Moreira David, Lopes-Nunes Jéssica, Santos Fátima Milhano, Campello Maria Paula Cabral, Oliveira Maria Cristina, Paulo António, Tomaz Cândida, Cruz Carla
CICS-UBI-Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilhã, Portugal.
Functional Proteomics Laboratory, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Calle Darwin 3, Campus de Cantoblanco, 28049 Madrid, Spain.
Pharmaceuticals (Basel). 2023 May 16;16(5):751. doi: 10.3390/ph16050751.
AT11-L0 is an aptamer derivative of AS1411 composed of G-rich sequences that can adopt a G-quadruplex (G4) structure and target nucleolin (NCL), a protein that acts as a co-receptor for several growth factors. Hence, this study aimed to characterize the AT11-L0 G4 structure and its interaction with several ligands for NCL targeting and to evaluate their capacity to inhibit angiogenesis using an in vitro model. The AT11-L0 aptamer was then used to functionalize drug-associated liposomes to increase the bioavailability of the aptamer-based drug in the formulation. Biophysical studies, such as nuclear magnetic resonance, circular dichroism, and fluorescence titrations, were performed to characterize the liposomes functionalized with the AT11-L0 aptamer. Finally, these liposome formulations with the encapsulated drugs were tested on the human umbilical vein endothelial cell (HUVEC) model to assess their antiangiogenic capacity. The results showed that the AT11-L0 aptamer-ligand complexes are highly stable, presenting melting temperatures from 45 °C to 60 °C, allowing for efficient targeting of NCL with a in the order of nM. The aptamer-functionalized liposomes loaded with ligands C and dexamethasone did not show cytotoxic effects in HUVEC cells compared with the free ligands and AT11-L0, as assessed by cell viability assays. AT11-L0 aptamer-functionalized liposomes encapsulating C and dexamethasone did not present a significant reduction in the angiogenic process when compared with the free ligands. In addition, AT11-L0 did not show anti-angiogenic effects at the concentrations tested. However, C shows potential as an angiogenesis inhibitor, which should be further developed and optimized in future experiments.
AT11-L0是AS1411的适体衍生物，由富含G的序列组成，可形成G-四链体（G4）结构，并靶向核仁素（NCL），NCL是一种作为多种生长因子共受体的蛋白质。因此，本研究旨在表征AT11-L0的G4结构及其与几种用于靶向NCL的配体的相互作用，并使用体外模型评估它们抑制血管生成的能力。然后，AT11-L0适体被用于使与药物相关的脂质体功能化，以提高制剂中基于适体的药物的生物利用度。进行了诸如核磁共振、圆二色性和荧光滴定等生物物理研究，以表征用AT11-L0适体功能化的脂质体。最后，在人脐静脉内皮细胞（HUVEC）模型上测试了这些包封药物的脂质体制剂，以评估它们的抗血管生成能力。结果表明，AT11-L0适体-配体复合物高度稳定，解链温度在45℃至60℃之间，能够以纳摩尔级的亲和力有效靶向NCL。通过细胞活力测定评估，与游离配体和AT11-L0相比，负载配体C和地塞米松的适体功能化脂质体在HUVEC细胞中未显示细胞毒性作用。与游离配体相比，包封C和地塞米松的AT11-L0适体功能化脂质体在血管生成过程中没有显著降低。此外，在测试浓度下，AT11-L0没有显示出抗血管生成作用。然而，C显示出作为血管生成抑制剂的潜力，应在未来实验中进一步开发和优化。
