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2. SummaryB cell-based cellular vaccines represent a promising approach to active immunotherapy of cancer complementing the use of dendritic cells, especially in pediatric patients and patients with low bone marrow reserves. B cells can be easily prepared in large numbers and readily home to secondary lymphoid organs, the primary site of induction of cytotoxic T lymphocyte (CTL) responses. However, most B cell-based vaccines tested so far failed to induce functional and protective CTLs in in vivo models. Here we demonstrate that B cells activated via the Toll like receptor-9 (TLR-9) and CD40 up-regulate surface expression of MHC and costimulatory molecules, produce IL-12, and exhibit potent antigen-presenting properties in vitro. Importantly, while administration of peptide-coated or transiently transfected B cells fails to induce immune responses, therapeutic immunization with low numbers of genetically modified B cells stably expressing antigen results in an induction of functional CTLs and protection against the growth of tumor in an animal model. Following activation, B cells partially loose their ability to home to organized lymphoid tissue due to the shedding of CD62L; however, this property can be restored by expression of protease-resistant mutant of CD62L. In summary, the data presented in this report suggest that genetically modified activated B cells represent a promising candidate for a cancer vaccine eliciting functional systemic CTLs.

Harnessing the ability of cytotoxic T lymphocytes (CTLs) to recognize and eradicate tumor or pathogen-infected cells is a critical goal of modern immune-based therapies. Although multiple immunization strategies efficiently induce high levels of antigen-specific CTLs, the initial increase is typically followed by a rapid contraction phase resulting in a sharp decline in the frequency of functional CTLs. We describe a novel approach to immunotherapy based on a transplantation of low numbers of antigen-expressing hematopoietic stem cells (HSCs) following nonmyeloablative or partially myeloablative conditioning. Continuous antigen presentation by a limited number of differentiated transgenic hematopoietic cells results in an induction and prolonged maintenance of fully functional effector T cell responses in a mouse model. Recipient animals display high levels of antigen-specific CTLs four months following transplantation in contrast to dendritic cell-immunized animals in which the response typically declines at 4–6 weeks post-immunization. Majority of HSC-induced antigen-specific CD8+ T cells display central memory phenotype, efficiently kill target cells in vivo, and protect recipients against tumor growth in a preventive setting. Furthermore, we confirm previously published observation that high level engraftment of antigen-expressing HSCs following myeloablative conditioning results in tolerance and an absence of specific cytotoxic activity in vivo. In conclusion, the data presented here supports potential application of immunization by limited transplantation of antigen-expressing HSCs for the prevention and treatment of cancer and therapeutic immunization of chronic infectious diseases such as HIV-1/AIDS.

Key Worde: Peripheral blood stem cell Mobilizer . Dextran sulfate CFU-GM CFU-GEMM BFU-E. AbstractThe munbers of MNC and CFU-GM in peripheral blood began to rise I hour after intravenous injection of DS and achieved peak value within 3-5 hours. The toxicity test suggested that the critical drawback of DS for clinical application was hypersensitive reaction rather than L D toxicity. The threshold molecular weight for causing hypersensitive reaction was slightly higher than 10000D. Compared to other kind of mobilizers, such as Cy, Ara-C, DNR, etc., DS may not only be used in patients, but also be used in normal individuals. The safe and effective dose of DS for human being is about 15-20 mg/ kg.

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