Jiang Jing, Liu Xiaobin, Deng Leixiu, Zhang Peipei, Wang Guangjun, Wang Shifu, Liu Honghao, Su Yunpeng
Department of Pharmacology, Binzhou Medical University, Shandong Province, Yantai 256603, China.
Shandong Simcere-Medgenn Biopharmaceutical Co., Ltd., Shandong Province, Yantai 264006, China.
Eur J Pharmacol. 2014 Oct 5;740:722-32. doi: 10.1016/j.ejphar.2014.06.002. Epub 2014 Jun 11.
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) exhibits potent antitumor activity in a wide range of cancers without deleterious side effects on normal tissues. Several TRAIL derivatives have been developed to improve its pharmacokinetics and therapeutic effects through strategies such as adding a leucine zipper to increase the circulation half-life. To obtain clinical grade LZ-TRAIL for phase I clinical trial, a single batch of 30 L bioreactor culture was performed using the Escherichia coli BL21 (DE3) strain expressing the recombinant LZ-TRAIL. A robust LZ-TRAIL production fermentation process was developed, which could be scaled up from 5L to 50 L, and had a titer of approximately 1.4 g/l. A four-step purification strategy was carried out to obtain a final product with over 95% purity and 45% yield. The final material was filter sterilized, aseptically vialed, and stored at 4°C, and comprehensively characterized using multiple assays (vialed product was sterile, purity was 95%, aggregates were <5%, potency revealed IC50 of 9 nM on MDA-MB-231 cells, and the endotoxin level was <0.25 U/mg). The purity, composition, and functional activities of the molecule were confirmed. in vivo investigations indicated that LZ-TRAIL has better antitumor potency in three Xenograft tumor models compared to TRAIL (95-281). LZ-TRAIL also showed improved pharmacokinetic and safety profiles in cynomolgus monkeys without abnormalities associated with drug exposure. In conclusion, the scalable synthesis of LZ-TRAIL is useful for production of phase I clinical trial material. These preclinical investigations warrant further clinical development of this product for cancer therapy.
肿瘤坏死因子相关凋亡诱导配体（TRAIL）在多种癌症中展现出强大的抗肿瘤活性，且对正常组织无有害副作用。已研发出多种TRAIL衍生物，通过诸如添加亮氨酸拉链以延长循环半衰期等策略来改善其药代动力学和治疗效果。为获得用于I期临床试验的临床级LZ - TRAIL，使用表达重组LZ - TRAIL的大肠杆菌BL21（DE3）菌株进行了一批30 L生物反应器培养。开发了一种稳健的LZ - TRAIL生产发酵工艺，该工艺可从5 L扩大至50 L，滴度约为1.4 g/l。实施了四步纯化策略，以获得纯度超过95%、产率为45%的最终产品。最终材料经过滤除菌、无菌分装，并储存在4°C，使用多种检测方法进行了全面表征（分装产品无菌，纯度为95%，聚集体<5%，对MDA - MB - 231细胞的效力显示IC50为9 nM，内毒素水平<0.25 U/mg）。确认了该分子的纯度、组成和功能活性。体内研究表明，与TRAIL相比，LZ - TRAIL在三种异种移植肿瘤模型中具有更好的抗肿瘤效力（95 - 281）。LZ - TRAIL在食蟹猴中还显示出改善的药代动力学和安全性特征，且无与药物暴露相关的异常情况。总之，LZ - TRAIL的可扩展合成对于生产I期临床试验材料很有用。这些临床前研究为该产品用于癌症治疗的进一步临床开发提供了依据。
