In vitro studies of DC matured with TLR7/8 agonists, with or without poly (I:C) as a TLR3 agonist, resulted in substantial secretion of bioactive IL-12(p70) and high potential to activate innate and adaptive immune responses. We also have described DC maturation cocktails using quinoline-like molecules, R848 or CL075, in 3-day and 7-day mDC.
#NOD SCID GAMMA MICE TRIAL#
Encouraging results have also been obtained in the first clinical trial using TLR-stimulated DC as well as studies using TLR agonists as immune stimulatory adjuvants. The discovery that TLR agonists can optimally activate murine DC to secrete IL-12 led to studies of the impact of TLR agonists on human mDC cytokine production. Many clinical trials have utilized a four-component cocktail (4C) containing IL-1β, IL-6, TNF-α and PGE 2 for DC maturation. Published studies demonstrated that monocyte-derived DC generated over 2 or 3 days give comparable or enhanced immune responses in vitro compared to 7-day mDC. These mostly rely on a 6-day protocol using IL-4 and GM-CSF to induce immature DC (iDC), followed by a 24 h maturation phase. Various methods have been developed for preparation of monocyte-derived mDC for clinical studies. Monocyte-derived DC are commonly used in DC vaccine strategies. Many current cancer vaccines focus on mature DC (mDC) loaded with tumor-associated antigens (TAA) and injected intradermally to activate CD8 + cytotoxic T lymphocytes (CTL). In addition, ex vivo analyses of human CD3 + T cells recovered from the spleens of these mice are also possible, including studies on lymphocyte subsets, Th1/Th2 polarization, presence of regulatory T cells and the impact of DC vaccination on their functions.ĭendritic cell (DC) vaccines hold high therapeutic potential for induction of antitumor immunity in cancer patients. This humanized mouse model system enables comparisons among different DC vaccine types to be rapidly assessed in vivo. These findings led to a ranking of the DC vaccine effects in vivo that reflected the hierarchy previously found for these mature DC variations in vitro. Moreover, consistent with our in vitro observations, vaccination using mature DC activated with TLR3 and TLR7/8 agonists resulted in enhanced immune responses in vivo. Human monocyte-derived DC generated in vitro in 3 days induced better MART-1-specific immune responses in the autologous donor T cells present in the humanized NSG mice. After two weekly vaccinations, the splenocyte populations containing human lymphocytes were recovered 7 days later and assessed for MART-1-specific immune responses using MHC-multimer-binding assays and functional assessment of specific killing of melanoma tumor cell lines.
#NOD SCID GAMMA MICE FULL#
Three DC vaccine formulations were compared that varied generation time in vitro (3 days versus 7 days) and signals for maturation (with or without Toll-like receptor (TLR)3 and TLR7/8 agonists) using MART-1 as a surrogate antigen, by electroporating mature DC with in vitro transcribed RNA encoding full length protein. NSG recipients of human PBMC were engrafted over 14 days and then vaccinated twice with autologous DC via intravenous injection. This led to selection of a simple and rapid protocol for engraftment and vaccine evaluation that encompassed 4 weeks. Two reconstitution regimes of NOD/scid IL2Rg null (NSG) mice with adult human peripheral blood mononuclear cells (PBMC) were evaluated for engraftment using 4-week and 9-week schedules. Here we describe a humanized mouse model to assess the efficacy of various human dendritic cell (DC) preparations. However, the ability to carry out comparisons of new cellular vaccines in vivo would be of substantial interest for design of clinical studies. To date very few systems have been described for preclinical investigations of human cellular therapeutics in vivo.
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