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Targeting on the amyloid-β (Aβ) is a promising immunotherapeutic strategy for Alzheimer's disease (AD). However, Aβ(1-15) sequence alone induces low antibody response and poor protection against AD. We describe here the immunological characterization and protective efficacy of several recombinant chimeric vaccines with hexavalent foldable Aβ(1-15) (6Aβ15) fused to PADRE or toxin-derived carrier proteins. Immunization with these chimeric antigens generated robust Th2 immune responses with high anti-Aβ42 antibody titers in different mice, which recognized neurotoxic Aβ42 oligomers, but did not stimulate Aβ42-specific T cell responses. These 6Aβ15 chimeric vaccines markedly reduced Aβ pathology and prevented development of behavioral deficits in immunized older AD mice. Importantly, toxin-derived carrier proteins as molecular adjuvants of chimeric vaccines could substantially boost immune responses and overcome Aβ- and old age-associated hypo-responsiveness, and elicit long-term Aβ-specific antibody response, which in turn inhibited Aβ-mediated pathology and improved acquisition and retention of spatial memory in immunized AD mice. These data indicate that toxin fragments as molecular adjuvants are promising new tools for the rational design and development of prototype chimeric vaccines for AD and this type of chimeric vaccine design has the added advantage of overcoming hypo-responsiveness in elderly AD patients with pre-existing memory Th cells from tetanus toxin.

// Dadi Zeng 1, * , Qiang Sun 1, * , Ang Chen 1 , Jiangfeng Fan 1 , Xiaopeng Yang 1 , Lei Xu 1 , Peng Du 1 , Weiyi Qiu 1 , Weicai Zhang 1 , Shuang Wang 1 , Zhiwei Sun 1 1 Beijing Institute of Biotechnology, Fengtai District, Beijing 100071, China * These two authors contributed equally to this work Correspondence to: Zhiwei Sun, email: szwyhhh@aliyun.com Shuang Wang, email: 18910810680@163.com Weicai Zhang, email: drzhangweicai@163.com Keywords: CD47, anti-CD47 antibody, phagocytosis, cancer therapy, cell-in-cell Received: March 17, 2016 Accepted: October 17, 2016 Published: November 15, 2016 ABSTRACT CD47/SIRPα interaction serves as an immune checkpoint for macrophage-mediated phagocytosis. Mouse anti-CD47 blocking antibodies had demonstrated potent efficacy in the treatment of both leukemic and solid tumors in preclinical experimentations, and therefore had moved forward rapidly into clinical trials. However, a fully human blocking antibody, which meets clinical purpose better, has not been reported for CD47 up to date. In this study, we reported the isolation of a fully human anti-CD47 blocking antibody, ZF1, from a phage display library. ZF1 displayed high specificity and affinity for CD47 protein, which were comparable to those for humanized anti-CD47 blocking antibody B6H12. Importantly, ZF1 treatment could induce robust, or even stronger than B6H12, phagocytosis of leukemic cancer cells by macrophage in vitro , and protect BALB/c nude mice from cancer killing by engrafted leukemic cells (CCRF and U937) to a similar extent as B6H12 did. Thus, these data provide primary early pre-clinical support for the development of ZF1 as a fully human blocking antibody to treat human leukemia by targeting CD47 molecule.

Active amyloid-beta (Aβ) immunotherapy is under investigation to prevent or treat Alzheimer disease (AD). We describe here the immunological characterization and protective effect of DNA epitope chimeric vaccines using 6 copies of Aβ1-15 fused with PADRE or toxin-derived carriers. These naked 6Aβ15-T-Hc chimeric DNA vaccines were demonstrated to induce robust anti-Aβ antibodies that could recognize Aβ oligomers and inhibit Aβ oligomer-mediated neurotoxicity, result in the reduction of cerebral Aβ load and Aβ oligomers, and improve cognitive function in AD mice, but did not stimulate Aβ-specific T cell responses. Notably, toxin-derived carriers as molecular adjuvants were able to substantially promote immune responses, overcome Aβ-associated hypo-responsiveness, and elicit long-term Aβ-specific antibody response in 6Aβ15-T-Hc-immunized AD mice. These findings suggest that our 6Aβ15-T-Hc DNA chimeric vaccines can be used as a safe and effective strategy for AD immunotherapy, and toxin-derived carrier proteins are effective molecular adjuvants of DNA epitope vaccines for Alzheimer's disease.