Munshi N C, Govindarajan R, Drake R, Ding L M, Iyer R, Saylors R, Kornbluth J, Marcus S, Chiang Y, Ennist D, Kwak L, Reynolds C, Tricot G, Barlogie B
University of Arkansas for Medical Sciences, Little Rock 72205, USA.
Blood. 1997 Feb 15;89(4):1334-40.
A graft-versus-leukemia (GVL) effect has been considered a major factor responsible for cures in patients with hematologic malignancies undergoing allogeneic bone marrow transplantation; however, associated graft-versus-host disease (GVHD) results in significant morbidity and mortality. T-cell depletion reduces the incidence and severity of GVHD but eliminates, at least partially, the GVL effect. Reinfusion of donor T lymphocytes at relapse posttransplantation can induce a potent antitumor response, but GVHD still occurs in the majority of patients. Prior transduction of T lymphocytes with the suicide gene, the viral thymidine kinase (TK), permits specific cell kill on administration of ganciclovir (GCV). Therefore, infusion of TK-transduced T lymphocytes may induce GVL effect and allow for their subsequent selective elimination in case GVHD develops. To evaluate the efficacy and feasibility of this promising approach, anti-CD3-stimulated primary human lymphocytes cultured in interleukin-2 were TK-transduced by a retroviral vector carrying both TK and neomycin-resistance genes. After selection in G418, more than 90% of the cells contained the TK gene as shown by a semiquantitative polymerase chain reaction. In addition, 1 to 5 days of GCV exposure, at clinically achievable concentrations of 20 to 50 micromol/L, induced > or = 90% killing of G418-selected cells without affecting nontransduced cells. Correlation of the extent of T-cell kill and the proportion of TK-gene-transduced cells is consistent with the absence of a bystander effect. Transduced cells were CD3+ and either CD8+ or CD4+ and retained functional properties of untransduced cells. In vivo administration of GCV prevented tumor development after subcutaneous injection of TK-transduced murine myeloma cells (MOPC-11), whereas such an effect was not observed on injection of untransduced cells into the opposite flank. Our studies provide critical information that (1) adequate numbers of TK-transduced lymphocytes can be selected efficiently with > or = 90% purity, (2) selected cells remain functional, (3) 24 hours of exposure to GCV at clinically achievable concentration effects > or = 90% killing of selected cells, and (4) GCV is effective in vivo in killing TK-transduced cells. Based on these data, a clinical study has been initiated in patients with multiple myeloma with persistent or relapsing disease after T-cell-depleted allogeneic transplants.
移植物抗白血病（GVL）效应被认为是接受异基因骨髓移植的血液系统恶性肿瘤患者得以治愈的主要因素；然而，与之相关的移植物抗宿主病（GVHD）却会导致显著的发病率和死亡率。去除T细胞可降低GVHD的发生率和严重程度，但至少会部分消除GVL效应。在移植后复发时回输供体T淋巴细胞可诱导有效的抗肿瘤反应，但大多数患者仍会发生GVHD。预先用自杀基因——病毒胸苷激酶（TK）转导T淋巴细胞，在给予更昔洛韦（GCV）时可实现特异性细胞杀伤。因此，输注经TK转导的T淋巴细胞可能会诱导GVL效应，并在发生GVHD时允许对其进行后续选择性清除。为评估这种有前景的方法的疗效和可行性，将携带TK和新霉素抗性基因的逆转录病毒载体转导在白细胞介素-2中培养的抗CD3刺激的原代人淋巴细胞。在G418中筛选后，通过半定量聚合酶链反应显示，超过90%的细胞含有TK基因。此外，在临床可达到的20至50微摩尔/升浓度下，暴露于GCV 1至5天可诱导对G418筛选细胞的≥90%杀伤，而不影响未转导的细胞。T细胞杀伤程度与TK基因转导细胞比例的相关性与不存在旁观者效应一致。转导细胞为CD3 +，且为CD8 +或CD4 +，并保留了未转导细胞的功能特性。在皮下注射经TK转导的鼠骨髓瘤细胞（MOPC-11）后，体内给予GCV可预防肿瘤发生，而将未转导的细胞注射到对侧胁腹则未观察到这种效果。我们的研究提供了关键信息，即（1）可以高效选择纯度≥90%的足够数量的经TK转导的淋巴细胞，（2）所选细胞仍保持功能，（3）在临床可达到的浓度下暴露于GCV 24小时可实现对所选细胞≥90%的杀伤，以及（4）GCV在体内对杀伤经TK转导的细胞有效。基于这些数据，已启动一项针对T细胞去除的异基因移植后患有持续性或复发性疾病的多发性骨髓瘤患者的临床研究。
