A critical comparison of methods to prepare sterically hindered 3-aryl isoxazoles containing fused aromatic rings using the nitrile oxide cycloaddition (NOC) reveal that modification of the method of Bode, Hachisu, Matsuura, and Suzuki (BHMS), utilizing either triethylamine as base or sodium enolates of the diketone, ketoester, and ketoamide dipolarophiles, respectively, was the method of choice for this transformation.

A series of 7-amino- and 7-acetamidoquinoline-5,8-diones with aryl substituents at the 2-position were synthesized, characterized and evaluated as potential NAD(P)H:quinone oxidoreductase (NQO1)-directed antitumor agents. The synthesis of lavendamycin analogs is illustrated. Metabolism studies demonstrated that 7-amino-analogues were generally better substrates for NQO1 than 7-amido-analogues as were compounds with smaller heteroaromatic substituents at the C-2 position. Surprisingly, only two compounds, 7-acetamido-2-(8’-quinolinyl)quinoline-5,8-dione (11) and 7-amino-2-(2-pyridinyl)quinoline-5,8-dione (23) showed selective cytotoxicity toward the NQO1-expressing MDA468-NQ16 breast cancer cells versus the NQO1-null MDA468-WT cells. For all other compounds, NQO1 protected against quinoline-5,8-dione cytotoxicity. Compound 22 showed a potent activity against human breast cancer cells expressing or not expressing NQO1 with IC50 values of respectively 190 nM and 140 nM and a low NQO1 mediated reduction rate, which suggests that the mode of action of 22 differs from lavendamycin and involves an unidentified target(s).

Using the structure-activity relationship emerging from previous reports, and guided by pharmacokinetic properties, new AIMs have been prepared with both improved efficacy against human glioblastoma cells and cell permeability as determined by fluorescent confocal microscopy. We present our first unambiguous evidence for telomeric G4-forming oligonucleotide anisotropy by NMR resulting from direct interaction with AIMs, which is consistent with both our G4 melting studies by CD, and our working hypothesis. Finally, we show that AIMs induce apoptosis in SNB-19 cells.

The asymmetric unit of the title compound, C21H16ClNO4, contains two independent mol­ecules (A and B), each adopting a conformation wherein the isoxazole ring is roughly orthogonal to the anthrone ring. The dihedral angle between the mean plane of the isoxazole (all atoms) and the mean plane of the anthrone (all atoms) is 88.48 (3)° in one mol­ecule and 89.92 (4)° in the other. The ester is almost coplanar with the isoxazole ring, with mean-plane dihedral angles of 2.48 (15) and 8.62 (5)°. In both mol­ecules, the distance between the ester carbonyl O atom and the anthrone ketone C atom is about 3.3 Å. The anthrone ring is virtually planar (r.m.s. deviations of 0.070 and 0.065 Å) and adopts a shallow boat conformation in each mol­ecule, as evidenced by the sum of the six intra-B-ring torsion angles [41.43 (15) and 34.38 (15)° for molecules A and B, respectively]. The closest separation between the benzene moieties of anthrones A and B is 5.1162 (7) Å, with an angle of 57.98 (5)°, consistent with an edge-to-face π-stacking inter­action. In the crystal, weak C—H⋯O and C—H⋯N inter­actions link the mol­ecules, forming a three-dimensional network.

In the United States, the average survival rate for malignant brain tumors is approximately one year. Despite recent advancements in treatment, there remains a need for the development of new therapeutic agents to target and treat malignant brain tumors to increase survival and quality of life. Due to its pivotal role in cancer cell proliferation, telomerase has become an attractive target in the development of new anticancer agents. The enzymatic activity of telomerase is responsible for telomere maintenance and adds the guanine-rich DNA sequence repeats to the 3′ end of DNA strands in the telomere regions. Human telomeric DNA has the potential to adopt G-quartet conformations, which in turn can stack in various arrangements to form G quadruplexes (G4). To this end, molecules that bind telomeric G4 DNA and inhibit telomerase have substantial anticancer potential. The objective of this study was to evaluate the cytotoxicity of a novel series of anthracenyl isoxazole amides (AIMs), capable of selectively binding to G4 telomeric DNA, and determine the ability of the AIMs to penetrate cell membranes and enter the nuclei of cancer cells. These AIMs have demonstrated significant activity against brain tumor cell lines in the DTP NCI-60 cell line panel, and COMPARE analysis indicated that these molecules exerted their cytotoxicity via a unique mechanism. Two AIMs, referred to here as AIM-dimer and AIM-double tail (AIM-dt), showed single digit micromolar IC50 values (4.75 +/− 0.27 and 3.13 +/− 0.45, respectively) by MTT assay when tested in the human glioblastoma cell line SNB-19. As a followup to the cytotoxicity studies, apoptosis was evaluated by flow cytometry (Annexin V). The results indicated that both AIMs induced apoptosis in SNB-19 cells at higher levels than untreated cells after a four-hour treatment. Since the AIMs are known to autofluoresce, confocal microscopy was performed to determine if they were able to penetrate cell membranes of the SNB-19 cells. Contrary to bioavailability calculations made with Symyx Draw v3.1, which suggested that only the AIM-dt had acceptable properties for cell penetration, both compounds were found to enter the cells. Perhaps more intriguing, however, are the results from the laser scanning cytometry (iCys) studies. SNB-19 cells were co-treated with either AIM-dimer or AIM-dt and the nuclear stain propidium iodide. Images taken of each AIM clearly indicated co-localization of the AIM at the nucleus, suggesting that these AIMs are able to penetrate cell membranes and reach the nuclei and therefore have potential to bind DNA. Further studies will be focused on elucidating the mechanism of action of the AIMs as well as confirming their interactions at the level of G4 telomeric DNA. Supported by NIH Grant P20RR017670 (HDB). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1767. doi:1538-7445.AM2012-1767