Engineering novel designer biologics in plant cells for oral treatment of ulcerative colitis

在植物细胞中设计新型生物制剂用于口腔治疗溃疡性结肠炎

基本信息

批准号：
10202300
负责人：
Jianfeng Xu
金额：
$ 39.52万
依托单位：
ARKANSAS STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-17 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10202300
关键词：
Affect Anti-Inflammatory Agents Antibodies Arkansas Attention Autoimmune Behavior Biological Biological Assay Biological Response Modifier Therapy Biotechnology C-terminal Cell Culture Techniques Cell Wall Cell membrane Cell surface Cells Cellulose Charge Chemicals Chronic Clinical Colitis Colon Competence Crohn&apos s disease Digestion Digestive System Disorders Disease Drug Delivery Systems Drug Kinetics Drug Targeting Effectiveness Encapsulated Engineering Enzymes Epithelial Failure GPI Membrane Anchors Gastrointestinal tract structure Glycoproteins Glycosylphosphatidylinositols Goals Hydroxyproline Immune Immunosuppression Industry Inflammation Inflammatory Inflammatory Bowel Diseases Intestinal Diseases Liquid substance Mediating Medical Microfibrils Modification Mus N-terminal Oral Oral Administration Patients Pectins Pharmaceutical Preparations Pharmacologic Substance Pharmacology Pharmacotherapy Plants Polyethylene Glycols Polymers Polysaccharides Post-Translational Protein Processing Process Proteins Psychological reinforcement Public Health Recombinants Research Science Site Sodium Dextran Sulfate Stomach Surface Symptoms System TNF gene Tandem Repeat Sequences Testing Therapeutic Tissues Tobacco Treatment Efficacy Ulcerative Colitis Universities Work adverse drug reaction antitumor effect capsule cost cost effective cytokine design dextran sulfate sodium induced colitis drug efficacy effective therapy glycosylation gut bacteria improved inflammatory marker innovation laboratory experience mouse model novel protein degradation response seryl-proline side effect systemic toxicity therapeutic effectiveness therapeutic protein training opportunity undergraduate student

项目摘要

Project Summary/Abstract Inflammatory bowel disease (IBD), including the two most common subtypes: Crohn’s disease (CD) and ulcerative colitis (UC), represents a group of intestinal disorders that cause prolonged inflammation of the digestive tract. The current therapeutic strategies, including the conventional anti-inflammatory medications and the new biologic drugs targeting the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα), have limited therapeutic efficacy and adverse drug reactions resulting from systemic administration. Colon-targeted oral delivery of anti-TNFα agents is highly desirable for the treatment of IBD, as it improves the drugs’ efficacy while reducing the systemic toxicity. Plant cell culture has emerged as a safe and cost-effective bioproduction platform for therapeutic proteins. A unique feature of the plant cells is that they could serve not only as the “bio-factory,” but also the oral delivery vehicle for recombinant biologics. Recent advances have demonstrated that plant cell walls, made primarily of cellulose microfibrils, can act as an excellent natural capsule for the oral delivery of biologic drugs. This project aims to leverage two unique posttranslational modifications – “glycosyl- phosphatidylinositol (GPI) anchor” and “plant-specific hydroxyproline (Hyp)-O-glycosylation” – to strategically design and engineer novel anti-TNFα biomolecules in plant cells to develop a new class of oral biologic drugs for the treatment of UC. The designer anti-TNFα biomolecules consist of three functional domains: a N-terminal single-chain fragment variable (scFv) of an anti-TNFα antibody, a proprietary Hyp-O-glycosylation module comprised of tandem repeats of the “Ser-Pro” motif, or (SP)n (n= 5 to 30), and a C-terminal GPI anchor. While the GPI anchor will “display” the expressed anti-TNFα biomolecules at the plant cell surface (but still encapsulated within the cell wall) to presumably create a high local concentration of the biologics, the (SP)n glycomodule will stabilize the protein from degradation during both the bioproduction and oral delivery processes. Meanwhile, the negatively charged glycans decorated on the (SP)n glycomodule will target the anti-TNFα biomolecules to the inflamed epithelium where positively charged proteins are always built up. Designer anti- TNFα biomolecules consisting of different sized (SP)n glycomodules will be investigated for their accumulation in tobacco BY-2 cells, biological activity, and stability in a simulated gastric fluid, which will determine an optimal design for the biomolecules. In order to improve the pharmacokinetic behavior of the plant cell produced oral biologic drugs, strategies will be developed to reinforce the plant cell wall matrices by filling in the cell wall pores/channels with polymeric molecules, such as polyethylene glycol (PEG), to increase their capacity for protecting the encapsulated proteins. Finally, the therapeutic effectiveness of the orally administrated designer anti-TNFα biologic (optimal design) in mitigating UC symptoms will be assessed in a dextran sulfate sodium (DSS)-induced colitis mouse model. The immune-modulatory effects of the anti-TNFα biologics will be determined by a histopathological analysis and assay of the inflammatory markers. The proposed research will potentially develop a new platform to produce effective oral biologic drugs for the treatment of UC and other inflammatory diseases in the colon.

项目概要/摘要炎症性肠病 (IBD)，包括两种最常见的亚型：克罗恩病 (CD) 和溃疡性结肠炎 (UC) 代表一组肠道疾病，可导致肠道长期炎症目前的治疗策略包括常规抗炎药物和针对促炎细胞因子肿瘤坏死因子α（TNFα）的新型生物药物，其作用有限结肠靶向口服给药的治疗效果和不良反应。抗 TNFα 药物的递送对于 IBD 的治疗是非常理想的，因为它可以提高药物的疗效，同时降低系统毒性。植物细胞培养已成为一种安全且具有成本效益的生物生产平台。植物细胞的一个独特特征是它们不仅可以充当“生物工厂”，植物细胞也是重组生物制剂的口服递送载体。主要由纤维素微纤维制成的壁可以作为口服递送的极好的天然胶囊该项目旨在利用两种独特的翻译后修饰——“糖基- 磷脂酰肌醇（GPI）锚”和“植物特异性羟脯氨酸（Hyp）-O-糖基化”——战略性地在植物细胞中设计和改造新型抗 TNFα 生物分子，以开发新型口服生物药物用于治疗 UC 的设计抗 TNFα 生物分子由三个功能域组成：N 端抗 TNFα 抗体的单链片段可变区 (scFv)，一种专有的 Hyp-O-糖基化模块由“Ser-Pro”基序或 (SP)n（n= 5 至 30）的串联重复序列和 C 端 GPI 锚点组成。 GPI 锚点将在植物细胞表面“展示”表达的抗 TNFα 生物分子（但仍然封装在细胞壁内），从本质上创造生物制剂的高局部浓度，（SP）n 糖模块将稳定蛋白质在生物生产和口服递送过程中的降解。同时，(SP)n 糖模块上装饰的带负电荷的聚糖将靶向抗 TNFα 发炎的上皮细胞中总是会积聚带正电的蛋白质的生物分子。将研究由不同大小的 (SP)n 糖模块组成的 TNFα 生物分子的积累情况烟草 BY-2 细胞中的生物活性和模拟胃液中的稳定性，这将确定最佳生物分子的设计，以改善植物细胞产生的口服药代动力学行为。生物药物，将制定策略通过填充细胞壁来强化植物细胞壁基质具有聚合物分子（例如聚乙二醇（PEG））的孔/通道，以增加其容量最后，口服给药设计者的治疗效果。将在葡聚糖硫酸钠中评估抗 TNFα 生物制剂（最佳设计）缓解 UC 症状的效果 (DSS) 诱导的结肠炎小鼠模型，抗 TNFα 生物制剂的免疫调节作用将是。该研究将通过组织病理学分析和炎症标记物测定来确定。有可能开发一个新平台来生产有效的口服生物药物，用于治疗 UC 和其他疾病结肠炎症性疾病。