A NOVEL APPROACH TO GMP PRODUCTION, CRYOPRESERVATION AND DISTRIBUTION OF TOOTH DE

描述（由申请人提供）：本提案的总体目标是建立在我们对从拔牙中恢复的干细胞/祖细胞的收集、处理和低温生物学特性的理解的基础上，通过这些数据的最终结果，开发牙髓来源干细胞（DPSCs）冷冻库，用于研究和/或潜在移植或其他临床应用，作为根据21 CFR 1271中描述的当前良好组织规范（cGTP）处理的人类细胞产品。最近的研究对这种令人兴奋的产后干细胞来源进行了调查，发现了一群来自酶消化牙髓组织的克隆性和高度增殖细胞（Gronthos等人，2000；Perry等人，2008）。这些研究的结果表明，这些细胞可能有潜力用于干细胞介导的治疗以及组织工程应用（Seo et al., 2005）。为了进一步研究和潜在的治疗应用这些细胞，我们的团队已经开始评估这些细胞和它们各自的起源组织在低温保存处理中存活的能力。理想情况下，最佳的冷冻保存过程将是直接有效的，当应用于整个组织时，干细胞可以在解冻后提取。这样做的理由是为了保存临床样本，以便随后的干细胞恢复，因为有理由推测，在临床中对组织进行低温保存将比直接分离干细胞更实用，因为直接分离干细胞需要额外的设备和人员（Seo等人，2005；Perry等人，2008）。这些细胞的冷冻银行将为它们的使用增加极大的灵活性，除其他外，允许将细胞运送给研究人员和/或临床医生，并有足够的时间进行供体鉴定和/或潜在测试。为了实现这一总体目标，我们提出以下具体目标：(I)最终开发优化的cGTP协议，以开发用于分配和使用的牙干细胞库；（II）牙齿组织冷冻库的最终开发cGTP方案，用于后期干细胞/祖细胞的分离和扩增；（三）开发和验证用于临床使用的细胞包装和分发的封闭容器。该提案的最终目标是建立一个冷冻干细胞或组织库，根据最灵敏的检测方法，与未冷冻的组织相比，其细胞功能恢复率达到90%。Specific Aim III的目标是获得FDA批准的包装系统，用于临床使用来自牙齿的干细胞/祖细胞。该系统也将用于其他临床相关的干细胞来源，其中需要较小的体积。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to build on our understanding of the collection, processing and cryobiological characteristics of stem/progenitor cells recovered from extracted teeth with the ultimate culmination of these data to develop a cryobank of dental pulp derived stem cells (DPSCs) for research and/or potential transplant or other clinical utilization as a human cell product processed under current good tissue practice (cGTP) as described under 21 CFR 1271. Recent studies investigating this exciting source of post-natal stem cells have identified a population of clonogenic and highly proliferative cells derived from enzymatically digested dental pulp tissue (Gronthos et al., 2000; Perry et al., 2008). Results from these studies have indicated that these cells may have the potential to be utilized in stem cell mediated therapies as well as tissue engineering applications (Seo et al., 2005). To allow further study and potential therapeutic use of these cells, our group has begun to evaluate the ability of these cells and their respective tissues of origin to survive cryopreservation processing. Ideally, the optimum cryopreservation process will be straight forward and effective when applied to the tissue as a whole, with the idea that stem cells could be extracted post-thaw. The rationale for this would be to preserve clinical samples for subsequent stem cell recovery as it is reasonable to speculate that cryopreservation of tissues in the clinic will be more practical than direct primary isolation of stem cells, which would require additional equipment and personnel (Seo et al., 2005; Perry et al., 2008). A cryobank of these cells would add great flexibility to their use by allowing, among other things, shipment of cells to investigators and/or clinicians and adequate time for donor characterization and/or potential testing. To achieve this overall goal, we propose the following Specific Aims: (I) Final development of optimized, cGTP protocols for development of a tooth stem cell bank for distribution and use; (II) Final development cGTP protocols for cryobanking of tooth tissue for later stem/progenitor cell isolation and expansion; and (III) Development and validation of a closed container for packaging and distribution of cells for clinical use. The ultimate goal of the proposal is a cryobank of DPSCs or tissue with e90% consistent functional recovery of cells based on the most sensitive assays available when compared to their unfrozen counterparts. For Specific Aim III the goal will be an FDA approved packaging system useful for clinical utilization of stem/progenitor cells derived from teeth. This system will also be useful for other clinically relevant stem cell sources where smaller volumes are required. PUBLIC HEALTH RELEVANCE: Stem cells can develop into different tissues types in the body. "Post-natal" or "adult" stem cells offer an alternative to obtaining stem cells from embryos. Such stem cells exist in human teeth, but more research is required before these cells can have direct clinical use. A bank of frozen stem cells from teeth would be ideal to facilitate this research. This proposed project would optimize methods for processing and freezing these cells for ease of distribution for research, testing, and eventually clinical use, as well as present the ability for one to store tissue from their own teeth for potential use of their own cells if/when needed.