Short communicationSensitivity of different resistant tumour cell lines to the two novel compounds (2Z,4E)-2-methylsulfanyl-5-(1-naphthyl)-4-nitro-2,4-pentadienoate and (1E,3E)-1,4-bis(2-naphthyl)-2,3-dinitro-1,3-butadiene
Introduction
In an attempt to generate new antitumour compounds with good properties in terms of antiproliferative and pro-apoptotic activity as well as of low toxicity in vivo and ability to overcome the most frequent mechanisms of resistance responsible for the failure of a high number of chemotherapeutic cancer treatments, we synthesized (2Z,4E)-2-methylsulfanyl-5-(1-naphthyl)-4-nitro-2,4-pentadienoate (1-Naph-NMCB, Petrillo et al., 2008) and (1E,3E)-1,4-bis(2-naphthyl)-2,3-dinitro-1,3-butadiene (2-Naph-DNB, Viale et al., 2007). The first compound is basically the result of the application of the “molecular simplification strategy” (Manetti et al., 2000, Crisòstomo et al., 2006) on the lead compound (1E,3E)-1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene (1-Naph-DNB, Viale et al., 2004, Novi et al., 2004, Dell'Erba et al., 2005), whereas the second molecule is a structural isomer of the latter, characterized by a different spatial arrangement (Fig. 1).
When tested in vitro for its inhibition of cell proliferation and apoptotic activity 1-Naph-NMCB showed a significant activity at micromolar concentrations, in particular against the MDA-MB-231 breast cancer cell line, and with a significant general improvement compared to 1-Naph-DNB (Petrillo et al., 2008). In particular, for apoptosis, 1-Naph-NMCB showed an activity that was in some cases better than that observed for 1-Naph-DNB, as evaluated by the morphological analysis of nuclear segmentation, the staining with Annexin V and the analysis in western blot of p53 oncosuppressor protein (Petrillo et al., 2008). Moreover, both the analysis of formation of interstrand cross-links and the analysis of the inhibition of restriction enzyme cutting activity in λ phage DNA treated with the compound (unpublished data) demonstrate the binding of 1-Naph-NMCB to DNA, although lower than that observed for the parent compound 1-Naph-DNB (Petrillo et al., 2008).
In vivo studies allowed the toxicological (determination of lethal and maximal tolerated doses) and pharmacological evaluation of 1-Naph-NMCB. Our findings showed that our compound was characterized by negligible histological toxic effects and an antitumour activity which was in some cases even better than that showed by 1-Naph-DNB, and was characterized by differences in tumour selectivity (Petrillo et al., 2007).
2-Naph-DNB showed significant features in terms of inhibition of cell proliferation, with a better activity than 1-Naph-DNB in MDA-MB-231 cells and HGC-27 human gastric cell line, and induction of apoptosis which was linked to the activation of p53 protein and the formation of interstrand cross-links to DNA (Viale et al., 2007). The latter observation was also confirmed by experiments using treated λ phage DNA and restriction enzymes (unpublished data). As for 1-Naph-NMCB, also 2-Naph-DNB was characterized in vivo by low toxic effect at histological level.
All these features prompted us to verify the ability of our molecules to overcome the different mechanisms of resistance showed by some cell lines selected in vitro for their resistance to important anticancer drugs such as doxorubicin, taxol, cisplatin and 5-fluorouracil, also in order to gain further insights into their mechanisms of action.
Section snippets
Chemicals
The two naphthylnitrobutadienes 2-Naph-DNB and 1-Naph-NMCB were synthesized from 3,4-dinitrothiophene (3,4-DNT) (Viale et al., 2007) and methyl 4-nitrothiophene-2-carboxylate (2-COOMe-4-NT) (Petrillo et al., 2007) as already described (Scheme 1).
While 1-Naph-NMCB differs from the lead compound 1-Naph-DNB (Petrillo et al., 2008) since it contains only one of the two original naphthylnitroethenyl moieties, the isomeric 2-Naph-DNB simply differs from 1-Naph-DNB for a different spatial arrangement
Inhibition of cell proliferation and morphology of microtubules
In human A2780 cells, sensitive (doubling time 24 h) and resistant to doxorubicin (doubling time 27 h), both 1-Naph-NMCB and 2-Naph-DNB were active at the micromolar concentration level. Both molecules were less active than doxorubicin in sensitive A2780 cells, on the other hand their IC50s for resistant and sensitive cells were nearly similar. On the basis of the results showed in Table 1, the resistance indexes for doxorubicin, 1-Naph-NMCB and 2-Naph-DNB were 63.0, 1.8 and 1.2, respectively.
Discussion
As reported in a previous paper (Novi et al., 2004) the mechanisms of resistance of the cell lines used in our experiments are quite well characterized. All these cells were selected in vitro by prolonged exposure to the different selective anticancer drugs.
Briefly, the main mechanism of resistance of L1210/DDP cells consists of a decreased accumulation of cisplatin and a reduced DNA platination (Richon et al., 1987). The resistance of HCT-8/FU7dR cell lines was linked to the shortage of
Acknowledgements
This research was supported by funds from M.I.U.R. (FIRB 2001) and from the Universities of Genova and Bologna.
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