Institute of Drug Metabolism and Pharmaceutical Analysis and Zhejiang Provincial Key Laboratory of Anti-Cancer Drug Research, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310058, China.
College of Pharmaceutical Sciences, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Eur J Pharm Sci. 2019 Jun 15;134:81-92. doi: 10.1016/j.ejps.2019.04.015. Epub 2019 Apr 13.
Antibody fragments, as the products of engineered antibodies, exhibit great potential for cancer therapy and imaging. Antibody fragment drug conjugates (AFDCs), which conjugate the highly specific, low-immunity and small-sized antibody fragments with cytotoxic payloads, can overcome the limitations of traditional IgG format drugs in cancer therapy. In this study, a commercialized anti-CD20 monoclonal antibody, ofatumumab (OFA), was applied to generate two site-specific monomethyl auristain E (MMAE)-conjugated AFDCs (Fab-vcMMAE, Fab-CH3mut-vcMMAE) by Sortase A mediated transpeptidation. Compared with OFA-vcMMAE, the two AFDCs maintained most of the binding affinity and the ability of internalization. In vitro studies revealed that Fab-vcMMAE and OFA-vcMMAE had almost identical IC values against CD20-positive cell lines, while Fab-CH3-vcMMAE had a lower anti-tumor activity. In vivo studies showed that Fab-vcMMAE had a significantly higher maximum tolerated dose (MTDs), a 30-fold shorter half-life, and slightly lower antitumor activity within the MTDs than OFA-vcMMAE. The distribution study showed that both of the Fab and Fab-CH3mut had higher penetration rates into the tumors than OFA in a xenograft model. Additionally, no obvious difference in tumor drug accumulation was found between the Fab and OFA groups after the penetration process, but the Fab-CH3mut group exhibited less tumor drug accumulation, possibly contributing to the inferior anti-tumor activity of Fab-CH3mut-vcMMAE in vivo. Overall, we preliminarily demonstrated the characteristics of AFDCs by studying OFA-based AFDCs. Our results revealed that Fab is a promising carrier of MMAE to enhance the anti-tumor activity and increase the safety profile compared with OFA.
抗体片段作为工程抗体的产物，在癌症治疗和成像方面具有巨大的潜力。抗体片段药物偶联物（AFDCs）通过连接高度特异性、低免疫原性和小尺寸的抗体片段与细胞毒性有效载荷，可以克服传统 IgG 格式药物在癌症治疗中的局限性。在这项研究中，一种商业化的抗 CD20 单克隆抗体，奥法木单抗（OFA），被用于通过 Sortase A 介导的转肽作用生成两种位点特异性单甲基奥瑞他汀 E（MMAE）偶联的 AFDCs（Fab-vcMMAE、Fab-CH3mut-vcMMAE）。与 OFA-vcMMAE 相比，这两种 AFDCs 保持了大部分的结合亲和力和内化能力。体外研究表明，Fab-vcMMAE 和 OFA-vcMMAE 对 CD20 阳性细胞系的 IC 值几乎相同，而 Fab-CH3-vcMMAE 的抗肿瘤活性较低。体内研究表明，Fab-vcMMAE 的最大耐受剂量（MTD）显著更高，半衰期短 30 倍，在 MTD 范围内的抗肿瘤活性略低。分布研究表明，在异种移植模型中，Fab 和 Fab-CH3mut 进入肿瘤的穿透率均高于 OFA。此外，在穿透过程后，Fab 和 OFA 组之间的肿瘤药物积累没有明显差异，但 Fab-CH3mut 组的肿瘤药物积累较少，这可能是 Fab-CH3mut-vcMMAE 在体内抗肿瘤活性较低的原因。总的来说，我们通过研究基于 OFA 的 AFDCs 初步展示了 AFDCs 的特征。我们的结果表明，Fab 是 MMAE 的一种有前途的载体，与 OFA 相比，它可以增强抗肿瘤活性并增加安全性。
