Manufacturing of Growth Factors to Enable Cardiopoietic Stem Cell Therapy for Hea

制造生长因子以实现心脏造血干细胞治疗

• 批准号: 8766958
• 负责人: RIDONG CHEN
• 金额: $ 22.41万
• 依托单位: APT THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8766958
• 关键词: Affect; Animal Model; Animals; BMP4; Bioreactors; Blood; Bone Marrow; Cardiac; Cell Culture Techniques; Cell Line; Cell Therapy; Cell physiology; Cells; Clinical; Clinical Trials; Complex; Development; Disease; Endotoxins; Ensure; Equipment; Escherichia coli; Exhibits; Future; Growth; Growth Factor; Guidelines; Health Expenditures; Heart failure; Human; Human Development; Laboratories; Left Ventricular Function; Mesenchymal; Mesenchymal Stem Cells; Methods; Modification; Myocardial; Natural regeneration; Patients; Peptide Signal Sequences; Persons; Phase; Phase III Clinical Trials; Price; Process; Production; Proteins; Reagent; Recombinants; Recruitment Activity; Research; Risk; Secure; Serum; Sialic Acids; Solubility; Stem cells; Suspension substance; Suspensions; System; Technology; Testing; Therapeutic; Transforming Growth Factors; Work; Xeno; activin A; bone morphogenetic protein 4; cost; cost effective; cytokine; functional outcomes; human stem cells; immunogenic; improved; in vivo; killings; manufacturing process; meetings; outcome forecast; promoter; public health relevance; research and development; stem cell therapy; success

DESCRIPTION (provided by applicant): Cardiopoietic cell therapy significantly improved left ventricular function, blunted pathological remodeling, and functional outcomes in animal models and Phase I/II clinical trial of heart failure patients. There is ongoing Phase III trial. In this therapeutic approach, bone marrow-derived mesenchymal stem cells are engaged into cardiac stem cells by exposing to a cardiogenic cocktail of growth factors, including activin A, transforming growth factor-b1, and bone morphogenetic protein-4, which is the single largest cost component of clinical manufacturing. These cytokines all belong to the TGF-b superfamily, which are critical for cardiopoietic stem cell expansion and differentiation. However, due to the complex post-proteolytic modifications and poor solubility at neutral pH, yield from all the curren manufacturing process is very low, and as a result bulk volume is uneconomical and not readily available. Currently, commercial products are transiently produced in non-human cell systems (E coli, SF9, CHO, by, for example, R&D Systems and PeproTech) or HEK293 cells (by, for example, HumanZyme and StemR&D) as research-use only reagents. Today, there is no capacity anywhere in the world to produce these essential TGF-b superfamily cytokines at the scale and quality that will meet the requirements for future large-scale clinical ex vivo processing of cardiopoietic and other human stem cell products. Currently, these high-demand cytokines are costly, $6540-$9,800/mg, which severely hinders clinical scale processing of stem cell therapeutics. We have been focused on the development of a cost-effective and scalable expression system to produce authentic, recombinant TGF-b superfamily cytokines from stable HEK293SH cells that current human cell or non-human cell expression systems either cannot produce or cannot economically produce. The proprietary technology includes HEK293SH, a selected high-yield HEK293 cell line adapted to suspension for growth in serum-free and chemically defined medium, an optimized human promoter and signal peptide, optimization of post-translational processing, and proprietary purification and stabilization methods for these poor-solubility proteins. Currently, Activin A, TGFb1 and BMP4 are produced at >10-30 fold higher yield and exhibit greater stability, higher activity, and higher purity than any other products commercially available, resulting in profoundly lower raw material and process labor costs. In this study, we propose to further optimize the production and develop large-scale and cost-effective production and processing capabilities for generating serum-free and xeno-free TGFb superfamily cytokines, which will enable safe, robust, and cost-effective ex vivo processing of cardiopoietic mesenchymal stem cell therapeutics. The targeted price will be reduced from current $6540-$9,800/mg to $100/mg.

描述（由申请人提供）：心脏病细胞疗法可显着改善左心室功能，钝化病理重塑以及动物模型中的功能结果以及心力衰竭患者的I/II期临床试验。正在进行III期试验。在这种治疗方法中，骨髓来源的间充质干细胞通过暴露于生长因子的心源性鸡尾酒中，包括激活素A，转化生长因子-B1和骨形态发生蛋白4，这是临床制造的最大成本成分。这些细胞因子都属于TGF-B超家族，这对于心po型干细胞扩张和分化至关重要。但是，由于蛋白质的复杂修饰和中性pH值下的溶解度差，因此所有curren制造过程的产量非常低，因此大量体积不经济，并且不容易获得。当前，商业产品是在非人类细胞系统（E Coli，Sf9，Cho，由R＆D Systems和Peprotech）或HEK293细胞（例如，例如Humanzyme和STEMR＆D）中瞬时生产的。如今，世界上任何地方都没有能力以规模和质量生产这些必不可少的TGF-B超家族细胞因子，这将满足心脏和其他人类干细胞产品的未来大规模临床临床处理的要求。目前，这些高需求的细胞因子成本高昂，$ 6540- $ 9,800/mg，这严重阻碍了干细胞疗法的临床规模处理。我们一直专注于开发一种具有成本效益且可扩展的表达系统，以产生来自稳定的HEK293SH细胞的真实，重组TGF-B超家族细胞因子，该细胞是当前人类细胞或非人类细胞表达系统无法产生或无法经济上产生的。专有技术包括HEK293SH，这是一种选择的高收益HEK293细胞系，适用于悬浮液，用于无血清和化学定义的培养基的生长，一种优化的人类启动子和信号肽，优化了翻译后加工的优化，以及这些差的纯度固定性蛋白的专有纯化和稳定方法。目前，与市售的任何其他产品相比，激活素A，TGFB1和BMP4的产量高于10-30倍，并且表现出更高的稳定性，更高的活性和更高的纯度，从而产生了较低的原料和过程劳动力成本。在这项研究中，我们建议进一步优化生产，并开发大规模且具有成本效益的生产和加工能力，以生成无血清和无XENO的TGFB TGFB超家族细胞因子，这将使心脏室内充质细胞细胞细胞的安全，稳健且具有成本效益的外部处理。目标价格将从目前的$ 6540- $ 9,800/毫克降低到$ 100/mg。