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

描述（申请人提供）：这项提案的总体目标是建立在我们对收藏的理解基础上，从离体牙齿中回收的干/祖细胞的处理和低温生物学特性，这些数据的最终结果是开发用于研究和/或潜在的移植或其他临床用途，作为根据现行组织质量管理规范（cGTP）处理的人细胞产品，如21 CFR 1271所述。最近研究这种令人兴奋的出生后干细胞来源的研究已经鉴定了来源于酶消化的牙髓组织的克隆形成和高度增殖的细胞群（Gronthos等人，2000;佩里等人，2008年）。这些研究的结果表明，这些细胞可能具有用于干细胞介导的治疗以及组织工程应用的潜力（Seo et al.，2005年）的报告。为了进一步研究和潜在的治疗用途，这些细胞，我们的小组已经开始评估这些细胞和它们各自的组织的起源生存冷冻保存处理的能力。理想情况下，最佳的冷冻保存过程将是直接和有效的，当应用于组织作为一个整体，与思想，干细胞可以提取解冻后。这样做的基本原理是保存临床样品用于随后的干细胞回收，因为可以合理地推测，在临床中冷冻保存组织将比直接初级分离干细胞更实用，直接初级分离干细胞将需要额外的设备和人员（Seo等人，2005;佩里等人，2008年）。这些细胞的冷冻库将通过允许将细胞运送给研究者和/或临床医生以及足够的时间用于供体表征和/或潜在测试等来为其使用增加极大的灵活性。为了实现这一总体目标，我们提出了以下具体目标：（I）最终开发优化的cGTP方案，用于开发用于分配和使用的牙齿干细胞库;（II）最终开发用于牙齿组织冷冻库的cGTP方案，用于以后的干细胞/祖细胞分离和扩增;以及（III）开发和验证用于包装和分配临床使用的细胞的封闭容器。该提案的最终目标是DPSC或组织的冷冻库，与未冷冻的对应物相比，基于可用的最灵敏的测定，具有e90%一致的细胞功能恢复。对于特定目标III，目标将是FDA批准的包装系统，可用于临床利用牙齿来源的干细胞/祖细胞。该系统也将用于需要较小体积的其他临床相关干细胞来源。 公共卫生相关性：干细胞可以发育成体内不同的组织类型。“出生后”或“成人”干细胞提供了从胚胎中获得干细胞的替代方案。这种干细胞存在于人类牙齿中，但在这些细胞直接用于临床之前还需要更多的研究。一个冷冻的牙齿干细胞库将是促进这项研究的理想选择。这个拟议的项目将优化处理和冷冻这些细胞的方法，以便于研究，测试和最终临床使用，并提供储存来自自己牙齿的组织的能力，以便在需要时使用自己的细胞。

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.