhancement of traditional anticancer drug effects has been also reported by several authors. Our recent research focused on the identification of new epigenetic agent-containing drug combinations to be employed in the therapy of leukemia. The results showed that the new combination of an histone deacetylase (HDAC) inhibitor and the ribonucleotide reductase (RR) inhibitor 3’-methyl-adenosine (3’-Me-Ado) is endowed with a significant antitumor activity against promyelocytic leukemia cells. This effect was dependent not only by the HDAC inhibitors-mediated enhancement of drug-induced apoptosis, but also by the ability of these pleiotropic agents to inhibit the activity of the RR enzyme. Studies are ongoing to verify the feasibility of creating new bifunctional agents containing both an RR and an HDAC inhibitor within the same molecule, so that they can reproduce the effects of this combination and furnish an easier and effective therapeutic modality in view of a possible future use in the clinics.

Indazole nucleus represents a very attractive scaffol to obtain new molecole endowed with antineoplastic activity. On the basis of these literature data we have designed some indazole derivatives such as N-indazolylbenzamides and N-indazolyl-N’-phenylureas as potential CDK1 inhibitors. In fact the above compounds contain the structural feature, common to the majority of CDK inhibitors, requested to make hydrogen bonds with the molecular forks present in the hinge region of CDKs. The N-indazolylbenzamides 1 were obtained by reacting aminoindazoles and substituted benzoylchlorides. Among the synthesized compounds some derivatives 1 resulted to be CDK1 inhibitors showing IC50 values in the range 4.4-10.0 M

Combination therapy is the mainstay of anticancer therapy due to the significant synergistic effects achievable. Now that anticancer drug research turned toward a more molecular targeted approach, the design of dual-target drugs appears to be a new promising strategy with the potential to improve the therapeutic efficacy of the single drug and to reduce the probability of drug induced resistance and cross resistance. In our previous work, we found that 3’-C-methyl-adenosine (3’-Me-Ado), developed by us as a potent ribonucleotide reductase (RR) inhibitor with antitumor activity against both human leukemia and carcinoma cell lines, elicited significant growth inhibitory and apoptotic synergistic effects in promyelocytic leukemia cells in combination with some hydroxamic acid–derived histone deacetylase (HDAC) inhibitors. Also the antiepileptic agent valproic acid (VPA) was shown to inhibit HDAC in tumors and to synergize with a number of anticancer drugs including RR inhibitors. Since VPA is currently used and well tolerated in the clinics, we pursued the development of co-drugs by combining the structures of 3’-Me-Ado and VPA in a single molecule. Thus, the 3’-C-methyladenosine-2’(5’)-O-valproic esters (A157, A167) and the 2’,5’-diester analog (A160) were synthesized and evaluated for their antitumor potential in HL60 and HuT78 cells. Cytotoxicity was studied by the Trypan blue dye exclusion test, whereas apoptosis was evaluated by fluorescence microscopy after staining with acridine orange and etidium bromide. Preliminary results showed that the compounds induced cytotoxic effects with IC50 ranging from 11 and 600 μM and from 32 and 900 μM in HL60 and HuT78 respectively, demonstrating a higher potency when compared with VPA alone (IC50 of about 2.5 mM). However, their apoptotic activity was modest, with the most potent compound A160 showing an AC50 higher than 100 μM in HL60 cells. The western blotting study of the level of acetylated histone H3 demonstrated that the compounds were not capable of inhibiting significantly the HDAC enzymes. Instead, the evaluation of cell cycle distribution by flow cytometry did show a recruitment of treated cells in the G1 phase of the cell cycle, suggesting that the compounds could retain the RR inhibiting activity of 3’-Me-Ado. Further studies are in progress to clarify the mechanism of cytotoxic activity of the synthesized co-drugs.

Monoclonal antibodies (mAbs) constitute the most rapidly growing class of human therapeutics and the second largest class of drugs after vaccines. The treatment of B-cell malignancies and HER2/Neu(+) breast cancer has benefited considerably from the use of therapeutic mAbs, either alone or in combination with standard chemotherapy. Frequent relapses, however, demonstrate that the bioactivity of these mAbs is still suboptimal. The concept of improving the anti-tumor activity of mAbs is well established and potentiating the cytotoxicity induced by anticancer mAbs can be achieved by strategies that target the downstream cytolytic effector cells. The recruitment of Fcγ receptor-dependent functions appears well suited in this regard, because several lines of evidence suggest that enhancing antibody-dependent cellular cytotoxicity (ADCC) induced by therapeutic mAbs may directly improve their clinical efficacy. The cytolytic effector cells involved in ADCC are FcγR-expressing natural killer (NK) cells, but also γδ T cells can be amplified and finetuned for stronger ADCC activity. γδ T cells are raising a considerable interest in the immunotherapy community given their intrinsic antitumor activity that can be boosted by stimulation with synthetic phosphoantigens (PAgs), or with drugs that cause their accumulation into target cells, like aminobisphosphonates (N-BPs), and low doses interleukin (IL)-2. The field is interesting, and several papers have already explored this approach in solid and haematological malignancies. Thus, we propose that enhancing the efficacy of mAbs by combination with γδ T cell activation may have considerable therapeutic potential for a variety of malignancies, most especially for patients whose FcγR alleles impair ADCC.

Recently our research group has reported the synthesis of some 3-amino-N-phenyl-1H-indazole-1-carboxamides able to inhibit at low micromolar concentrations the cell growth of many neoplastic cell lines. The above compounds are unsubstituted in the indazole nucleus and this gives the hope to obtain more active compounds if appropriate substituents are beared by the above nucleus. So, several new N-phenyl-1H-indazole-1-carboxamides 1c-h and 4l,m were prepared by reacting phenyl isocyanates 3a,b with 3-amino-1H-indazoles 2c,e,g, or 1H-indazole 2l respectively. Chemical trasformations of compounds 1a,b and 1g,h gave 3-acetamido-N-phenyl-1H-indazole-1-carboxamides 5a,b, and 3,5-diamino-N-phenyl-1H-indazole-1-carboxamides 4i,j respectively. Finally, 3,5-diacetamido-N-phenyl-1H-indazole-1-carboxamides 6a,b were prepared by acethylation of 4i,j. Some of synthesized compounds were evaluated for their in vitro antiproliferative activity against 60 human tumoral cell lines derived from nine clinically isolated cancer types (leukemia, non-small cell lung, colon, CNS, melanoma, ovarian, renal, prostate and breast). Compound 1c, the most active of the series, was able to inhibit cell growth showing GI50 values in the 0.041-33.6 μM range, being very effective against colon and melanoma cell lines. Compound 1c induced an increase of cells in G0-G1 peak and a decrease of cells in S and G2-M phases when used at a concentration of or higher than 40µM which is the TGI calculated after 24 h of 1c exposure. This could be due to a block of G1-S transition as indicated by the ability of 1c to increase the hypophosphorylate pRb/ total pRb ratio.

The aim of our study was to evaluate the capability of a crude extract of phenols from extra virgin olive oil of Moraiolo cultivar to induce apoptosis and/or differentiation in sensitive and resistant HL60 cell lines to anticancer drugs (Typical Multidrug Resistance). Our data highlight that the crude extract is able to induce apoptosis on both sensitive and resistant cells, whereas the exposure to a number of anticancer drugs does not induce apoptosis in resistant cells. In differentiation experiments we investigated the capability of crude extract of phenols to induce the expression of CD11 granulocytic or CD14 monocytic cell surface antigen in sensitive and resistant HL60 cell lines. At IC50 dose level (17 µg/ml and 32 µg/ml respectively for sensitive and resistant cell lines), the crude extract induced differentiation associated with the expression of CD14 monocytic cell surface antigen either in sensitive or resistant cell lines but not that of CD11 granulocityc cell surface antigen. Further investigations are in progress to better clarify the mechanism by which olive oil phenols induce differentiation differentiation on this cell line.

HDAC inhibitors are a new class of antitumor agents that were reported to enhance the cytotoxic effects of a number of classical anticancer drugs through multiple mechanisms. In particular, they are capable of modulating the expression of a series of key cellular genes leading to significant antiproliferative and apoptotic effects. However, which of the possible drug combinations would be the most effective and clinically useful is still to be determined. We treated the HL60 and NB4 promyelocitic leukaemia cells with a combination of the RR inhibitor 3’-Me-Ado and several hydroxamic acid–derived HDAC inhibitors, including the two recently synthesized molecules, MC1864 and MC1879, and the reference compound Trichostatin A (TSA). Initial results showed significant antiproliferative and apoptotic synergistic effects with the combinations. To investigate the molecular basis of this finding, we evaluated the effects of the combinations on cell cycle distribution and on the level of some proteins involved in the apoptotic process (p21, caspase-3, Bcl-2, Bax, AIF). Since HDAC inhibitors seemed to increase the G1-S transition block induced by 3’-Me-Ado, an effect on RR activity was hypothesized. Indeed, the HPLC evaluation of intracellular dNTP pools showed that both TSA and MC1864 induced a significant perturbation in dNTP pools, even if with a somewhat different pattern, suggesting that RR modulation could contribute to the observed synergism. Furthermore, while TSA was shown to activate the intrinsic apoptotic pathway, MC1864 induced a significant dose-dependent increase in ROS generation and the activation of an AIF-dependent apoptotic mechanism. Finally, cell exposure to the radical scavenger N-acetylcysteine determined a significant inhibition of MC1864- but not TSA-mediated synergistic effects. Hence, our findings are consistent with a possible role of HDAC inhibitor mediated-ROS induction in RR inhibition and in the potentiation of RR inhibitors-mediated apoptosis induction.