Kurtzberg Joanne, Buntz Susan, Gentry Tracy, Noeldner Pamela, Ozamiz April, Rusche Benjamin, Storms Robert W, Wollish Amy, Wenger David A, Balber Andrew E
Robertson Clinical and Translational Cell Therapy Program, Duke Translational Medicine Institute, Duke University Medical Center, Durham, North Carolina, USA.
Lysosomal Diseases Testing Laboratory, Thomas Jefferson University, Philadelphia, Pennsylvania, USA.
Cytotherapy. 2015 Jun;17(6):803-815. doi: 10.1016/j.jcyt.2015.02.006. Epub 2015 Mar 12.
Cord blood (CB) transplantation slows neurodegeneration during certain inherited metabolic diseases. However, the number of donor cells in the brain of patients does not appear to be sufficient to provide benefit until several months after transplant. We developed the cell product DUOC-01 to provide therapeutic effects in the early post-transplant period.
DUOC-01 cultures initiated from banked CB units were characterized by use of time-lapse photomicroscopy during the 21-day manufacturing process. Antigen expression was measured by means of flow cytometry and immunocytochemistry; transcripts for cytokines and enzymes by quantitative real-time polymerase chain reaction; activities of lysosomal enzymes by direct biochemical analysis; alloreactivity of DUOC-01 and of peripheral blood (PB) mononuclear cells (MNC) to DUOC-01 by mixed lymphocyte culture methods; and cytokine secretion by Bioplex assays.
DUOC-01 cultures contained highly active, attached, motile, slowly proliferating cells that expressed common (cluster of differentiation [CD]11b, CD14 and Iba1), M1 type (CD16, inducible nitric oxide synthase), and M2-type (CD163, CD206) macrophage or microglia markers. Activities of 11 disease-relevant lysosomal enzymes in DUOC-01 products were similar to those of normal PB cells. All DUOC-01 products secreted interleukin (IL)-6 and IL-10. Accumulation of transforming growth factor-β, IL-1β, interferon-γ and TNF-α in supernatants was variable. IL-12, IL-2, IL-4, IL-5 and IL-13 were not detected at significant concentrations. Galactocerebrosidase, transforming growth factor-β and IL-10 transcripts were specifically enriched in DUOC-01 relative to CB cells. PB MNCs proliferated and released cytokines in response to DUOC-01. DUOC-01 did not proliferate in response to mismatched MNC.
DUOC-01 has potential as an adjunctive cell therapy to myeloablative CB transplant for treatment of inherited metabolic diseases.
脐血（CB）移植可减缓某些遗传性代谢疾病患者的神经退行性变。然而，在移植后数月内，患者大脑中的供体细胞数量似乎不足以产生疗效。我们研发了细胞产品DUOC-01，以期在移植后早期发挥治疗作用。
在21天的生产过程中，利用延时显微镜对源自库存CB单位的DUOC-01培养物进行特性分析。通过流式细胞术和免疫细胞化学检测抗原表达；采用定量实时聚合酶链反应检测细胞因子和酶的转录本；通过直接生化分析检测溶酶体酶的活性；采用混合淋巴细胞培养方法检测DUOC-01及外周血（PB）单个核细胞（MNC）对DUOC-01的同种异体反应性；通过Bioplex检测法检测细胞因子分泌情况。
DUOC-01培养物包含高活性、贴壁、可移动、增殖缓慢的细胞，这些细胞表达常见的（分化簇[CD]11b、CD14和Iba1）、M1型（CD16、诱导型一氧化氮合酶）和M2型（CD163、CD206）巨噬细胞或小胶质细胞标志物。DUOC-01产品中11种与疾病相关的溶酶体酶的活性与正常PB细胞相似。所有DUOC-01产品均分泌白细胞介素（IL）-6和IL-10。上清液中转化生长因子-β、IL-1β、干扰素-γ和肿瘤坏死因子-α的积累情况各不相同。未检测到显著浓度的IL-12、IL-2、IL-4、IL-5和IL-13。相对于CB细胞，半乳糖脑苷脂酶、转化生长因子-β和IL-10转录本在DUOC-01中特异性富集。PB MNC对DUOC-01有增殖反应并释放细胞因子。DUOC-01对不匹配的MNC无增殖反应。
DUOC-01有潜力作为清髓性CB移植的辅助细胞疗法用于治疗遗传性代谢疾病。
