Preclinical Testing of Interleukin-7 in Primates

背景：大剂量化疗或全身照射后的造血细胞移植(HCT)后感染是常见的。这至少在一定程度上是由于CD4细胞重组缓慢。在成人患者中，T细胞谱系受到限制，表型原始T细胞数多年来一直很低。这主要是由于成年患者再生T细胞的能力有限(从造血干细胞)。假设：白介素7(IL7)可加速T细胞的再生。研究设计与方法：利用黄色荧光蛋白(YFP)基因转导的CD34细胞和绿色荧光蛋白(GFP)基因转导的T细胞，对照射后的狒狒进行自体HCT移植。移植后，IL7治疗的动物与安慰剂治疗的动物相比，将通过1.计数YFP T细胞，2.通过检测多种抗原免疫后各种抗原特异性T细胞来确定功能T细胞的谱系，3.计数表型原始的(如CD45RA高的)CD4T细胞，4.评估胸腺的大小和组织学，包括RAG和TDT免疫染色，胸腺细胞亚群和T细胞受体重排切除环(TREC)水平，以及5.测定外周T细胞中的TREC水平，来评估IL7组和安慰剂组动物的从头T细胞生成。为了排除IL7处理的动物血液中YFP/CD45RA High/TREC T细胞计数增加的预期结果是由于IL7诱导的干细胞来源的T细胞从器官转移到血液的可能性，血液和淋巴组织中都将对YFP、CD45RA High和TREC细胞进行计数。我们还将通过计数GFP T细胞来评估IL7是否刺激灵长类T细胞的外周扩张。此外，我们将确定IL7在灵长类动物中的有效剂量(从头开始刺激再生)以及潜在的毒性，包括骨质疏松症或淋巴增生性疾病。这些结果将用于临床研究的设计。

Background: Infections are frequent after high dose chemotherapy or total body irradiation followed by hematopoietic cell transplantation (HCT). This is due at least in part to the slow reconstitution of CD4 cells. In adult patients, the T cell repertoire is restricted and counts of phenotypically nave T cells are low for years. This is primarily due to the limited ability of adult patients to regenerate T cells de novo (from hematopoietic stem cells). Hypothesis: Interleukin-7 (IL7) hastens the regeneration of T cells de novo. Research design and Methods: Autologous HCT transplantation of irradiated baboons will be performed, using yellow fluorescent protein (YFP) gene-transduced CD34 cells and green fluorescent protein (GFP) gene-transduced T cells. After transplant, de novo T cell generation in IL7-treated versus placebo-treated animals will be evaluated by 1. enumerating YFP+ T cells, 2. determining functional T cell repertoire through the detection of various antigen-specific T cells after immunization with multiple antigens, 3. enumerating phenotypically nave (e.g., CD45RA high) CD4 T cells, 4. assessing the size and histology of the thymus, including RAG and TdT immunostains, thymocyte subsets and T cell receptor rearrangement excision circle (TREC) levels in thymocytes, and 5. determining TREC levels in peripheral T cells. To rule out the possibility that the expected result of increased YFP+/CD45RA high/TREC+ T cell counts in the blood of the IL7-treated animals is due to IL7-induced shift of stem cell-derived T cells from organs to the blood, YFP+, CD45RA high and TREC+ cells will be enumerated in both the blood and lymphatic tissues. We will also evaluate whether IL7 stimulates peripheral expansion of primate T cells by enumerating GFP+ T cells. Moreover, we will determine the efficacious (de novo regeneration-stimulating) dose of IL7 in primates and potentially toxicity including osteoporosis or a lymphoproliferative disorder. These results will be used for the design of clinical studies.