Among ethanol extracts of 10 edible berries, bilberry extract was found to be the most effective at inhibiting the growth of HL60 human leukemia cells and HCT116 human colon carcinoma cells in vitro. Bilberry extract induced apoptotic cell bodies and nucleosomal DNA fragmentation in HL60 cells. The proportion of apoptotic cells induced by bilberry extract in HCT116 was much lower than that in HL60 cells, and DNA fragmentation was not induced in the former. Of the extracts tested, that from bilberry contained the largest amounts of phenolic compounds, including anthocyanins, and showed the greatest 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity. Pure delphinidin and malvidin, like the glycosides isolated from the bilberry extract, induced apoptosis in HL60 cells. These results indicate that the bilberry extract and the anthocyanins, bearing delphinidin or malvidin as the aglycon, inhibit the growth of HL60 cells through the induction of apoptosis. Only pure delphinidin and the glycoside isolated from the bilberry extract, but not malvidin and the glycoside, inhibited the growth of HCT116 cells.

DNase I footprinting has revealed that zinc(II) complexes with macrocyclic tetraamines (1,4,7,10-tetraazacyclododecane,cyclen) appended with one or two aryl-methyl group(s), ((9-acridinyl)methyl-, (4-quinolyl)methyl-, 1,7-bis(4-quinolyl)methyl-, (1-naphthyl)methyl-, and 1,7-bis(1-naphthyl)methyl-cyclen) selectively bind to native double-stranded DNA (150 base pairs), at AT-rich regions like classical minor groove binders (distamycin A and 4,6-diamidino-2-phenylindole (DAPI)). The selectivity and affinity depend on the stacking ability and number of the aromatic ring. Zn2+ is an essential metal ion for the DNA binding, which cannot be replaced by other metal ions such as Cu2+ or Ni2+. The DNA binding by these Zn2+−cyclen derivatives was inhibited by captopril having a stronger affinity for the fifth coordination site of the Zn2+−cyclen complexes. Micrococcal nuclease footprinting, moreover, revealed that those Zn2+−cyclen derivatives bound only to the thymine groups in the A−T base pairs, while distamycin A and DAPI simultaneously bound to the thymine and adenine groups in the A−T base pairs. Distamycin A and the Zn2+−(4-quinolyl)methyl-cyclen reversibly competed for common AT-rich regions of minor groove. The DNA binding mode by the Zn2+−cyclen derivatives was due to the selective and strong complex formation between the Zn2+−cyclen moiety and the imide-deprotonated thymine at neutral pH.

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The characteristic binding mode of zinc(II) complexes of macrocyclic tetraamines (1,4,7,10-tetraazacyclododecane, cyclen) appended with one or two arylmethyl group(s) [(4-quinolyl)methyl-, 1,7-bis(4-quinolyl)methyl-, (1-naphthyl)methyl-, 1,7-bis(1-naphthyl)methyl-, and (9-acridinyl)methyl-cyclen] to double-stranded calf thymus DNA and synthetic DNAs [poly(dA)-poly(dT), poly(dA-dT)2, poly(dI).poly(dC), poly(dI-dC)2, poly(dG) poly(dC), and poly(dG-dC)2] has been examined by spectrophotometric methods, Tm measurement, and inhibition of these DNA-directed transcriptions in vitro. Various hypochromic and bathochromic effects on the pendant aromatic absorption spectra of the complexes were observed in titration with the native and synthetic DNA. The binding constants Kapp (=[bound cyclen derivatives]/[unbound cyclen derivatives][DNA phosphates] M(-1)), at 25 degrees C in 10 mM EPPS (pH 8.0) containing 0.1 M Na+, were determined and compared with those of the corresponding Zn2+ -free ligands. The results showed that the Zn2+ -cyclen complexes interact with the DNA more strongly than the corresponding diprotonated ligands, leading to a stronger stacking of the pendant aromatic rings. The binding of Zn2+ -(9-acridinyl)methyl-cyclen to calf thymus DNA was competed by an AT-selective, minor groove binder, distamycin, but not by a major groove binder, methyl green. In an unusual interaction of excess Zn2+ -(9-acridinyl)methyl-cyclen with poly(dA).poly(dT), the Zn2+ -cyclen moiety went into the minor groove to make coordination bonds with the deprotonated imides of the thymines, resulting in disruption of the poly(dA).poly(dT) duplex. Thymine-containing DNA-directed transcription with Escherichia coli RNA polymerase in vitro was inhibited by the Zn2+ -(9-acridinyl)methyl-cyclen. The 50% inhibition concentrations of the transcription (IC50) were 22-45 microM with poly(dA).poly(dT) or poly(dA-dT)2 as templates, while with poly(dG-dC)2 as a template the IC50 value was 110 microM.

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