Develop novel assays for assessing cellular and gene therapies

细胞和基因治疗产品必须进行无菌性、稳定性、纯度和效力测试。此外，对临床细胞治疗产品的同一性、一致性和可比性进行测试也很重要。 测试细胞和基因疗法具有挑战性。这些疗法通常是从一个人那里收集的，因此可用于测试的材料数量有限。它们通常在生产后立即或不久输血，因此完成测定的时间非常有限。这些疗法中的许多是具有多种功能的复杂细胞。对这些疗法的临床有效性至关重要的细胞功能通常是未知的。传统上，诸如流式细胞术、ELISA、ELISPOT和细胞培养的分析测定已用于分析细胞和基因疗法。虽然这些测定已被证明是非常有用的，但可以用这些测定分析的因素的数量和类型是有限的。 我们一直在研究使用基因和微小RNA表达分析细胞疗法。这些测定可能仅需要使用少量细胞，并且可用于评估整个转录组的表达。我们一直在测试全局基因和微小RNA表达谱的能力，以确定这些测定法用于评估细胞疗法的稳定性、纯度和效力的实用性。我们已经表明，基因表达谱可以检测储存细胞的变化，并检测外周血白细胞（T细胞，B细胞和单核细胞）和造血干细胞之间的差异。基因表达谱分析还能够检测未成熟和成熟树突状细胞（DC）之间的差异，并且可用于比较使用成熟剂的不同组合产生的成熟DC。我们将很快开始分析临床DC产品，以比较基因表达与临床结果，以鉴定可能用于一致性、稳定性、同一性和效价测试的生物标志物。 骨髓基质细胞（BMSC）被用于调节免疫系统和许多再生医学应用，包括骨修复。骨髓间充质干细胞是一种异质性细胞群，少数骨髓间充质干细胞是骨骼干细胞，对再生医学的临床应用非常重要。我们一直在研究BMSCs以确定识别骨骼干细胞的标志物。 我们正在使用单细胞分析和下一代DNA测序来识别骨骼干细胞的标记。 嵌合抗原受体（CAR）T细胞被用于治疗许多血液恶性肿瘤，然而，这些疗法的接受者之间的临床结果不同，其中一些变异性可能是由于变异性，因此CAR T细胞产品之间的效力差异。 我们正在使用基因表达分析，mRNA分析，单细胞分析和下一代测序来确定与这些细胞的临床效力相关的因素。

Cell and gene therapy products must be tested for sterility, stability, purity and potency. In addition, its important to test clinical cell therapy products for identity, consistency and comparability. Testing cellular and gene therapies is challenging. These therapies are generally collected from a single person so the quantity of material available to test is limited. They are typically transfused immediately or shortly after they are produced so there is a very limited amount of time to complete the assays. Many of these therapies are complex cells that have multiple functions. The cell functions that are critical to the clinical effectiveness of these therapies are often not known. Traditionally, analytic assays such as flow cytometry, ELISA, ELISPOT and cell culture have been used to analyze cellular and gene therapies. While these assays have proven to be very useful, the number and types of factors that can be analyzed with these assays is limited. We have been investigating the use of gene and micro RNA expression assays for the analysis of cellular therapies. These assays can require the use of only small quantities of cells and can be used to assess the expression of the entire transcriptome. We have been testing the ability of global gene and micro RNA expression profiling to determine the utility of these assays for assessing the stability, purity and potency of cellular therapies. We have shown that gene expression profiling can detect changes in stored cells and detect differences between peripheral blood leukocytes (T cells, B cells and monocytes) and hematopoietic stem cells. Gene expression profiling has also been able to detect differences between immature and mature dendritic cells (DCs) and has been useful for comparing mature DCs produced using different combinations of maturation agents. We will soon begin analyzing clinical DC products to compare gene expression with clinical outcome in order to identify biomarkers that might be useful of consistency, stability, identity and potency testing. Bone Marrow Stromal Cells (BMSCs) are being use to modulate the immune system and for a number of regenerative medicine applications including bone repair. BMSCs are a heterogeneous population of cells and a small number of BMSCs are skeletal stem cells which are important for regenerative medicine clinical applications. We have been investing BMSCs to determine markers that identify skeletal stem cells. We are using single cell analysis and next generation DNA sequencing to identify markers for the skeletal stem cells. Chimeric Antigen Receptor (CAR) T cells are being used to treat a number of hematologic malignancies, however, clinical outcomes have varied among recipients of these therapies and some of this variability is likely due to variability, and hence, differences in potency among CAR T cell products. We are using gene expression analysis, mRNA analysis, single cell analysis and next generation sequencing to identify factors associated with the clinical potency of these cells.