Annexin–V FITC/PI assay.

<p>MDA-MB-231 cells were treated with the complexes AuD6 and AuD8 (20 µM) for 16 and 24 h. Then, cells were labeled with Annexin–V FITC and PI and analyzed by flow cytometry in order to evaluate the percentage of apoptotic cells. Apoptotic cells at early stage occur in the lower right quadrant while apoptotic cells at late stage set in the up-right part. The percentage in the lower left quadrant is due to viable cells whereas the upper left part to non-apoptotic cell death.</p

Western blot and morphological analysis (concentration-dependent study).

<p><b>A</b>, Western blot analysis of breast cancer MDA-MB-231 cell extracts. Cells were treated with the complexes AuD6 and AuD8 at the indicated concentrations for 24 h. <b>B</b>, Western blot analysis of the p27 protein amount in MDA-MB-231 cell extract after treatment with the compound AuD8 at the indicated concentrations for 24 h. The solvent DMSO was used as a control while GAPDH as a loading control. <b>C</b>, Apoptotic morphological changes of MDA-MB-231 cells after treatment with AuD6 and AuD8 at the indicated concentrations for 24 h (phase contrast imaging, 100× magnification).</p

<i>In vitro</i> inhibition of proteasome.

<p>IC₅₀ values (µM±SD) obtained for the three proteasomal activities on the purified 20S proteasome and on MDA-MB-231 cell extract after 2 h incubation.</p

Western blot <i>in vivo</i> analysis.

<p>Female nude mice bearing MDA-MB-231 tumors were treated with either vehicle (control) or the compounds AuD6 and AuD8 at 1 mg kg⁻¹ d⁻¹. Tumors were collected and the corresponding tissues prepared for Western blot analysis after either 13-day treatment (<b>A</b>) or 27-day treatment (<b>B</b>) [I and II denote distinct experiments]. Tissues were also prepared for the assays of caspase-3 activity (<b>C</b>) and of proteasomal CT-like activity (<b>D</b>) after 27 days of treatment.</p

Chemical structures of investigated gold(III)-based compounds AuD6 (A) and AuD8 (B).

<p>Chemical structures of investigated gold(III)-based compounds AuD6 (A) and AuD8 (B).</p

Antitumor activity <i>in vivo</i> on MDA-MB-231 xenografts.

<p>Female nude mice bearing MDA-MB-231 tumors were treated with either vehicle (control) or the compounds AuD6 and AuD8 at 1 mg kg⁻¹ d⁻¹. <b>A,</b> Inhibition of xenograft growth by both complexes. Tumor volumes were measured every other day using a caliper. Points represent the mean ± SD (bars) of seven mice per group. The insert depicts representative tumors from each treatment group; * = p<0.05. <b>B</b>, If only the most responsive mice are considered, the xenograft growth inhibition is greater. The insert shows average weights of mice over time; ** = p<0.01. <b>C</b>, Immunohistochemical p27 and TUNEL staining of tumor samples indicates proteasome inhibition and apoptosis as a result of both compounds. Stronger p27 staining is observed following AuD8 treatment, and more TUNEL positive cells are observed following AuD6 treatment. Brown colored cells are considered positive.</p

Toward the Selective Delivery of Chemotherapeutics into Tumor Cells by Targeting Peptide Transporters: Tailored Gold-Based Anticancer Peptidomimetics

Complexes [Au^IIIX₂(dtc-Sar-AA-O­(<i>t</i>-Bu))] (AA = Gly, X = Br (<b>1</b>)/Cl (<b>2</b>); AA = Aib, X = Br (<b>3</b>)/Cl (<b>4</b>); AA = l-Phe, X = Br (<b>5</b>)/Cl (<b>6</b>)) were designed on purpose in order to obtain gold­(III)-based anticancer peptidomimetics that might specifically target two peptide transporters (namely, PEPT1 and PEPT2) upregulated in several tumor cells. All the compounds were characterized by means of FT-IR and mono- and multidimensional NMR spectroscopy, and the crystal structure of [Au^IIIBr₂(dtc-Sar-Aib-O­(<i>t</i>-Bu))] (<b>3</b>) was solved and refined. According to in vitro cytotoxicity studies, the Aib-containing complexes <b>3</b> and <b>4</b> turned out to be the most effective toward all the human tumor cell lines evaluated (PC3, DU145, 2008, C13, and L540), reporting IC₅₀ values much lower than that of cisplatin. Remarkably, they showed no cross-resistance with cisplatin itself and were proved to inhibit tumor cell proliferation by inducing either apoptosis or late apoptosis/necrosis depending on the cell lines. Biological results are here reported and discussed in terms of the structure–activity relationship

Circularly Polarized Luminescence from New Heteroleptic Eu(III) and Tb(III) Complexes

The complexes [Eu(bpcd)(tta)], [Eu(bpcd)(Coum)], and [Tb(bpcd)(Coum)] [tta = 2-thenoyltrifluoroacetyl-acetonate, Coum = 3-acetyl-4-hydroxy-coumarin, and bpcd = N,N′-bis(2-pyridylmethyl)-trans-1,2-diaminocyclohexane-N,N′-diacetate] have been synthesized and characterized from photophysical and thermodynamic points of view. The optical and chiroptical properties of these complexes, such as the total luminescence, decay curves of the Ln(III) luminescence, electronic circular dichroism, and circularly polarized luminescence, have been investigated. Interestingly, the number of coordinated solvent (methanol) molecules is sensitive to the nature of the metal ion. This number, estimated by spectroscopy, is >1 for Eu(III)-based complexes and <1 for Tb(III)-based complexes. A possible explanation for this behavior is provided via the study of the minimum energy structure obtained by density functional theory (DFT) calculations on the model complexes of the diamagnetic Y(III) and La(III) counterparts [Y(bpcd)(tta)], [Y(bpcd)(Coum)], and [La(bpcd)(Coum)]. By time-dependent DFT calculations, estimation of donor–acceptor (D–A) distances and of the energy position of the S1 and T1 ligand excited states involved in the antenna effect was possible. These data are useful for rationalizing the different sensitization efficiencies (ηsens) of the antennae toward Eu(III) and Tb(III). The tta ligand is an optimal antenna for sensitizing Eu(III) luminescence, while the Coum ligand sensitizes better Tb(III) luminescence {ϕovl = 55%; ηsens ≥ 55% for the [Tb(bpcd)(Coum)] complex}. Finally, for the [Eu(bpcd)(tta)] complex, a sizable value of glum (0.26) and a good quantum yield (26%) were measured