Jean-Serge Remy

Recently, it has been shown that the efficiency of antitumoral drugs can be enhanced when combined with therapeutic siRNAs. In the present study, an original platform based on polydiacetylenic micelles containing a cationic head group able to efficiently deliver a small interfering RNA (siRNA) targeting the PLK-1 gene while offering a hydrophobic environment for encapsulation of lipophilic drugs such as camptothecin is developed. We demonstrate that the co-delivery of these two agents with our micellar system results in a synergistic tumor cell killing of cervical and breast cancer cell lines in vitro. The combined drugs are active in a subcutaneous in vivo cancer model. Altogether, the results show that our nanometric micellar delivery system can be used for the development of new drug–siRNA combo-therapies.

Polydiacetylenic nanofibers (PDA-Nfs) obtained by photopolymerization of surfactant 1 were optimized for intracellular delivery of small interfering RNAs (siRNAs). PDA-Nfs/siRNA complexes efficiently silenced the oncogene Lim-1 in the renal cancer cells 786-O in vitro. Intraperitoneal injection of PDA-Nfs/siLim1 downregulated Lim-1 in subcutaneous tumor xenografts obtained with 786-O cells in nude mice. Thus, PDA-Nfs represent an innovative system for in vivo delivery of siRNAs.

Oligospermine–siRNA conjugates are able to induce efficient luciferase gene silencing upon carrier-free transfection. These conjugates are readily accessible by a versatile automated chemistry that we developed using a DMT-spermine phosphoramidite reagent. In this article, we used this chemistry to study a wide range of structural modifications of the oligospermine–siRNA conjugates, i.e., variation of conjugate positions and introduction of chemical modifications to increase nuclease resistance. At first we examined gene silencing activity of a series of siRNA–tris(spermine) conjugates with and without chemical modifications in standard carrier assisted conditions. The three spermine units attached at one of the two ends of the sense strand or at the 3′-end of the antisense strand are compatible with gene silencing activity whereas attachment of spermine units at the 5′-end of the antisense strand abolished the activity. 2′-O-Methylated nucleotides introduced in the sense strand are compatible while not in the antisense strand. Thiophosphate links could be used without activity loss at the 3′-end of both strands and at the 5′-end of the sense strand to conjugate oligospermine. Consequently a series of oligospermine–siRNA conjugates containing 15 to 45 spermines units in various configurations were chosen, prepared, and examined in carrier-free conditions. Attachment of 30 spermine units singly at the 5′-end of the sense strand provides the most potent carrier-free siRNA. Longevity of luciferase gene silencing was studied using oligospermine–siRNA conjugates. Five day long efficiency with more than 80% gene expression knockdown was observed upon transfection without vector. Oligospermine–siRNA conjugates targeting cell-constitutive natural lamin A/C gene were prepared. Efficient gene silencing was observed upon carrier-free transfection of siRNA conjugates containing 20 or 30 spermine residues grafted at the 5′-end of the sense strand.

PH-Sensitive linkers designed to undergo selective hydrolysis at acidic pH compared to physiological pH can be used for the selective release of therapeutics at their site of action. In this paper, the hydrolytic cleavage of a wide variety of molecular structures that have been reported for their use in pH-sensitive delivery systems was examined. A wide variety of hydrolytic stability profiles were found among the panel of tested chemical functionalities. Even within a structural family, a slight modification of the substitution pattern has an unsuspected outcome on the hydrolysis stability. This work led us to establish a first classification of these groups based on their reactivities at pH 5.5 and their relative hydrolysis at pH 5.5 vs. pH 7.4. From this classification, four representative chemical functions were selected and studied in-vitro. The results revealed that only the most reactive functions underwent significant lysosomal cleavage, according to flow cytometry measurements. These last results question the acid-based mechanism of action of known drug release systems and advocate for the importance of an in-depth structure-reactivity study, using a tailored methodology, for the rational design and development of bio-responsive linkers.

The use of multivalent carbohydrate compounds to block cell-surface lectin receptors is a promising strategy to inhibit the entry of pathogens into cells and could lead to the discovery of novel antiviral agents. One of the main problems with this approach, however, is that it is difficult to make compounds of an adequate size and multivalency to mimic natural systems such as viruses. Hexakis adducts of [60]fullerene are useful building blocks in this regard because they maintain a globular shape at the same time as allowing control over the size and multivalency. Here we report water-soluble tridecafullerenes decorated with 120 peripheral carbohydrate subunits, so-called ‘superballs’, that can be synthesized efficiently from hexakis adducts of [60]fullerene in one step by using copper-catalysed azide–alkyne cycloaddition click chemistry. Infection assays show that these superballs are potent inhibitors of cell infection by an artificial Ebola virus with half-maximum inhibitory concentrations in the subnanomolar range.

Inhibition of excessive Toll-like receptor 4 (TLR4) signaling is a therapeutic approach pursued for many inflammatory diseases. We report that Mannoside Glycolipid Conjugates (MGCs) selectively blocked TLR4-mediated activation of human monocytes and monocyte-derived dendritic cells (DCs) by lipopolysaccharide (LPS). They potently suppressed pro-inflammatory cytokine secretion and maturation of DCs exposed to LPS, leading to impaired T cell stimulation. MGCs did not interfere with LPS and could act in a delayed manner, hours after LPS stimulation. Their inhibitory action required both the sugar heads and the lipid chain, although the nature of the sugar and the structure of the lipid tail could be modified. They blocked early signaling events at the cell membrane, enhanced internalization of CD14 receptors, and prevented colocalization of CD14 and TLR4, thereby abolishing NF-κB nuclear translocation. When the best lead conjugate was tested in a mouse model of LPS-induced acute lung inflammation, it displayed an anti-inflammatory action by suppressing the recruitment of neutrophils. Thus, MGCs could serve as promising leads for the development of selective TLR4 antagonistic agents for inflammatory diseases.

PH-sensitive linkers designed to undergo selective hydrolysis at acidic pH compared to physiological pH can be used for selective release of therapeutics selectively at targets and orthoesters have been demonstrated to be good candidates for such linkers. Following an HPLC screening, a Spiro Diorthoester (SpiDo) derivative was identified as a potent acid-labile group for the development of pH-sensitive targeted systems. After incorporation of this linker into activatable FRET-based probe and side-by-side comparison to a well-known alkylhydrazone linker, this SpiDo linker has shown a fast and pH sensitive hydrolysis for mild acidic conditions, a pH sensitive lysosomal hydrolysis, and high stability in human plasma.