Institute of Cell Biology, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, 1000, Ljubljana, Slovenia.
Clinical Institute of Genomic Medicine, University Medical Centre Ljubljana, Ljubljana, Slovenia.
Histochem Cell Biol. 2023 Nov;160(5):435-452. doi: 10.1007/s00418-023-02224-2. Epub 2023 Aug 3.
Urinary bladder cancer can be treated by intravesical application of therapeutic agents, but the specific targeting of cancer urothelial cells and the endocytotic pathways of the agents are not known. During carcinogenesis, the superficial urothelial cells exhibit changes in sugar residues on the apical plasma membranes. This can be exploited for selective targeting from the luminal side of the bladder. Here we show that the plant lectins Jacalin (from Artocarpus integrifolia), ACA (from Amaranthus caudatus) and DSA (from Datura stramonium) selectively bind to the apical plasma membrane of low- (RT4) and high-grade (T24) cancer urothelial cells in vitro and urothelial tumours ex vivo. The amount of lectin binding was significantly different between RT4 and T24 cells. Endocytosis of lectins was observed only in cancer urothelial cells and not in normal urothelial cells. Transmission electron microscopy analysis showed macropinosomes, endosome-like vesicles and multivesicular bodies filled with lectins in RT4 and T24 cells and also in cells of urothelial tumours ex vivo. Endocytosis of Jacalin and ACA in cancer cells was decreased in vitro after addition of inhibitor of macropinocytosis 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and increased after stimulation of macropinocytosis with epidermal growth factor (EGF). Clathrin, caveolin and flotillin did not colocalise with lectins. These results confirm that the predominant mechanism of lectin endocytosis in cancer urothelial cells is macropinocytosis. Therefore, we propose that lectins in combination with conjugated therapeutic agents are promising tools for improved intravesical therapy by targeting cancer cells.
膀胱癌可以通过膀胱内应用治疗剂进行治疗，但癌症尿路上皮细胞的特异性靶向和药物的内吞途径尚不清楚。在癌变过程中，表面尿路上皮细胞在上皮细胞顶膜上的糖残基发生变化。这可以被利用来从膀胱的腔侧进行选择性靶向。在这里，我们表明植物凝集素 Jacalin（来自 Artocarpus integrifolia）、ACA（来自 Amaranthus caudatus）和 DSA（来自 Datura stramonium）在体外选择性结合低级别（RT4）和高级别（T24）癌症尿路上皮细胞和尿路上皮肿瘤的顶膜。RT4 和 T24 细胞之间的凝集素结合量有显著差异。只有在癌症尿路上皮细胞中观察到凝集素的内吞作用，而在正常尿路上皮细胞中则没有。透射电子显微镜分析显示，在 RT4 和 T24 细胞以及离体的尿路上皮肿瘤细胞中，存在填充有凝集素的巨胞饮体、内体样小泡和多泡体。在添加巨胞饮体抑制剂 5-(N-乙基-N-异丙基)amiloride（EIPA）后，癌症细胞中 Jacalin 和 ACA 的内吞作用在体外减少，而在用表皮生长因子（EGF）刺激巨胞饮作用后增加。网格蛋白、窖蛋白和 flotillin 与凝集素没有共定位。这些结果证实，凝集素在癌症尿路上皮细胞中的主要内吞作用机制是巨胞饮作用。因此，我们提出，凝集素与共轭治疗剂结合是通过靶向癌细胞来提高膀胱内治疗的有前途的工具。
