完全人源化的抗癌胚抗原（CEA）Gold 4表位的IgG和IgM抗体，设计用于结直肠癌的放射免疫治疗（RIT）。

Fully human IgG and IgM antibodies directed against the carcinoembryonic antigen (CEA) Gold 4 epitope and designed for radioimmunotherapy (RIT) of colorectal cancers.

作者信息

Garambois Véronique, Glaussel Fabienne, Foulquier Elodie, Ychou Marc, Pugnière Martine, Luo Robin X, Bezabeh Binyam, Pèlegrin André

机构信息

INSERM, EMI0227 Centre de Recherche en Cancérologie, CRLC Val d'Aurelle-Paul Lamarque, F-34298 Cedex 5 Montpellier, France.

出版信息

BMC Cancer. 2004 Oct 15;4:75. doi: 10.1186/1471-2407-4-75.

DOI:10.1186/1471-2407-4-75

PMID:15488142

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC526287/

Abstract

BACKGROUND

Human monoclonal antibodies (MAbs) are needed for colon cancer radioimmunotherapy (RIT) to allow for repeated injections. Carcinoembryonic antigen (CEA) being the reference antigen for immunotargeting of these tumors, we developed human anti-CEA MAbs.

METHODS

XenoMouse-G2 animals were immunized with CEA. Among all the antibodies produced, two of them, VG-IgG2kappa and VG-IgM, were selected for characterization in vitro in comparison with the human-mouse chimeric anti-CEA MAb X4 using flow cytometry, surface plasmon resonance, and binding to radiolabeled soluble CEA and in vivo in human colon carcinoma LS174T bearing nude mice.

RESULTS

Flow cytometry analysis demonstrated binding of MAbs on CEA-expressing cells without any binding on NCA-expressing human granulocytes. In a competitive binding assay using five reference MAbs, directed against the five Gold CEA epitopes, VG-IgG2kappa and VG-IgM were shown to be directed against the Gold 4 epitope. The affinities of purified VG-IgG2kappa and VG-IgM were determined to be 0.19 +/- 0.06 x 10(8) M(-1) and 1.30 +/- 0.06 x 10(8) M(-1), respectively, as compared with 0.61 +/- 0.05 x 10(8) M(-1) for the reference MAb X4. In a soluble phase assay, the binding capacities of VG-IgG2kappa and VG-IgM to soluble CEA were clearly lower than that of the control chimeric MAb X4. A human MAb concentration of about 10(-7) M was needed to precipitate approximatively 1 ng 125I-rhCEA as compared with 10(-9) M for MAb X4, suggesting a preferential binding of the human MAbs to solid phase CEA. In vivo, 24 h post-injection, 125I-VG-IgG2kappa demonstrated a high tumor uptake (25.4 +/- 7.3%ID/g), close to that of 131I-X4 (21.7 +/- 7.2%ID/g). At 72 h post-injection, 125I-VG-IgG2kappa was still concentrated in the tumor (28.4 +/- 11.0%ID/g) whereas the tumor concentration of 131I-X4 was significantly reduced (12.5 +/- 4.8%ID/g). At no time after injection was there any accumulation of the radiolabeled MAbs in normal tissues. A pertinent analysis of VG-IgM biodistribution was not possible in this mouse model in which IgM displays a very short half-life due to poly-Ig receptor expression in the liver.

CONCLUSION

Our human anti-CEA IgG2kappa is a promising candidate for radioimmunotherapy in intact form, as F(ab')2 fragments, or as a bispecific antibody.

摘要

背景

结肠癌放射免疫疗法（RIT）需要人源单克隆抗体（MAb）以实现重复注射。癌胚抗原（CEA）是这些肿瘤免疫靶向的参考抗原，我们研发了人抗CEA单克隆抗体。

方法

用CEA免疫XenoMouse-G2动物。在产生的所有抗体中，选择其中两种，即VG-IgG2κ和VG-IgM，与鼠-人嵌合抗CEA单克隆抗体X4进行体外特性分析，采用流式细胞术、表面等离子体共振以及与放射性标记的可溶性CEA结合，并在荷人结肠癌LS174T的裸鼠体内进行分析。

结果

流式细胞术分析表明，单克隆抗体可与表达CEA的细胞结合，而不与表达NCA的人粒细胞结合。在使用针对5个Gold CEA表位的5种参考单克隆抗体的竞争性结合试验中，VG-IgG2κ和VG-IgM显示针对Gold 4表位。纯化的VG-IgG2κ和VG-IgM的亲和力分别测定为0.19±0.06×10⁸ M⁻¹和1.30±0.06×10⁸ M⁻¹，而参考单克隆抗体X4为0.61±0.05×10⁸ M⁻¹。在可溶性相试验中，VG-IgG2κ和VG-IgM与可溶性CEA的结合能力明显低于对照嵌合单克隆抗体X4。沉淀约1 ng ¹²⁵I-rhCEA需要约10⁻⁷ M的人单克隆抗体浓度，而单克隆抗体X4为10⁻⁹ M，这表明人单克隆抗体优先与固相CEA结合。在体内，注射后24小时，¹²⁵I-VG-IgG2κ显示出高肿瘤摄取率（25.4±7.3%ID/g），接近¹³¹I-X4（21.7±7.2%ID/g）。注射后72小时，¹²⁵I-VG-IgG2κ仍集中在肿瘤中（28.4±11.0%ID/g），而¹³¹I-X4的肿瘤浓度显著降低（12.5±4.8%ID/g）。注射后任何时候放射性标记的单克隆抗体在正常组织中均无蓄积。由于肝脏中多聚Ig受体的表达，IgM半衰期很短，因此在该小鼠模型中无法对VG-IgM的生物分布进行相关分析。