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Chandran P.L.,U.S. National Institutes of Health | Dimitriadis E.K.,U.S. National Institutes of Health | Lisziewicz J.,EMMUNITY Inc. | Speransky V.,U.S. National Institutes of Health | Horkay F.,U.S. National Institutes of Health
Soft Matter | Year: 2014

Mannobiose-modified polyethylenimines (PEI) are used in gene therapy to generate nanoparticles of DNA that can be targeted to the antigen-presenting cells of the immune system. We report that the sugar modification alters the DNA organization within the nanoparticles from homogenous to shell-like packing. The depth-dependent packing of DNA within the nanoparticles was probed using AFM nano-indentation. Unmodified PEI-DNA nanoparticles display linear elastic properties and depth-independent mechanics, characteristic of homogenous materials. Mannobiose-modified nanoparticles, however, showed distinct force regimes that were dependent on indentation depth, with 'buckling'-like response that is reproducible and not due to particle failure. By comparison with theoretical studies of spherical shell mechanics, the structure of mannobiosylated particles was deduced to be a thin shell with wall thickness in the order of few nanometers, and a fluid-filled core. The shell-core structure is also consistent with observations of nanoparticle denting in altered solution conditions, with measurements of nanoparticle water content from AFM images, and with images of DNA distribution in Transmission Electron Microscopy. © the Partner Organisations 2014. Source

Toke E.R.,Genetic Immunity Kft | Toke E.R.,EMMUNITY Inc. | Lorincz O.,Genetic Immunity Kft | Lorincz O.,EMMUNITY Inc. | And 25 more authors.
Gene Therapy | Year: 2014

There is no clinically available cancer immunotherapy that exploits Langerhans cells (LCs), the epidermal precursors of dendritic cells (DCs) that are the natural agent of antigen delivery. We developed a DNA formulation with a polymer and obtained synthetic 'pathogen-like' nanoparticles that preferentially targeted LCs in epidermal cultures. These nanoparticles applied topically under a patch-elicited robust immune responses in human subjects. To demonstrate the mechanism of action of this novel vaccination strategy in live animals, we assembled a high-resolution two-photon laser scanning-microscope. Nanoparticles applied on the native skin poorly penetrated and poorly induced LC motility. The combination of nanoparticle administration and skin treatment was essential both for efficient loading the vaccine into the epidermis and for potent activation of the LCs to migrate into the lymph nodes. LCs in the epidermis picked up nanoparticles and accumulated them in the nuclear region demonstrating an effective nuclear DNA delivery in vivo. Tissue distribution studies revealed that the majority of the DNA was targeted to the lymph nodes. Preclinical toxicity of the LC-targeting DNA vaccine was limited to mild and transient local erythema caused by the skin treatment. This novel, clinically proven LC-targeting DNA vaccine platform technology broadens the options on DC-targeting vaccines to generate therapeutic immunity against cancer. © 2014 Macmillan Publishers Limited. Source

Kolonics A.,Hungarian Academy of Sciences | Kolonics A.,R and D Ultrafast Lasers Ltd | Csiszovszki Z.,Genetic Immunity Kft | Csiszovszki Z.,EMMUNITY Inc. | And 8 more authors.
Experimental Dermatology | Year: 2014

Epidermal Langerhans cells (LCs) function as professional antigen-presenting cells of the skin. We investigated the LC-targeting properties of a special mannose-moiety-coated pathogen-like synthetic nanomedicine DermaVir (DV), which is capable to express antigens to induce immune responses and kill HIV-infected cells. Our aim was to use multiphoton laser microscopy (MLM) in vivo in order to visualize the uptake of Alexa-labelled DV (AF546-DV) by LCs. Knock-in mice expressing enhanced green fluorescent protein (eGFP) under the control of the langerin gene (CD207) were used to visualize LCs. After 1 h, AF546-DV penetrated the epidermis and entered the eGFP-LCs. The AF546-DV signal was equally distributed inside the LCs. After 9 h, we observed AF546-DV signal accumulation that occurred mainly at the cell body. We demonstrated in live animals that LCs picked up and accumulated the nanoparticles in the cell body. © 2014 John Wiley & Sons A/S. Source

Verstrepen B.E.,Biomedical Primate Research Center | Oostermeijer H.,Biomedical Primate Research Center | Fagrouch Z.,Biomedical Primate Research Center | Van Heteren M.,Biomedical Primate Research Center | And 14 more authors.
PLoS ONE | Year: 2014

The mosquito-borne West Nile virus (WNV) causes human and animal disease with outbreaks in several parts of the world including North America, the Mediterranean countries, Central and East Europe, the Middle East, and Africa. Particularly in elderly people and individuals with an impaired immune system, infection with WNV can progress into a serious neuroinvasive disease. Currently, no treatment or vaccine is available to protect humans against infection or disease. The goal of this study was to develop a WNV-vaccine that is safe to use in these high-risk human target populations. We performed a vaccine efficacy study in non-human primates using the contemporary, pathogenic European WNV genotype 1a challenge strain, WNV-Ita09. Two vaccine strategies were evaluated in rhesus macaques (Macaca mulatta) using recombinant soluble WNV envelope (E) ectodomain adjuvanted with Matrix-M, either with or without DNA priming. The DNA priming immunization was performed with WNV-DermaVir nanoparticles. Both vaccination strategies successfully induced humoral and cellular immune responses that completely protected the macaques against the development of viremia. In addition, the vaccine was well tolerated by all animals. Overall, The WNV E protein adjuvanted with Matrix-M is a promising vaccine candidate for a non-infectious WNV vaccine for use in humans, including at-risk populations. Copyright: © 2014 Verstrepen et al. Source

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