Kalousková Barbora, Skořepa Ondřej, Cmunt Denis, Abreu Celeste, Krejčová Kateřina, Bláha Jan, Sieglová Irena, Král Vlastimil, Fábry Milan, Pola Robert, Pechar Michal, Vaněk Ondřej
Department of Biochemistry, Faculty of Science, Charles University, Hlavova 2030, 12840 Prague, Czech Republic.
Institute of Molecular Genetics, Czech Academy of Sciences, Vídeňská 1083, 14220 Prague, Czech Republic.
Biomedicines. 2021 Nov 2;9(11):1597. doi: 10.3390/biomedicines9111597.
Targeted cancer immunotherapy is a promising tool for restoring immune surveillance and eradicating cancer cells. Hydrophilic polymers modified with coiled coil peptide tags can be used as universal carriers designed for cell-specific delivery of such biologically active proteins. Here, we describe the preparation of pHPMA-based copolymer conjugated with immunologically active protein B7-H6 via complementary coiled coil VAALEKE (peptide E) and VAALKEK (peptide K) sequences. Receptor B7-H6 was described as a binding partner of NKp30, and its expression has been proven for various tumor cell lines. The binding of B7-H6 to NKp30 activates NK cells and results in Fas ligand or granzyme-mediated apoptosis of target tumor cells. In this work, we optimized the expression of coiled coil tagged B7-H6, its ability to bind activating receptor NKp30 has been confirmed by isothermal titration calorimetry, and the binding stoichiometry of prepared chimeric biopolymer has been characterized by analytical ultracentrifugation. Furthermore, this coiled coil B7-H6-loaded polymer conjugate activates NK cells and, in combination with coiled coil scFv, enables their targeting towards a model tumor cell line. Prepared chimeric biopolymer represents a promising precursor for targeted cancer immunotherapy by activating the cytotoxic activity of natural killer cells.
靶向癌症免疫疗法是恢复免疫监视和根除癌细胞的一种有前景的工具。用卷曲螺旋肽标签修饰的亲水性聚合物可用作通用载体，用于细胞特异性递送此类生物活性蛋白。在此，我们描述了通过互补卷曲螺旋VAALEKE（肽E）和VAALKEK（肽K）序列与免疫活性蛋白B7-H6共轭的基于聚（N-（2-羟丙基）甲基丙烯酰胺）（pHPMA）共聚物的制备。受体B7-H6被描述为NKp30的结合伴侣，并且已在各种肿瘤细胞系中证实其表达。B7-H6与NKp30的结合激活NK细胞，并导致Fas配体或颗粒酶介导的靶肿瘤细胞凋亡。在这项工作中，我们优化了卷曲螺旋标记的B7-H6的表达，通过等温滴定量热法证实了其与激活受体NKp30结合的能力，并通过分析超速离心法表征了制备的嵌合生物聚合物的结合化学计量。此外，这种负载卷曲螺旋B7-H6的聚合物缀合物激活NK细胞，并与卷曲螺旋单链抗体片段（scFv）结合，使其能够靶向模型肿瘤细胞系。制备的嵌合生物聚合物通过激活自然杀伤细胞的细胞毒性活性，代表了靶向癌症免疫疗法的一种有前景的前体。
