Triplex metallohelices have enantiomer-dependent mechanisms of action in colon cancer cells.

Self-assembled enantiomers of an asymmetric di-iron metallohelix differ in their antiproliferative activities against HCT116 colon cancer cells such that the compound with Λ-helicity at the metals becomes more potent than the Δ compound with increasing exposure time. From concentration- and temperature-dependent 57Fe isotopic labelling studies of cellular accumulation we postulate that while the more potent Λ enantiomer undergoes carrier-mediated efflux, for Δ the process is principally equilibrative. Cell fractionation studies demonstrate that both enantiomers localise in a similar fashion; compound is observed mostly within the cytoskeleton and/or genomic DNA, with significant amounts also found in the nucleus and membrane, but with negligible concentration in the cytosol. Cell cycle analyses using flow cytometry reveal that the Δ enantiomer induces mild arrest in the G1 phase, while Λ causes a very large dose-dependent increase in the G2/M population at a concentration significantly below the relevant IC50. Correspondingly, G2-M checkpoint failure as a result of Λ-metallohelix binding to DNA is shown to be feasible by linear dichroism studies, which indicate, in contrast to the Δ compound, a quite specific mode of binding, probably in the major groove. Further, spindle assembly checkpoint (SAC) failure, which could also be responsible for the observed G2/M arrest, is established as a feasible mechanism for the Λ helix via drug combination (synergy) studies and the discovery of tubulin and actin inhibition. Here, while the Λ compound stabilizes F-actin and induces a distinct change in tubulin architecture of HCT116 cells, Δ promotes depolymerization and more subtle changes in microtubule and actin networks.

Comment on "Delivering a photosensitive transplatin prodrug to overcome cisplatin drug resistance" by H. Song, W. Li, R. Qi, L. Yan, X. Jing, M. Zheng and H. Xiao, Chem. Commun., 2015, 51, 11493.

Consistency of literature results with the transformation of trans-diamminedichloridoplatinum(ii) (transplatin) into cis-diammine-dichloridoplatinum(ii) (cisplatin) under UVA irradiation claimed in the article recently published in this journal is questioned on the basis of previous and new experimental data.

Conformational analysis of site-specific DNA cross-links of cisplatin-distamycin conjugates.

The requirement for novel platinum antitumor drugs led to the concept of synthesis of novel platinum drugs based on targeting cisplatin to various carrier molecules. We have shown [Loskotova, H., and Brabec, V. (1999) Eur. J. Biochem. 266, 392-402] that attachment of DNA minor-groove-binder distamycin to cisplatin changes several features of DNA-binding mode of the parent platinum drug. Major differences comprise different conformational changes in DNA and a considerably higher interstrand cross-linking efficiency. The studies of the present work have been directed to the analysis of oligodeoxyribonucleotide duplexes containing single, site-specific adducts of platinum-distamycin conjugates. These uniquely modified duplexes were analyzed by Maxam-Gilbert footprinting, phase-sensitive gel electrophoresis bending assay and chemical probes of DNA conformation. The results have indicated that the attachment of distamycin to cisplatin mainly affects the sites involved in the interstrand cross-links so that these adducts are preferentially formed between complementary guanine and cytosine residues. This interstrand cross-link bends the helix axis by approximately 35 degrees toward minor groove, unwinds DNA by approximately 95 degrees and distorts DNA symmetrically around the adduct. In addition, CD spectra of restriction fragments modified by the cisplatin-distamycin conjugates have demonstrated that distamycin moiety in the interstrand cross-links of these compounds interacts with DNA. This interaction facilitates the formation of these adducts. Hence, the structural impact of the specific interstrand cross-link detected in this study deserves attention when biological behavior of cisplatin derivatives targeted by oligopeptide DNA minor-groove-binders is evaluated.

Mutations of conserved arginines in the membrane domain of erythroid band 3 lead to a decrease in membrane-associated band 3 and to the phenotype of hereditary spherocytosis.

To elucidate the molecular basis of band 3 deficiency in a recently defined subset of patients with autosomal dominant hereditary spherocytosis (HS), we screened band 3 cDNA for single-strand conformation polymorphism (SSCP). In 5 of 17 (29%) unrelated HS subjects with band 3 deficiency, we detected substitutions R760W, R760Q, R808C, and R870W that were all coinherited with the HS phenotype. The involved arginines are highly conserved throughout evolution. To examine whether or not the product of the mutant allele is inserted into the membrane, we studied one HS subject who was doubly heterozygous for the R760Q mutation and the K56E (band 3sMEMPHIS) polymorphism that results in altered electrophoretic mobility of the band 3 Memphis proteolytic fragments. We detected only the band 3MEMPHIS in the erythrocyte membrane indicating that the protein product of the mutant, R760Q, band 3 allele is absent from the red blood cell membrane. These findings suggest that the R760Q substitution, and probably the other arginine subsitutions, produce band 3 deficiency either by precluding incorporation of the mutant protein into the red blood cell membrane or by leading to loss of mutant protein from differentiating erythroid precursors.

[Treatment of iron overload states with oral administration of the chelator agent, L1 (Deferiprone)]. / Lécba stavu s pretízením zelezem podáváním perorálního chelátoru L 1 (Deferiprone).

Nine iron overloaded patients were treated with L1--Deferiprone (1,2-dimethyl-3-hydroxypyrid-4-one) in daily dose 3 g (40-50 mg/kg) for 12 weeks. In 7 patients the efficiency of L 1 treatment was compared to the therapeutic effect of the same dose of desferrioxamine (Desferal). A significant increase in urinary iron excretion was observed after administration of both chelating agents. Iron excretion after L 1 treatment was approximately 65% of that obtained with Desferal. The amount of excreted iron correlated with the amount of iron stores before chelation. A significant decrease in transferrin saturation, serum and red cell ferritin was observed after treatment with Desferal, L 1 administration caused a significant decrease only in serum ferritin level. However, all the parameters reflecting iron stores remained increased when compared to normal values after 12 weeks of chelation therapy. An incomplete absorption from gut and some reutilization of chelated iron may be responsible for less potent iron chelation by L 1 in comparison to Desferal. A low tolerance of the drug together with repeated nausea and vomiting were the most frequent adverse effects observed in the course of L1 administration. L 1 treatment had to be discontinued due to repeated vomiting in one patient and due to progressive granulocytopenia and thrombocytopenia in another patient. Because of the side effects more clinical studies with L 1 are needed before its introduction in wide clinical practice.

[Iron stores in patients with chronic kidney failure treated with recombinant human erythropoietin]. / Sledování zásob zeleza u nemocných s chronickým selháním ledvin lécených rekombinantním lidským erytropoetinem.

Nine patients with anemia of chronic renal failure were treated with recombinant human erythropoietin (rHuEPO) in dose 50-150 IU/kg/week. After 8 weeks the treatment was maintained with 30-50 IU/kg/week for one year. A significant increase of hemoglobin (Hb) level and red blood cell (RBC) count was observed in all patients. Administration of rHUEPO maintained Hb level higher than 100 milligrams and RBC count above 3.0 x 10(12)/l. Iron stores decreased in all patients. Parameters reflecting either the real amount of iron available for erythropoiesis or iron stores in erythroid precursors, i.e. red cell ferritin (eF), free erythrocyte protoporphyrin (FEP) and transferrin saturation (satTRF) were the most reliable tools for diagnosis of iron deficient erythropoiesis. Serum ferritin (sF) was not decreased in most patients, however, sF level was below 50 micrograms/l in all patients at the time of diagnosis of iron deficiency. Iron supplementation in a daily dose allowing absorption of 100mg of elementary iron was sufficient to cover the increased demand for iron in rHuEPO treated patients.

Evaluation of cytotoxic and antitumor effects of a tetravalent analog of carboplatin.

The cytostatic and cytocidal effects of a newly synthesized tetravalent platinum drug (CBDCA-ox) on two experimental ascites tumors as well as on normal tissues of the mouse were investigated. It was found that CBDCA-ox exhibited only negligible acute toxicity in comparison with its reduced analog. Also hemotoxicity of this tetravalent drug was markedly lower than that of CBDCA. The comparison of antitumor activity of CBDCA and CBDCA-ox tested against LS/BL lymphosarcoma and EAT tumor cells strongly supports the view that CBDCA-ox deserves further preclinical testing.