Department of Basic Biomedical Sciences, University of South Dakota Sanford School of Medicine, Vermilion, SD, USA.
Sioux Falls VA Health Care System, Sioux Falls, SD, USA.
Drug Des Devel Ther. 2020 Dec 4;14:5393-5403. doi: 10.2147/DDDT.S274308. eCollection 2020.
Carbamoylated erythropoietin (CEPO) is a chemically engineered, nonhematopoietic derivative of erythropoietin (EPO) that retains its antidepressant and pro-cognitive effects, which are attributed to the increased expression of neurotrophic factors like brain derived neurotrophic factor (BDNF), in the central nervous system. However, the chemical modification process which produces CEPO from erythropoietin (EPO) requires pure EPO as raw material, is challenging to scale-up and can also cause batch-to-batch variability. To address these key limitations while retaining its behavioral effects, we designed, expressed and analyzed a triple, glutamine, substitution recombinant mimetic of CEPO, named QPO.
We employ a combination of computational structural biology, molecular, cellular and behavioral assays to design, produce, purify and test QPO.
QPO was shown to be a nonhematopoietic polypeptide with significant antidepressant-like and pro-cognitive behavioral effects in rodent assays while significantly upregulating BDNF expression in-vitro and in-vivo. The in-silico binding affinity analysis of QPO bound to the EPOR/EPOR homodimer receptor shows significantly decreased binding to Active Site 2, but not Active Site 1, of EPOR.
The results of the behavioral and gene expression analysis imply that QPO is a successful CEPO mimetic protein and potentially acts via a similar neurotrophic mechanism, making it a drug development target for psychiatric disorders. The decreased binding to Active Site 2 could imply that this active site is not involved in neuroactive signaling and could allow the development of a functional innate repair receptor (IRR) model. Substituting the three glutamine substitution residues with arginine (RPO) resulted in the loss of behavioral activity, indicating the importance of glutamine residues at those positions.
氨基甲酰化促红细胞生成素（CEPO）是一种经过化学修饰的、非造血衍生的促红细胞生成素（EPO），它保留了其抗抑郁和促认知的作用，这归因于在中枢神经系统中神经营养因子（如脑源性神经营养因子（BDNF））的表达增加。然而，从促红细胞生成素（EPO）中产生 CEPO 的化学修饰过程需要纯 EPO 作为原料，难以规模化，并且还可能导致批间变异性。为了解决这些关键限制，同时保留其行为效应，我们设计、表达和分析了一种三重、谷氨酰胺取代的 CEPO 重组模拟物，命名为 QPO。
我们采用计算结构生物学、分子、细胞和行为分析相结合的方法来设计、生产、纯化和测试 QPO。
QPO 被证明是一种非造血多肽，在啮齿动物试验中具有显著的抗抑郁样和促认知行为效应，同时显著上调体外和体内的 BDNF 表达。QPO 与 EPOR/EPOR 同源二聚体受体结合的计算机结合亲和力分析表明，与 EPOR 的活性位点 2结合显著降低，但与活性位点 1 结合没有降低。
行为和基因表达分析的结果表明，QPO 是一种成功的 CEPO 模拟蛋白，可能通过类似的神经营养机制发挥作用，使其成为精神疾病药物开发的靶点。与活性位点 2 的结合减少可能意味着该活性位点不参与神经活性信号传递，并可能允许开发功能性固有修复受体（IRR）模型。用精氨酸（RPO）替代三个谷氨酰胺取代残基会导致行为活性丧失，这表明这些位置的谷氨酰胺残基很重要。
