Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, Buffalo, New York, 14214, USA.
AAPS J. 2022 Mar 25;24(3):47. doi: 10.1208/s12248-022-00698-x.
This work describes use of anti-carcinoembryonic antigen antibodies (10H6, T84.66) for targeted delivery of an endosomal escape peptide (H6CM18) and gelonin, a type I ribosome inactivating protein. The viability of colorectal cancer cells (LS174T, LoVo) was assessed following treatment with gelonin or gelonin immunotoxins, with or without co-treatment with T84.66-H6CM18. Fluorescent microscopy was used to visualize the escape of immunoconjugates from endosomes of treated cells, and efficacy and toxicity were assessed in vivo in xenograft tumor-bearing mice following single- and multiple-dose regimens. Application of 25 pM T84.66-H6CM18 combined with T84.66-gelonin increased gelonin potency by ~ 1,000-fold and by ~ 6,000-fold in LS174T and LoVo cells. Intravenous 10H6-gelonin at 1.0 mg/kg was well tolerated by LS174T tumor-bearing mice, while 10 and 25 mg/kg doses led to signs of toxicity. Single-dose administration of PBS, gelonin conjugated to T84.66 or 10H6, T84.66-H6CM18, or gelonin immunotoxins co-administered with T84.66-H6CM18 were evaluated. The combinations of T84.66-gelonin + 1.0 mg/kg T84.66-H6CM18 and 10H6-gelonin + 0.1 mg/kg T84.66-H6CM18 led to significant delays in LS174T growth. Use of a multiple-dose regimen allowed further anti-tumor effects, significantly extending median survival time by 33% and by 69%, for mice receiving 1 mg/kg 10H6-gelonin + 0.1 mg/kg T84.66-H6CM18 (p = 0.0072) and 1 mg/kg 10H6-gelonin + 1 mg/kg T84.66-H6CM18 (p = 0.0017). Combined administration of gelonin immunoconjugates with antibody-targeted endosomal escape peptides increased the delivery of gelonin to the cytoplasm of targeted cells, increased gelonin cell killing in vitro by 1,000-6,000 fold, and significantly increased in vivo efficacy.
本研究描述了使用抗癌胚抗原抗体（10H6、T84.66）将内体逃逸肽（H6CM18）和蓖麻毒素导入细胞内。将 LS174T、LoVo 结肠癌细胞与蓖麻毒素或蓖麻毒素免疫毒素进行孵育，检测细胞活力，比较共处理 T84.66-H6CM18 对细胞活力的影响。荧光显微镜观察免疫偶联物从处理细胞的内体逃逸的情况，评估单剂量和多剂量方案下在荷瘤裸鼠体内的疗效和毒性。25 pM T84.66-H6CM18 联合 T84.66-蓖麻毒素可使 LS174T 和 LoVo 细胞中蓖麻毒素的效力增加 1000 倍和 6000 倍。1.0 mg/kg 静脉注射 10H6-蓖麻毒素对 LS174T 荷瘤小鼠的耐受性良好，而 10 和 25 mg/kg 剂量会导致毒性迹象。评估 PBS、T84.66 偶联的蓖麻毒素或 10H6、T84.66-H6CM18 的单剂量给药以及 T84.66-H6CM18 共给药的免疫毒素。T84.66-蓖麻毒素+1.0 mg/kg T84.66-H6CM18 和 10H6-蓖麻毒素+0.1 mg/kg T84.66-H6CM18 的联合用药显著延缓了 LS174T 的生长。多剂量方案的使用可进一步增强抗肿瘤作用，使接受 1.0 mg/kg 10H6-蓖麻毒素+0.1 mg/kg T84.66-H6CM18（p=0.0072）和 1.0 mg/kg 10H6-蓖麻毒素+1.0 mg/kg T84.66-H6CM18（p=0.0017）的小鼠的中位生存时间分别显著延长 33%和 69%。
