Abderraouf Kenani

Cytotoxicity, intracellular distribution and uptake of doxorubicin and doxorubicin coupled to cell-penetrating peptides in different cell lines: a comparative study.

One of the major obstacles which are opposed to the success of anticancer treatment is the cell resistance that generally develops after administration of commonly used drugs. In this study, we try to overcome the tumour cell resistance of doxorubicin (Dox) by developing a cell-penetrating peptide (CPP)-anticancer drug conjugate in aim to enhance its intracellular delivery and that its therapeutic effects. For this purpose, two cell-penetrating peptides, penetratin (pene) and tat, derived from the HIV-1 TAT protein, were chemically conjugated to Dox. The cytotoxicity, intracellular distribution and uptake were accessed in CHO cells (Chinese Hamster Ovarian carcinoma cells), HUVEC (Human Umbilical Vein Endothelial Cells), differentiated NG108.15 neuronal cell and breast cancer cells MCF7drug-sensitive or MDA-MB 231 drug-resistant cell lines. The conjugates showed different cell killing activity and intracellular distribution pattern by comparison to Dox as assessed respectively by MTT-based colorimetric cellular cytotoxicity assay, confocal fluorescence microscopy and FACS analysis. After treatment with 3 microM with Dox-CPPs for 2h, pene increase the Dox cytotoxicity by 7.19-fold in CHO cells, by 11.53-fold in HUVEC cells and by 4.87-fold in MDA-MB 231 cells. However, cytotoxicity was decreased in NG108.15 cells and MCF7. Our CPPs-Dox conjugate proves the validity of CPPs for the cytoplasmic delivery of therapeutically useful molecules and also a valuable strategy to overcome drug resistance.

Copper(II)- and iron(II)-complexes of methyl 2-(2-aminoethyl)-aminomethyl-pyridine-6-carboxyl-histidinate (AMPHIS), a peptide mimicking the metal-chelating moiety of bleomycin. An ESR investigation

Methyl 2-(2-aminoethyl)-aminomethyl-pyridine-6-carboxyl-histidinate (AMPHIS), a synthetic analogue of the chelating part of bleomycin (BLM), has been studied for its metal binding properties. Electron spin resonance parameters of AMPHIS-Cu(II) and BLM-Cu(II) have been found to be closely similar likewise spectra of oxygen radicals spin-adducts induced by AMPHIS-Fe(II)-O2 and BLM-Fe(II)-O2 systems. Thus, AMPHIS could constitute a very useful tool for the study of BLM mode of action.

A convenient method for the cleavage of the d-mannosyl-l-gulose disaccharide from bleomycin-A2

In order to elucidate the biological role of the sugar residue of the antitumor drug bleomycin, this was deglycosylated by beta-elimination under mild alkaline conditions, and by solvolysis with hydrogen fluoride. The latter procedure proved to be better because it led to the complete deglycosylation without modification of the peptide, thus allowing further biological investigations of this component.

Doxorubicin coupled to penetratin promotes apoptosis in CHO cells by a mechanism involving c-Jun NH2-terminal kinase

Doxorubicin (Dox) has demonstrated potent activity in treating malignant lymphomas but its therapeutic efficacy is hampered by induction of cardiotoxicity. This side effect is related to the ability of the drug to generate reactive oxygen species in cells. Previously, we demonstrated that coupling Dox to penetratin (Pen), a cell penetrating peptide, represent a valuable strategy to overcome drug resistance in CHO cells. In the present study, we evaluated the consequences of the conjugation of Dox to Pen in term of apoptosis induction. When tested on CHO cells, Dox-Pen generated a typical apoptotic phenotype but at lower dose that needed for unconjugated Dox. Cell death induction was associated with chromatin condensation, caspase activation, Bax oligomerisation and release of cytochrome c. By using reactive oxygen species and c-jun NH2-terminal kinase (JNK) inhibitors, we prevented Dox- and Dox-Pen-induced CHO cell death. The chimeric soluble DR5 receptor that inhibits TRAIL induced cell death does not prevent Dox or Dox-Pen-induced cytotoxicity. These observations indicate that conjugation of Dox to cell penetrating peptide does not impair the ability of the drug to trigger cell death through activation of the intrinsic pathway involving c-Jun NH2-terminal kinase but could exhibit less toxic side effects and could warrant its use in clinic.

Comparative subcellular distribution of the copper complexes of bleomycin-A2 and deglycobleomycin-A2

We have compared the cellular uptake and subcellular localization of 14C-labeled bleomycin-A2 (BLM) and deglycobleomycin-A2 in living KB3 cells. Both drugs exhibit poor internalization into cells but reveal significantly different intracellular distribution, with low and high accumulation into the cell nuclei for BLM and deglyco-BLM, respectively. The results indicate that the carbohydrate chain does not constitute a limiting factor for BLM permeation and is directly implicated in the intracellular distribution of the drug into cells.

Metal-complexing, DNA-binding and DNA-cleaving properties of a synthetic molecule AMBIGLU, a simplified model for the study of bleomycin

Abstract On the basis of the previous studies on simple synthetic molecules structurally related to the anti-tumor drug bleomycin-A 2 (BLM-A 2 ), the roles of the main parts of the parent compound, a metal-chelating peptide, a DNA-binding heterocycle and a protecting activating sugar residue, were delineated. A new synthetic compound, AMBIGLU was designed taking into account these results. The synthesis, the copper-chelating properties, the radical production, the DNA-binding and DNA-cleaving ability are described here and compared to those of BLM-A 2 .

Original paperMetal-complexing, DNA-binding and DNA-cleaving properties of a synthetic molecule AMBIGLU, a simplified model for the study of bleomycinComplexation des métaux, liaison à l'ADN et coupure de l'ADN par une molécule synthétique, AMBIGLU, un modèle simplifié pour l'étude de la bléomycine

Abstract On the basis of the previous studies on simple synthetic molecules structurally related to the anti-tumor drug bleomycin-A 2 (BLM-A 2 ), the roles of the main parts of the parent compound, a metal-chelating peptide, a DNA-binding heterocycle and a protecting activating sugar residue, were delineated. A new synthetic compound, AMBIGLU was designed taking into account these results. The synthesis, the copper-chelating properties, the radical production, the DNA-binding and DNA-cleaving ability are described here and compared to those of BLM-A 2 .

Analogue versus digital recognition of DNA by bleomycin: an effect of the carbohydrate moiety

We have sought to determine the influence of the carbohydrate moiety of the antitumour antibiotic bleomycin on the sequence‐specific cleavage of DNA. Both bleomycin A2 and deglycobleomycin A2 produce different cleavage patterns with DNA in which the 2‐amino group has been removed from guanine, added to adenine, or both, as well as on a designed DNA fragment containing a few defined cleavage sites. Although each drug cleaves DNA primarily at Gp and Gp sites, the cleavage at these sites is frequently found to be stronger with deglycobleomycin compared with bleomycin A2. Conversely, in most cases the cleavage at secondary sites, in particular at ApT steps, is significantly reduced or even abolished with deglycobleomycin. The results indicate that the gulose‐mannose moiety of bleomycin A2 plays a significant role in the recognition of preferred nucleotide sequences and confirm the view that both secondary structure and interaction with guanine are involved in determining sequence‐specific cleavage of DNA by bleomycin.

DNA-binding and DNA-cleaving properties of a synthetic model AGAGLU related to the antitumour drugs AMSA and bleomycin

We have previously described two synthetic models gathering a simplified model of the complexing part of Bleomycin (Blm) and the intercalating moiety of m-AMSA. These molecules, namely AGGA and AGAMGA, do not seem able to cleave DNA as Blm does. The present work is devoted to the study of a new derivative, AGAGLU, which includes in its structure a judiciously chosen connector between the two parts of the molecule. This compound, the chelating and DNA-binding properties of which are described here, has been shown to induce single-strand breakage of duplex DNA in a high level.

Polymorphism on Chromosome 9p21.3 Is Associated with Severity and Early-Onset CAD in Type 2 Diabetic Tunisian Population

Multiple association studies found that the human 9p21.3 chromosome locus is a risk factor for atherosclerosis. The purpose of this study was to investigate the association of the severity and early-onset of coronary artery disease with variant rs1333049 on chromosome 9p21.3 polymorphism and the impact of this variant on cardiovascular risk factors in type 2 diabetic patients. The study population consisted of a control CAD group (101 patients) and 273 consecutive type 2 diabetic patients. Severity and extent of coronary atherosclerosis were scored numerically using the Gensini scoring system. The diabetic population was divided into three groups according to Gensini score: Group 1: no stenosis; Group 2: moderate CAD; Group 3, severe CAD. The homozygous CC genotype of rs1333049 was significantly associated with CAD in Group 2 (OR: 1.36; p = 0.02) and Group 3 (OR: 5.77, p < 0.001) compared to Group 1 (OR: 0.18; p = 0.2) and control group (OR: 0.22; p = 0.21). Among diabetic patients with early-onset CAD, CC genotype carriers had significantly higher Gensini scores than non-CC genotype carriers (49 ± 21.3 versus 14.87 ± 25.22; p < 0.001). The homozygous CC genotype of rs1333049 confers a magnified risk of early-onset and severe CAD in type 2 diabetic Tunisian population.