Helble Michaela, Chu Jacqueline, Flowers Kaitlyn, Trachtman Abigail R, Huynh Alana, Kim Amber, Shupin Nicholas, Hojecki Casey E, Gary Ebony N, Solieva Shahlo, Parzych Elizabeth M, Weiner David B, Kulp Daniel W, Patel Ami
Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA 19104, USA; Department of Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA 19104, USA.
Mol Ther. 2025 Jan 8;33(1):152-167. doi: 10.1016/j.ymthe.2024.11.030. Epub 2024 Nov 19.
Monoclonal antibodies are an important class of biologics with over 160 Food and Drug Administration/European Union-approved drugs. A significant bottleneck to global accessibility of recombinant monoclonal antibodies stems from complexities related to their production, storage, and distribution. Recently, gene-encoded approaches such as mRNA, DNA, or viral delivery have gained popularity, but ensuring biologically relevant levels of antibody expression in the host remains a critical issue. Using a synthetic DNA platform, we investigated the role of antibody structure and sequence toward in vivo expression. SARS-CoV-2 antibody 2196 was recently engineered as a DNA-encoded monoclonal antibody (DMAb-2196). Utilizing an immunoglobulin heavy and light chain "chain-swap" methodology, we interrogated features of DMAb-2196 that can modulate in vivo expression through rational design and structural modeling. Comparing these results to natural variation of antibody sequences resulted in development of an antibody frequency score that aids in the prediction of expression-improving mutations by leveraging antibody repertoire datasets. We demonstrate that a single amino acid mutation identified through this score increases in vivo expression up to 2-fold and that combinations of mutations can also enhance expression. This analysis has led to a generalized pipeline that can unlock the potential for in vivo delivery of therapeutic antibodies across many indications.
单克隆抗体是一类重要的生物制品，有160多种已获美国食品药品监督管理局/欧盟批准的药物。重组单克隆抗体全球可及性的一个重大瓶颈源于其生产、储存和分发方面的复杂性。最近，诸如信使核糖核酸、脱氧核糖核酸或病毒递送等基因编码方法受到欢迎，但确保宿主中抗体表达达到生物学相关水平仍然是一个关键问题。我们使用一个合成脱氧核糖核酸平台，研究了抗体结构和序列对体内表达的作用。严重急性呼吸综合征冠状病毒2抗体2196最近被设计为一种脱氧核糖核酸编码的单克隆抗体（DMAb - 2196）。利用免疫球蛋白重链和轻链“链交换”方法，我们通过合理设计和结构建模探究了DMAb - 2196中可调节体内表达的特征。将这些结果与抗体序列的自然变异进行比较，得出了一个抗体频率评分，该评分通过利用抗体库数据集有助于预测改善表达的突变。我们证明，通过该评分确定的单个氨基酸突变可使体内表达提高至2倍，并且突变组合也可增强表达。这一分析形成了一个通用流程，可释放治疗性抗体在多种适应症中体内递送的潜力。
