Restoring Anticancer Immune Response by Targeting Tumor-Derived Exosomes With a HSP70 Peptide Aptamer.

BACKGROUND: Exosomes, via heat shock protein 70 (HSP70) expressed in their membrane, are able to interact with the toll-like receptor 2 (TLR2) on myeloid-derived suppressive cells (MDSCs), thereby activating them. METHODS: We analyzed exosomes from mouse (C57Bl/6) and breast, lung, and ovarian cancer patient samples and cultured cancer cells with different approaches, including nanoparticle tracking analysis, biolayer interferometry, FACS, and electron microscopy. Data were analyzed with the Student's t and Mann-Whitney tests. All statistical tests were two-sided. RESULTS: We showed that the A8 peptide aptamer binds to the extracellular domain of membrane HSP70 and used the aptamer to capture HSP70 exosomes from cancer patient samples. The number of HSP70 exosomes was higher in cancer patients than in healthy donors (mean, ng/mL ± SD = 3.5 ± 1.7 vs 0.17 ± 0.11, respectively, P = .004). Accordingly, all cancer cell lines examined abundantly released HSP70 exosomes, whereas "normal" cells did not. HSP70 had higher affinity for A8 than for TLR2; thus, A8 blocked HSP70/TLR2 association and the ability of tumor-derived exosomes to activate MDSCs. Treatment of tumor-bearing C57Bl/6 mice with A8 induced a decrease in the number of MDSCs in the spleen and inhibited tumor progression (n = 6 mice per group). Chemotherapeutic agents such as cisplatin or 5FU increase the amount of HSP70 exosomes, favoring the activation of MDSCs and hampering the development of an antitumor immune response. In contrast, this MDSC activation was not observed if cisplatin or 5FU was combined with A8. As a result, the antitumor effect of the drugs was strongly potentiated. CONCLUSIONS: A8 might be useful for quantifying tumor-derived exosomes and for cancer therapy through MDSC inhibition.

Heat shock proteins in hematopoietic malignancies.

Inducible heat shock proteins are molecular chaperones whose expression is increased after many different types of stress. They have a protective function helping the cell to cope with lethal conditions. Their basal expression is low in nonstressed, normal and nontransformed cells. However, in cancer cells and particularly in hematological malignancies, they are surprisingly abundant. Malignant cells have to rewire their metabolic requirements and therefore have a higher need for chaperones. This cancer cell addiction for HSPs is the basis for the use of HSP inhibitors in cancer therapy. HSPs have been shown to interact with different key apoptotic proteins. As a result, HSPs can essentially block the apoptotic pathways at several steps, most of them involving the activation of cystein proteases called caspases. Apoptosis and differentiation are physiological processes that share many common features, for instance, a controlled caspase activation and chromatin condensation are frequently observed. It is, therefore, not surprising that HSPs may be implicated in the differentiation process. HSPs may determine the fate of the cells by orchestrating the decision of apoptosis versus differentiation. This review will focus on the role of HSPs in hematological malignancies and the emerging therapeutic options that are being either proposed or used to target these protective proteins.

Heat shock proteins as danger signals for cancer detection.

First discovered in 1962, heat shock proteins (HSPs) are highly studied with about 35,500 publications on the subject to date. HSPs are highly conserved, function as molecular chaperones for a large panel of "client" proteins and have strong cytoprotective properties. Induced by many different stress signals, they promote cell survival in adverse conditions. Therefore, their roles have been investigated in several conditions and pathologies where HSPs accumulate, such as in cancer. Among the diverse mammalian HSPs, some members share several features that may qualify them as cancer biomarkers. This review focuses mainly on three inducible HSPs: HSP27, HPS70, and HSP90. Our survey of recent literature highlights some recurring weaknesses in studies of the HSPs, but also identifies findings that indicate that some HSPs have potential as cancer biomarkers for successful clinical applications.

Turbidity diagrams of polyanion/polycation complexes in solution as a potential tool to predict the occurrence of polyelectrolyte multilayer deposition.

Surface functionalization with polyelectrolyte multilayer films (PEM films) has become very popular owing to its simplicity and versatility. However, even if some research is already available, this field of surface chemistry lacks a systematic knowledge of how the polyelectrolyte structure and solution conditions influence the growth of PEM films. In this investigation, we focus on the possible relationship between turbidity of polycation and polyanion mixtures in solution, and the buildup of PEM films made from the same polyelectrolytes in the same physicochemical conditions, namely pH, temperature and ionic strength. It comes out that for six different polycation/polyanion combinations there is a clear correlation between the turbidity evolution of polycation/polyanion complexes with the salt concentration and the evolution of the film deposition with the same parameter. In this investigation, the complexes in solution were prepared in conditions where the ratio between the number of cationic to anionic groups was close to unity. Even if there is a correlation between turbidity in solution and PEM film deposition, we found some exceptions in the low salt concentration regime. This work is an extension of the preliminary works of Cohen Stuart (D. Kovacevic et al. Langmuir 18 (2002) 5607-5612) and Sukishvili et al. (S.A. Sukhishvili, E. Kharlampieva and V. Izumrudov, Macromolecules 39 (2006) 8873-8881).

Micro-stratified architectures based on successive stacking of alginate gel layers and poly(l-lysine)-hyaluronic acid multilayer films aimed at tissue engineering.

A micro-stratified 3D scaffold was designed by successive stacking of alginate gel layers (AGLs) and poly(l-lysine)-hyaluronic acid (PLL-HA) multilayer films. AGLs are obtained by complexation of alginate by Ca2+ ions. Alginate solutions are first sprayed onto a solid substrate inclined such that the excess of solution be removed by natural drainage. A CaCl2 solution is then either sprayed onto the substrate or the alginate covered substrate is dipped into a CaCl2 solution. The spraying of the CaCl2 solution leads to micro-porous AGLs, whereas the dipping in a CaCl2 aqueous solution leads to a more homogeneous gel layer without porosity. The second process also allows the formation of AGLs with a controlled thickness. With the goal of stacking different AGLs and PLL-HA films, the influence of a PLL-HA precursor film on the formation of AGLs is firstly investigated. It is found that when an alginate solution is sprayed on a PLL-HA multilayer built in the presence of CaCl2, the multilayer plays the role of reservoir of Ca2+ ions and of PLL chains, which both diffuse out of the multilayer film and complex alginate chains. This leads to the formation of a "pre-alginate gel". When this film is further dipped in the CaCl2 solution, an additional AGL forms, which is, however, free of PLL chains. Finally after the build-up of a PLL-HA film on the top of AGL, we succeeded in designing micro-stratified 3D scaffolds constituted by alternating strata of AGLs and PLL-HA films. This micro-stratified gel provides a new scaffold design with a perfectly controlled build-up: AGL aims to be a 3D scaffold for cell culture, and the PLL-HA multilayers should act as reservoirs for biologically active molecules.

Cell apoptosis control using BMP4 and noggin embedded in a polyelectrolyte multilayer film.

Programmed cell death (apoptosis) is a genetically regulated process of cell elimination essential during development. During development, programmed cell death is involved in the specific shaping of organs, in the elimination of cells having achieved their program, and in regulating the number of cells to differentiate. Tooth development includes these three aspects and was used here as a model to study the control of apoptosis. Bone morphogenetic proteins (BMPs) are currently considered as playing a major role in signaling apoptosis. This apoptosis could be stopped by treatments with a BMP antagonist ("Noggin"). We selected a model system made by a layer-by-layer approach using poly-L-glutamic acid (PlGA) and poly-L-lysine (PlL) films into which BMP4 and/or Noggin have been embedded. Our results indicate that in situ control of apoptosis during tooth differentiation mediated by both BMP4 and Noggin embedded in a polyelectrolyte multilayer film is possible. We show here for the first time that in the presence of BMP4 and Noggin embedded in a multilayered film, we can induce or inhibit cell death in tooth differentiation, and conserve their biological effects.