Filipovich A H, Vallera D, McGlave P, Polich D, Gajl-Peczalska K, Haake R, Lasky L, Blazar B, Ramsay N K, Kersey J
University of Minnesota, Minneapolis 55455.
Transplantation. 1990 Sep;50(3):410-5. doi: 10.1097/00007890-199009000-00011.
Twenty-nine patients with advanced leukemias (median age 34 years) received histocompatible sibling marrow that had been depleted of T cells by ex vivo incubation with anti-CD5 monoclonal antibody-ricin immunotoxin (T101-R) for the purpose of graft-versus-host disease prophylaxis. Donor cell engraftment was documented in 28/29 patients by DNA restriction fragment length polymorphisms. In this pilot study the dose of T101-R incubated with donor marrow was increased in a stepwise manner from 300 ng (10 patients) to 600 ng (5 patients) to 1000 ng immunotoxin (IT)/10(7) bone marrow mononuclear cells (14 patients) in an attempt to achieve more effective GvHD prophylaxis. A statistically significant reduction in acute GvHD was achieved for patients receiving marrow pretreated with 1000 ng of immunotoxin (34%) compared to recipients of BM treated with 300 ng immunotoxin (100%, P = 0.0004). T-depleted marrow samples were evaluated for residual T cell activity using several in vitro assays including proliferation to the purified mitogen PHA (HA-17) and in mixed lymphocyte culture (MLC), T cell cytotoxicity, a limiting dilution assay for detecting precursors of proliferating T cells (LDApPTL), and phenotypic analysis of viable T cells expanded in 16-day culture with interleukin 2. The extent of T cell depletion determined by LDA assay varied widely at each immunotoxin concentration used. Thus, there was no correlation between the dose of T cells infused and subsequent GvHD. Phenotyping of lymphocytes recovered from immunotoxin-treated marrow demonstrated that residual T cells were CD5 negative in all cases tested. The only in vitro parameter that predicted subsequent acute or chronic GvHD was the demonstration of viable CD5 negative lymphocytes with T cell phenotype (CD2, CD3, and/or CD7 positive) after 16-day culture with IL-2 of the T-depleted bone marrow. We observed that such CD5 negative cells expressing other T cell markers have cytotoxic function and speculate that these cells may be capable of mediating GvHD in allogeneic transplantation.
29例晚期白血病患者（中位年龄34岁）接受了通过与抗CD5单克隆抗体-蓖麻毒素免疫毒素（T101-R）体外孵育以清除T细胞的组织相容性同胞骨髓，目的是预防移植物抗宿主病。通过DNA限制性片段长度多态性在28/29例患者中记录到供体细胞植入。在这项初步研究中，与供体骨髓孵育的T101-R剂量从300 ng（10例患者）逐步增加到600 ng（5例患者）再到1000 ng免疫毒素（IT）/10⁷骨髓单个核细胞（14例患者），试图实现更有效的移植物抗宿主病预防。接受用1000 ng免疫毒素预处理骨髓的患者急性移植物抗宿主病有统计学意义的降低（34%），而接受用300 ng免疫毒素处理骨髓的患者为100%（P = 0.0004）。使用几种体外试验评估T细胞清除的骨髓样本的残余T细胞活性，包括对纯化的丝裂原PHA（HA-17）的增殖反应和混合淋巴细胞培养（MLC）、T细胞细胞毒性、检测增殖T细胞前体的有限稀释分析（LDApPTL）以及在含白细胞介素2的16天培养中对存活T细胞的表型分析。在每个使用的免疫毒素浓度下，通过LDA分析确定的T细胞清除程度差异很大。因此，输注的T细胞剂量与随后的移植物抗宿主病之间没有相关性。对从免疫毒素处理的骨髓中回收的淋巴细胞进行表型分析表明，在所有测试病例中残余T细胞均为CD5阴性。唯一能预测随后急性或慢性移植物抗宿主病的体外参数是在T细胞清除的骨髓用IL-2培养16天后显示具有T细胞表型（CD2、CD3和/或CD7阳性）的存活CD5阴性淋巴细胞。我们观察到这种表达其他T细胞标志物的CD5阴性细胞具有细胞毒性功能，并推测这些细胞可能能够在同种异体移植中介导移植物抗宿主病。
