Bera T K, Viner J, Brinkmann E, Pastan I
Laboratory of Molecular Biology, National Cancer Institute, NIH, Bethesda, Maryland 20892-4255, USA.
Cancer Res. 1999 Aug 15;59(16):4018-22.
We have generated a stable bivalent Fv molecule [(dsFv)2] of the anti-erbB2 monoclonal antibody e23 in which the V(H) and V(L) domains of the Fv are linked to each other by a disulfide bond and the two Fvs are connected by a 15-amino acid linker (T. K. Bera et al., J. Mol. Biol., 281: 475-483, 1998). The e23 (dsFv)2 molecule is linked to a truncated form of Pseudomonas exotoxin (PE38) to generate a bivalent disulfide-stabilized immunotoxin e23 (dsFv)2-PE38. Compared to the monovalent immunotoxin, the (dsFv)2 immunotoxin showed greatly increased cytotoxicity to four cancer cell lines expressing low levels of erbB2 but not to four other cell lines with high erbB2 expression. e23 (dsFv)2-PE38 was administered i.v. to mice, and its half-life was determined. The t(1/2)alpha and t(1/2)beta were 20 and 325 min, respectively, whereas the corresponding values for the monovalent dsFv immunotoxin were shorter, 6 and 52 min. The antitumor activities of the monovalent and bivalent immunotoxin were compared using mice bearing A431 tumors. Despite the fact that e23 (dsFv)2-PE38 was 13-fold more active than e23 dsFv-PE38 on A431 cells in cell culture, its antitumor activity in mice was <2-fold that of the monovalent immunotoxin. These data show that a large increase in avidity does not always lead to an increase in cytotoxic activity. Furthermore, in one of the cases in which cytotoxic activity in vitro was greatly enhanced, there was only a small increase in antitumor activity.
我们构建了抗erbB2单克隆抗体e23的稳定二价Fv分子[（dsFv）2]，其中Fv的V（H）和V（L）结构域通过二硫键相互连接，两个Fv通过一个15个氨基酸的接头相连（T.K.贝拉等人，《分子生物学杂志》，281:475 - 483，1998）。e23（dsFv）2分子与截短形式的铜绿假单胞菌外毒素（PE38）相连，以产生二价二硫键稳定的免疫毒素e23（dsFv）2 - PE38。与单价免疫毒素相比，（dsFv）2免疫毒素对四种低水平表达erbB2的癌细胞系的细胞毒性大大增加，但对其他四种高表达erbB2的细胞系则没有。将e23（dsFv）2 - PE38静脉注射给小鼠，并测定其半衰期。α半衰期（t1/2α）和β半衰期（t1/2β）分别为20分钟和325分钟，而单价dsFv免疫毒素的相应值较短，分别为6分钟和52分钟。使用荷A431肿瘤的小鼠比较单价和二价免疫毒素的抗肿瘤活性。尽管在细胞培养中e23（dsFv）2 - PE38对A431细胞的活性比e23 dsFv - PE38高13倍，但其在小鼠中的抗肿瘤活性却不到单价免疫毒素的2倍。这些数据表明，亲和力的大幅增加并不总是导致细胞毒性活性的增加。此外，在体外细胞毒性活性大大增强的一个案例中，抗肿瘤活性仅略有增加。
