Eric Vivès

Cell-penetrating and cell-targeting peptides in drug delivery

During the last decade, the potential of peptides for drug delivery into cells has been highlighted by the discovery of several cell-penetrating peptides (CPPs). CPPs are very efficient in delivering various molecules into cells. However, except in some specific cases, their lack of cell specificity remains the major drawback for their clinical development. At the same time, various peptides with specific binding activity for a given cell line (cell-targeting peptides) have also been reported in the literature. One of the goals of the next years will be to optimize the tissue and cell delivery of therapeutic molecules by means of peptides which combine both targeting and internalization advantages. In this review, we describe the main strategies that are currently in use or likely to be employed in the near future to associate both targeting and delivery properties.

Cell membrane is a more sensitive target than cytoplasm to dense ionization produced by auger electrons.

Abstract Pouget, J-P., Santoro, L., Raymond, L., Chouin, N., Bardiès, M., Bascoul-Mollevi, C., Huguet, H., Azria, D., Kotzki, P-O., Pè legrin, M., Vivès, E. and Pèlegrin, A. Cell Membrane is a More Sensitive Target than Cytoplasm to Dense Ionization Produced by Auger Electrons. Radiat. Res. 170, 192–200 (2008). To improve radioimmunotherapy with Auger electron emitters, we assessed whether the biological efficiency of 125I varied according to its localization. A-431 and SK-OV-3 carcinoma cells were incubated with increasing activities (0–4 MBq/ml) of 125I-labeled vectors targeting the cell membrane, the cytoplasm or the nucleus. We then measured cell survival by clonogenic assay and the mean radiation dose to the nucleus by assessing the cellular medical internal radiation dose (MIRD). The relationship between survival and the radiation dose delivered was investigated with a linear mixed regression model. For each cell line, we obtained dose–response curves for the three targets and the reference values (i.e., the dose leading to 75, 50 or 37% survival). When cell survival was expressed as a function of the total cumulative decays, nuclear 125I disintegrations were more harmful than disintegrations in the cytoplasm or at the cell membrane. However, when survival was expressed as a function of the mean radiation dose to the nucleus, toxicity was significantly higher when 125I was targeted to the cell membrane than to the cytoplasm. These findings indicate that the membrane is a more sensitive target than the cytoplasm for the dense ionization produced by Auger electrons. Moreover, cell membrane targeting is as cytotoxic as nuclear targeting in SK-OV-3 cells. We suggest that targeting the membrane rather than the cytoplasm may contribute to the development of more efficient radioimmunotherapies based on Auger electron radiation, also because most of the available vectors are directed against cell surface antigens.

Impact of the guanidinium group on hybridization and cellular uptake of cationic oligonucleotides.

The grafting of cationic groups to synthetic oligonucleotides (ONs) in order to reduce the charge repulsion between the negatively charged strands of a duplex or triplex, and consequently to increase a complexs stability, has been extensively studied. Guanidinium groups, which are highly basic and positively charged over a wide pH range, could be an efficient ON modification to enhance their affinity for nucleic acid targets and to improve cellular uptake. A straightforward post‐synthesis method to convert amino functions attached to ONs (on sugar, nucleobase or backbone) into guanidinium tethers has been perfected. In comparison to amino groups, such cationic groups anchored to α‐oligonucleotide phosphoramidate backbones play important roles in duplex stability, particularly with RNA targets. This high affinity could be explained by dual recognition resulting from Watson–Crick or Hoogsteen base pairing combined with cationic/anionic backbone recognition between strands involving H‐bond formation and salt bridging. Molecular‐dynamics simulations corroborate interactions between the cationic backbones of the α‐ONs and the anionic backbones of the nucleic acid targets. Moreover, ONs with guanidinium modification increased cellular uptake relative to negatively charged ONs. The cellular localization of these new cationic phosphoramidate ONs is mainly cytoplasmic. The uptake of these ON analogues might occur through endocytosis.

General overview of radioimmunotherapy of solid tumors.

Radioimmunotherapy (RIT) represents an attractive tool for the treatment of local and/or diffuse tumors with radiation. In RIT, cytotoxic radionuclides are delivered by monoclonal antibodies that specifically target tumor-associated antigens or the tumor microenvironment. While RIT has been successfully employed for the treatment of lymphoma, mostly with radiolabeled antibodies against CD20 (Bexxar(®); Corixa Corp., WA, USA and Zevalin(®); Biogen Idec Inc., CA, USA and Schering AG, Berlin, Germany), its use in solid tumors is more challenging and, so far, few trials have progressed beyond Phase II. This review provides an update on antibody-radionuclide conjugates and their use in RIT. It also discusses possible optimization strategies to improve the clinical response by considering biological, radiobiological and physical features.

Uptake and quantification of intracellular concentration of lipophilic pro-oligonucleotides in HeLa cells.

Several lipophilic prodrugs of oligonucleotides (T12 and T20) bearing enzymolabile protecting groups and labeled with fluorescein were synthesized. Their cellular uptake was studied by three different approaches using fluorescence: fluorescence microscopy, flow cytometry and spectrofluorometry. The corresponding prooligonucleotides (pro-oligos) were rapidly and efficiently taken up by HeLa cells and were found homogeneously in the cytoplasm and in the nucleus. The uptake was proportional to their relative lipophilicity and likely proceeded through a passive diffusion mechanism. Uptake followed a dose-response curve. This prooligo approach led to a 2-log increase of uptake in comparison with a T20 phosphorothioate oligonucleotide. Finally, an intracellular concentration of pro-oligo was estimated between 4 and 6 microM for an external concentration of 1 microM and up to 27 microM for an incubation at 10 microM.

Peptides in receptor-mediated radiotherapy: from design to the clinical application in cancers

Short peptides can show high affinity for specific receptors overexpressed on tumor cells. Some of these are already used in cancerology as diagnostic tools and others are in clinical trials for therapeutic applications. Therefore, peptides exhibit great potential as a diagnostic tool but also as an alternative or an additional antitumoral approach upon the covalent attachment of a therapeutic moiety such as a radionuclide or a cytotoxic drug. The chemistry offers flexibility to graft onto the targeting-peptide either fluorine or iodine directly, or metallic radionuclides through appropriate chelating agent. Since short peptides are straightforward to synthesize, there is an opportunity to further improve existing peptides or to design new ones for clinical applications. However, several considerations have to be taken into account to optimize the recognition properties of the targeting-peptide to its receptor, to improve its stability in the biological fluids and its residence in the body, or to increase its overall therapeutic effect. In this review, we highlight the different aspects which need to be considered for the development of an efficient peptide receptor-mediated radionuclide therapy in different neoplasms.

Cyclization of Peptides through a Urea Bond: Application to the Arg‐Gly‐Asp Tripeptide

Various synthetic cyclopeptides bind different cellular proteins with high affinity and specificity. In this study, we designed a new series of cyclic tetrapeptides containing the RGD sequence, a ligand for the αvβ3 integrin receptor, in which the ring closure was performed through a urea bond between the α‐amino group of the peptide and either the α‐ or the ε‐amino group of an additional lysine. Interestingly, we showed that the urea‐closed peptide had a higher affinity for αvβ3 receptors than a reference pentacyclopeptide. Moreover, the synthetic strategy allows coupling of the resulting cyclic tetrapeptide through the carboxylic acid moiety of its lysine residue to fluorescent molecules or drugs. In addition, this strategy could be easily adapted for the cyclization of any other peptides.