Federal Research Center of Fundamental and Translational Medicine (FRC FTM), Timakova str., 2, 630117 Novosibirsk, Russia.
N.N. Vorozhtsov Novosibirsk Institute of Organic Chemistry SB RAS, Lavrentiev Ave., 9, 630090 Novosibirsk, Russia.
Molecules. 2023 Dec 8;28(24):8014. doi: 10.3390/molecules28248014.
Recombinant human interferon alpha-2b (rIFN) is widely used in antiviral and anticancer immunotherapy. However, the high efficiency of interferon therapy is accompanied by a number of side effects; this problem requires the design of a new class of interferon molecules with reduced cytotoxicity. In this work, IFN was modified via genetic engineering methods by merging it with the blood plasma protein apolipoprotein A-I in order to reduce acute toxicity and improve the pharmacokinetics of IFN. The chimeric protein was obtained via biosynthesis in the yeast . The yield of ryIFN-ApoA-I protein when cultivated on a shaker in flasks was 30 mg/L; protein purification was carried out using reverse-phase chromatography to a purity of 95-97%. The chimeric protein demonstrated complete preservation of the biological activity of IFN in the model of vesicular stomatitis virus and SARS-CoV-2. In addition, the chimeric form had reduced cytotoxicity towards Vero cells and increased cell viability under viral load conditions compared with commercial IFN-a2b preparations. Analysis of the pharmacokinetic profile of ryIFN-ApoA-I after a single subcutaneous injection in mice showed a 1.8-fold increased half-life of the chimeric protein compared with ryIFN.
重组人干扰素 alpha-2b(rIFN) 被广泛用于抗病毒和抗癌免疫治疗。然而，干扰素治疗的高效性伴随着许多副作用；这个问题需要设计一类新的干扰素分子，降低其细胞毒性。在这项工作中，通过基因工程方法将 IFN 与血浆蛋白载脂蛋白 A-I 融合，从而降低急性毒性并改善 IFN 的药代动力学。通过在酵母中生物合成获得嵌合蛋白。在摇瓶中摇床培养时，ryIFN-ApoA-I 蛋白的产量为 30mg/L；通过反相色谱法纯化至 95-97%的纯度。嵌合蛋白在水疱性口炎病毒和 SARS-CoV-2 的模型中完全保留了 IFN 的生物学活性。此外，与商业 IFN-a2b 制剂相比，嵌合形式对 Vero 细胞的细胞毒性降低，在病毒载量条件下细胞活力增加。对小鼠单次皮下注射后 ryIFN-ApoA-I 的药代动力学特征进行分析表明，与 ryIFN 相比，嵌合蛋白的半衰期延长了 1.8 倍。
