Site Selective Antibody-Oligonucleotide Conjugation via Microbial Transglutaminase
Site Selective Antibody-Oligonucleotide Conjugation via Microbial Transglutaminase
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DOI：
10.3390/molecules24183287
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发表时间：
2019-09-01
期刊：
影响因子：
4.6
通讯作者：
Dowdy, Steven F.
中科院分区：
文献类型：
--
作者：
Huggins, Ian J.;Medina, Carlos A.;Dowdy, Steven F.
核酸疗法（NATs），包括sirna和反义寡核苷酸（ASOs），具有巨大的潜力来治疗不可药物的基因组。将sirna和ASOs靶向特定的细胞类型已经显著提高了疗效并降低了毒性。事实上，tris-GalNAc与sirna和ASOs的结合已显示出针对肝细胞驱动的疾病的临床疗效。然而，由于一些原因，用寡核苷酸疗法靶向非肝脏疾病仍然存在问题，包括靶向特定细胞类型和内体逃逸。靶向sirna和ASOs的单克隆抗体（mAb）具有将这些药物输送到各种特定细胞和组织类型的潜力。然而，大多数偶联策略依赖于赖氨酸或半胱氨酸残基的随机化学偶联，导致偶联物的异质性和药物：抗体比率（DAR）的分布。为了生产每个单抗具有两个sirna的均匀的DAR-2偶联物，我们开发了一种新的两步偶联程序，包括微生物转谷氨酰胺酶（mtga）用叠氮化物功能化的连接肽标记抗体c端，该连接肽随后可以通过叠氮化物-炔环加成偶联到含二苄基环辛基（DBCO）的寡核苷酸。使用这种策略产生的抗体- sirna（和ASO）偶联物（arc）是可溶性的，化学定义的靶向寡核苷酸疗法，具有极大增加靶向细胞类型数量的潜力。
Nucleic Acid Therapeutics (NATs), including siRNAs and AntiSense Oligonucleotides (ASOs), have great potential to drug the undruggable genome. Targeting siRNAs and ASOs to specific cell types of interest has driven dramatic improvement in efficacy and reduction in toxicity. Indeed, conjugation of tris-GalNAc to siRNAs and ASOs has shown clinical efficacy in targeting diseases driven by liver hepatocytes. However, targeting non-hepatic diseases with oligonucleotide therapeutics has remained problematic for several reasons, including targeting specific cell types and endosomal escape. Monoclonal antibody (mAb) targeting of siRNAs and ASOs has the potential to deliver these drugs to a variety of specific cell and tissue types. However, most conjugation strategies rely on random chemical conjugation through lysine or cysteine residues resulting in conjugate heterogeneity and a distribution of Drug:Antibody Ratios (DAR). To produce homogeneous DAR-2 conjugates with two siRNAs per mAb, we developed a novel two-step conjugation procedure involving microbial transglutaminase (MTGase) tagging of the antibody C-terminus with an azide-functionalized linker peptide that can be subsequently conjugated to dibenzylcyclooctyne (DBCO) bearing oligonucleotides through azide-alkyne cycloaddition. Antibody-siRNA (and ASO) conjugates (ARCs) produced using this strategy are soluble, chemically defined targeted oligonucleotide therapeutics that have the potential to greatly increase the number of targetable cell types.