Faculty of Biotechnology, Department of Protein Engineering, University of Wroclaw, Joliot-Curie 14a, 50-383 Wroclaw, Poland.
Int J Mol Sci. 2022 Jul 31;23(15):8514. doi: 10.3390/ijms23158514.
Precise anticancer therapies employing cytotoxic conjugates constitute a side-effect-limited, highly attractive alternative to commonly used cancer treatment modalities, such as conventional chemotherapy, radiotherapy or surgical interventions. Receptor tyrosine kinases are a large family of N-glycoproteins intensively studied as molecular targets for cytotoxic conjugates in various cancers. At the cell surface, these receptors are embedded in a dense carbohydrate layer formed by numerous plasma membrane glycoproteins. The complexity of the cell surface architecture is further increased by galectins, secreted lectins capable of recognizing and clustering glycoconjugates, affecting their motility and activity. Cell surface N-glycosylation is intensively remodeled by cancer cells; however, the contribution of this phenomenon to the efficiency of treatment with cytotoxic conjugates is largely unknown. Here, we evaluated the significance of N-glycosylation for the internalization and toxicity of conjugates targeting two model receptor tyrosine kinases strongly implicated in cancer: HER2 and FGFR1. We employed three conjugates of distinct molecular architecture and specificity: Affibody-vcMMAE (targeting HER2), vcMMAE-KCK-FGF1.E and T-Fc-vcMMAE (recognizing different epitopes within FGFR1). We demonstrated that inhibition of N-glycosylation reduced the cellular uptake of all conjugates tested and provided evidence for a role of the galectin network in conjugate internalization. In vitro binding studies revealed that the reduced uptake of conjugates is not due to impaired HER2 and FGFR1 binding. Importantly, we demonstrated that alteration of N-glycosylation can affect the cytotoxic potential of conjugates. Our data implicate a key role for cell surface N-glycosylation in the delivery of cytotoxic conjugates into cancer cells.
精准的抗癌疗法采用细胞毒性偶联物，为癌症的常用治疗方法（如传统化疗、放疗或手术干预）提供了一种副作用受限、极具吸引力的替代方案。受体酪氨酸激酶是一个大型 N-糖蛋白家族，作为各种癌症中细胞毒性偶联物的分子靶点，受到了深入研究。在细胞表面，这些受体嵌入在由许多质膜糖蛋白形成的密集碳水化合物层中。半乳糖凝集素（能够识别和聚集糖缀合物、影响其运动和活性的分泌型凝集素）进一步增加了细胞表面结构的复杂性。癌细胞强烈重塑细胞表面 N-糖基化；然而，这种现象对细胞毒性偶联物治疗效率的贡献在很大程度上仍是未知的。在这里，我们评估了 N-糖基化对靶向两种强烈参与癌症的模型受体酪氨酸激酶（HER2 和 FGFR1）的偶联物内化和毒性的重要性。我们使用了三种具有不同分子结构和特异性的偶联物：靶向 HER2 的 Affibody-vcMMAE、靶向 FGFR1 不同表位的 vcMMAE-KCK-FGF1.E 和 T-Fc-vcMMAE。我们证明了 N-糖基化的抑制减少了所有测试偶联物的细胞内摄取，并为半乳糖凝集素网络在偶联物内化中的作用提供了证据。体外结合研究表明，偶联物摄取减少不是由于 HER2 和 FGFR1 结合受损所致。重要的是，我们证明了 N-糖基化的改变会影响偶联物的细胞毒性潜力。我们的数据表明，细胞表面 N-糖基化在将细胞毒性偶联物递送到癌细胞中起着关键作用。
