A bispecific recombinant immunotoxin, DT2219, targeting human CD19 and CD22 receptors in a mouse xenograft model of B-cell leukemia/lymphoma
A bispecific recombinant immunotoxin, DT2219, targeting human CD19 and CD22 receptors in a mouse xenograft model of B-cell leukemia/lymphoma
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DOI：
10.1158/1078-0432.ccr-04-2290
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发表时间：
2005-05-15
影响因子：
11.5
通讯作者：
Chen, H
中科院分区：
文献类型：
--
作者：
Vallera, DA;Todhunter, DA;Chen, H
一种新型双特异性单链融合蛋白 DT2219，由白喉毒素 (DT390) 的催化和易位结构域与两个识别 CD19 和 CD22 的重复 sFv 亚基融合组成，并在大肠杆菌中表达。通过掺入一段人肌肉醛缩酶并使用基于 N-月桂酰肌氨酸钠去污剂的重折叠程序，解决了重折叠步骤中的产量、纯度和聚集问题。通过将抗 CD19 和抗 CD22 组合在同一单链分子上解决了功效降低的问题。 DT2219 比单独使用抗 CD19 和抗 CD22 sFv 制成的单体或二价免疫毒素具有更高的抗癌活性，并且与抗 CD19 和抗 CD22 亲本单克隆抗体相比，它与患者白血病细胞和 CD19(+)CD22(+) Daudi 或 Raji 细胞的结合水平更高。所得的DT2219经过突变以增强其亲合力，在体外对Daudi细胞具有细胞毒性（IC50 = 0.3 nmol/L）。在体内，DT2219在侧腹肿瘤治疗模型中有效，在该模型中显着抑制肿瘤生长（P < 0.05），在全身模型中显着延长已建立Daudi的严重联合免疫缺陷小鼠的生存期（P < 0.008）与对照相比。 DT2219在识别B细胞恶性肿瘤方面具有更广泛的反应性，具有更强的杀伤力，并且比使用个体免疫毒素需要更少的毒素，这值得作为治疗B白血病/淋巴瘤的新药进行进一步研究。
A novel bispecific single-chain fusion protein, DT2219, was assembled consisting of the catalytic and translocation domains of diphtheria toxin (DT390) fused to two repeating sFv subunits recognizing CD19 and CD22 and expressed in Escherichia coli. Problems with yield, purity, and aggregation in the refolding step were solved by incorporating a segment of human muscle aldolase and by using a sodium N-lauroyl-sarcosine detergent-based refolding procedure. Problems with reduced efficacy were addressed by combining the anti-CD19 and anti-CD22 on the same singlechain molecule. DT2219 had greater anticancer activity than monomeric or bivalent immunotoxins made with anti-CD19 and anti-CD22 sFv alone and it showed a higher level of binding to patient leukemia cells and to CD19(+)CD22(+) Daudi or Raji cells than did anti-CD19 and anti-CD22 parental monoclonal antibodies. The resulting DT2219, mutated to enhance its avidity, was cytotoxic to Daudi cells in vitro (IC50 = 0.3 nmol/L), In vivo, DT2219 was effective in a flank tumor therapy model in which it significantly inhibited tumor growth (P < 0.05) and in a systemic model in which it significantly prolonged survival of severe combined immunodeficient mice with established Daudi (P < 0.008) compared with controls. DT2219 has broader reactivity in recognizing B-cell malignancies, has more killing power, and requires less toxin than using individual immunotoxin, which warrants further investigation as a new drug for treating B leukemia/lymphoma.