From beta-nitrothiophenes to ring-fused nitrobenzenes: an overall ring-enlargement process via a facile, aromatization-driven, thermal 6pi electrocyclization.

[reaction: see text] In prosecution of previous work on the thermal cyclization of 1-aryl-4-methanesulfonyl-2-nitro-3-phenylsulfonyl-1,3-butadienes (7), the 3-unsubstituted derivatives 8, deriving from the initial ring opening of 3-nitrothiophene (2), have been likewise found herein to undergo cyclization, followed by aromatization, in analogous mild experimental conditions, leading to the ring-fused homo- or heteroaromatic nitro derivatives 10. The concerted electrocyclic nature of the process is strongly supported by the outcome of tests based on the variation of the polarity of the solvent or of the electron density on the aryl of 8. Thus, the successful application of the process to the non-phenylsulfonyl-activated 8 significantly widens the scope of a synthetically valuable overall ring-opening/ring-closing procedure from nitrothiophenes. Support to the recently renewed interest in thermal 6pi electrocyclizations as a tool for the construction of the benzene ring is furthermore provided.

Inhibition of cell proliferation, cytotoxicity and induction of apoptosis of 1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene in gastrointestinal tumour cell lines and preliminary evaluation of its toxicity in vivo.

Our preliminary data suggested that 1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene [Viale M, Ottone M, Chiavarina B, Mariggiò MA, Prevosto C, Dell'Erba C, et al. Preliminary evaluation in vitro of the inhibition of cell proliferation, cytotoxicity and induction of apoptosis by 1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene. Invest New Drug 2004;22:359-67] (Naph-DNB), possesses good characteristics in terms of inhibition of cell proliferation in two cell lines derived from colon and gastric cancers. On this basis and to confirm the specificity of our compound towards gastrointestinal malignancies, we have analyzed the inhibition of cell proliferation, the cytotoxicity and the induction of apoptosis by Naph-DNB in seven cell lines derived from human colon (DLD-1, Lovo, HCT-8 and Colo 741), stomach (HGC-27) and pancreas (Panc-1 and Hup-T4) tumours. For the sake of comparison, cells have also been exposed to four anticancer drugs utilized for the treatment of gastrointestinal malignancies (oxaliplatin, irinotecan, gemcitabine and 5-fluorouracil). Moreover, toxicological data have been obtained in order to define the lethal dose (LD) and maximal tolerated dose (MTD) values and the spectrum of tissue alterations caused by the intraperitoneal (i.p.) and intravenous (i.v.) administration of Naph-DNB. IC50 data obtained by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay suggest that Naph-DNB is generally more active than two or more of the anticancer drugs above in most cell lines: it displayed the lowest activity only in HGC-27 cells, although data concerning the IC75 parameter enlighten a significantly better activity than irinotecan and 5-fluorouracil. Using the equitoxic concentrations IC50 and IC75, we have also evaluated the ability of Naph-DNB and of the other anticancer drugs to kill cells and to induce apoptosis. Our data show that at these concentrations Naph-DNB has a cytotoxic activity comparable or even better than that of some anticancer drugs. Similarly, Naph-DNB induces apoptosis better than the other anticancer drugs in HCT-8 and HGC-27 cells, while in Lovo and Panc-1 cells the induction is comparable. On the basis of toxicological data we defined the LD10, LD50, LD90 (i.p., 17.6, 36.1 and 54.1 mg kg(-1), respectively; i.v., 6.1, 14.1 and 22.0 mg kg(-1), respectively) and the MTD (i.p., 15 mg kg(-1); i.v., 5 mg kg(-1)) parameters. Histochemical analysis has shown that, in general, the administration of even toxic doses of Naph-DNB does not cause great structural injuries, although it can have some effects on the metabolism of glicogen and iron in organs as liver and spleen. In conclusion, our preclinical studies in vitro suggest that Naph-DNB may represent a good anticancer compound for the treatment of generally unresponsive tumours such as those of pancreas, stomach and colon. Moreover, the analysis of its toxic effects has allowed the definition of LD and MTD parameters, which will be used in further experiments in vivo for the definition of its antitumour activity.

Preliminary evaluation in vitro of the inhibition of cell proliferation, cytotoxicity and induction of apoptosis by 1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene.

The pattern of inhibition of cell proliferation and cytotoxicity in vitro by 1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene (Naph-DNB) was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and the trypan blue (TB) dye exclusion assays in nine murine and human cell lines of different histologic origin. In our culture conditions Naph-DNB showed a good inhibiting activity against all cell lines tested, with IC(50)s varying within a narrow micromolar range of concentrations (2.0 +/- 0.2-14.3 +/- 2.3 microM). In particular, murine P388 (leukemia), human Jurkat (leukemia), A2780, PA-1 (ovarian carcinoma) and Saos-2 (osteosarcoma) cells showed the highest sensitivity to the inhibiting potential of Naph-DNB, while human A549 (non small cell lung cancer, NSCLC), MDA-MB-231 (breast cancer), HGC-27 (gastric cancer) and HCT-8 (colon carcinoma) were the least sensitive cell lines. Moreover, the analysis of cytotoxicity of Naph-DNB evaluated by the TB test showed that this compound was able to kill cells with IC(50)s ranging from 1.7 to 39.2 microM. The study of the induction of apoptosis was carried out by 4'-6-diamidine-2'-phenylindole (DAPI) staining of segmented nuclei, western blot of p53 protein and TdT-mediated dUTP-biotin nick end labeling (TUNEL) method, while the interaction with DNA was evaluated through the analysis of interstrand cross-link (ISCL) formation. Our data show that in all cell lines tested Naph-DNB was able to form ISCLs, to upregulate p53 oncosuppressor-protein and to induce apoptosis. Moreover, TUNEL analysis also suggested that Naph-DNB, similarly to other anticancer drugs, was able to block cells in the G (0)/ G (1) phase of the cell cycle. In conclusion our data suggest that Naph-DNB may be an effective novel lead molecule for the design of new anticancer compounds.

1,4-Bis(1-naphthyl)-2,3-dinitro-1,3-butadiene a novel anticancer compound effective against tumor cell lines characterized by different mechanisms of resistance.

The inhibition of cell proliferation by 1,4-bis (1-naphthyl)-2,3-dinitro-1,3-butadiene (Naph-DNB) was evaluated in vitro against 4 cell lines (L1210/DDP, A2780/DX3, HCT-8/FU7dR, A549-T12) selected for their resistance to cisplatin, doxorubicin, 5-fluorouracil and taxol, and their wild-type counterparts. Naph-DNB is a novel anti-cancer compound obtained years ago within a research project of Organic Chemistry aimed at synthesizing 2,3-dinitrobutadiene derivatives. Because of its chemical structure, Naph-DNB was suggested to interact with nucleic acids, in particular DNA, and the other cellular macromolecules. This hypothesis made us consider Naph-DNB as a candidate for studies concerning its antitumour activity. We used the MTT assay to test the inhibition of cell proliferation after incubation of the cell lines with Naph-DNB for 72 h. For comparison, resistant and wild-type cell lines were also tested against those anticancer drugs used in vitro for their selection. In these culture conditions Naph-DNB retained its inhibiting activity against all resistant cells with IC50 values similar to those obtained in corresponding wild-type cell lines. Moreover, Naph-DNB was twice as effective as 5-fluorouracil against wild-type HCT-8 cells. Our previous findings about the interaction of Naph-DNB with DNA through the formation of interstrand cross-links suggested a mechanism of action similar to that of platinum/alkylating agents or topoisomerase inhibitors (intercalating agents). Our present data obtained by the K-SDS precipitation assay in A2780 and A549 cells showed that Naph-DNB is not able to form a stable topoisomerase-DNA complex as is the case for topoisomerase inhibitors. In conclusion, our results indicate that Naph-DNB is able to overcome some of the classical mechanisms of resistance selected by some anticancer drugs mainly used in clinical setting.

Access to ring-fused homo- and heteroaromatic derivatives via an initial ring-opening of 3-nitro-4-(phenylsulfonyl)thiophene.

Within an overall ring-opening/ring-forming protocol, the (E,E)-4-methylthio-2-nitro-3-phenylsulfonyl-1-pyrrolidino-1,3-butadiene (7) [derived from the initial opening of 3-nitro-4-(phenylsulfonyl)thiophene (6) with pyrrolidine and silver nitrate in EtOH] is revealed to be an excellent precursor of nitro(phenylsulfonyl) derivatives of ring-fused aromatic (naphthalene, phenanthrene) or heteroaromatic (benzothiophene) compounds whose substitution pattern cannot be easily achieved by conventional methods. The key step is represented by a thermal electrocyclic rearrangement of (E,E)-1-aryl-4-methylsulfonyl-2-nitro-3-phenylsulfonyl-1,3-butadienes (9), which, thanks to proper geometric and electronic factors, occurs in unprecedentedly mild conditions and is followed by an irreversible, concerted syn beta-elimination of methanesulfinic acid to aromatize the newly formed cyclohexadienic ring.