Immunobiology Division, Center of Molecular Immunology, P.O. Box 16040, Havana, 11600, Cuba.
Glycoconj J. 2013 Oct;30(7):687-99. doi: 10.1007/s10719-013-9473-y. Epub 2013 Apr 2.
Humans, in contrast to other mammals, do not synthesize N-glycolyl-neuraminic acid (Neu5Gc) due to a deletion in the gene (cmah) encoding the enzyme responsible for this conversion, the cytidine monophospho-N-acetyl-neuraminic acid hydroxylase (CMP-Neu5Ac hydroxylase). The detection of considerable amounts of Neu5Gc-sialoconjugates, in particular gangliosides, in human malignancies makes these antigens attractive targets for immunotherapy, in particular with monoclonal antibodies (mAbs). We have previously described a GM3(Neu5Gc) ganglioside-specific mAb, named 14F7, with the ability to kill tumor cells in a complement-independent manner. Silencing the cmah gene in GM3(Neu5Gc)-expressing L1210 mouse lymphocytic leukemia B cells caused the abrogation of this cytotoxic effect. We now show that cmah-silenced L1210 cells (cmah-kd) express a high level of GM3(Neu5Ac) and have an impaired ability for anchorage-independent cell growth and tumor development in vivo. No evidences of increased immunogenicity of the cmah-kd cell line were found. These results provide new evidences on the role of GM3(Neu5Gc), or Neu5Gc-sialoconjugates in general, in tumor biology. As an important tool in this study, we used the humanized version (here referred to as 7C1 mAb) of a recently described, rationally-designed mutant of 14F7 mAb that is able to bind to both GM3(Neu5Gc) and GM3(Neu5Ac). In contrast to its parental antibody, the humanized 14F7 (14F7hT) mAb, 7C1 mAb was able to kill not only GM3(Neu5Gc)-expressing L1210 wild type cells, but also GM3(Neu5Ac)-expressing cmah-kd cells, which endorses this antibody as a potential agent for cancer immunotherapy.
与其他哺乳动物不同，人类由于编码负责这种转化的酶（胞苷单磷酸-N-乙酰神经氨酸羟化酶）的基因（cmah）缺失，不能合成 N-羟乙酰神经氨酸（Neu5Gc）。在人类恶性肿瘤中检测到相当数量的 Neu5Gc-唾液酸缀合物，特别是神经节苷脂，使这些抗原成为免疫治疗的有吸引力的靶标，特别是使用单克隆抗体（mAb）。我们之前描述了一种 GM3（Neu5Gc）神经节苷脂特异性 mAb，命名为 14F7，具有以补体非依赖方式杀死肿瘤细胞的能力。在表达 GM3（Neu5Gc）的 L1210 小鼠淋巴细胞白血病 B 细胞中沉默 cmah 基因导致这种细胞毒性作用被阻断。我们现在表明，沉默 cmah 的 L1210 细胞（cmah-kd）表达高水平的 GM3（Neu5Ac），并且在体内锚定独立细胞生长和肿瘤发展的能力受损。未发现 cmah-kd 细胞系免疫原性增加的证据。这些结果为 GM3（Neu5Gc）或 Neu5Gc-唾液酸缀合物在肿瘤生物学中的作用提供了新的证据。作为这项研究中的重要工具，我们使用了最近描述的、经过合理设计的 14F7 mAb 突变体的人源化版本（在此称为 7C1 mAb），该突变体能够结合 GM3（Neu5Gc）和 GM3（Neu5Ac）。与亲本抗体相反，人源化 14F7（14F7hT）mAb，7C1 mAb 不仅能够杀死表达 GM3（Neu5Gc）的 L1210 野生型细胞，还能够杀死表达 GM3（Neu5Ac）的 cmah-kd 细胞，这使该抗体成为癌症免疫治疗的潜在药物。
