Delivery of htsFLT01 to Y79 Cells Using MiRGD Peptide and Graphene Quantum Dots
Sina Goli garmestani1 , Zahra-Soheila Soheili 1 *, Saman Hosseinkhani 2 , Hamid Ahmadieh 3 , Hamid Latifi-Navid 1 , Somayeh Piroozmand 1 , Naeimeh Bayatkhani 1 , Shahram Samiei 4
- Department of Molecular Medicine, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran
- Department of Nanobiotechnology, Faculty of Biological Sciences,Tarbiat Modares University, Tehran, Iran
- Ophthalmic Research Center, Research Institute for Ophthalmology and Vision Science, Shahid Beheshti University of Medical Sciences, Tehran, Iran
- Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran
Abstract: Retinoblastoma is a prevalent intraocular malignancy in children. Vascular Endothelial Growth Factor (VEGF) serves as a key proangiogenic factor secreted by nearly all solid tumors. The htsFLT01 fusion protein effectively neutralizes both mouse and human VEGF and PlGF. MiRGD peptides, containing the iRGD motif, enhance tissue penetration by binding to αv integrins in cancerous tissues. Graphene Quantum Dots (GQDs) have emerged as promising agents in bio-imaging due to their biocompatibility, low cytotoxicity, and tunable fluorescence properties.
Methods: In this study, htsFLT01 plasmids were prepared using the Favorgen Maxi preparation kit. MiRGD peptides were purified through Ni-NTA chromatography, followed by desalting via dialysis. GQDs were synthesized via the hydrothermal method using citric acid and urea, and their absorption and emission wavelengths (330 nm and 440 nm, respectively) were confirmed via BioTek Cytation. The functional surface groups of GQDs were characterized by FTIR, and their size and charge were analyzed using Dynamic Light Scattering (DLS). Gel retardation assays demonstrated stable complex formation, further confirmed by agarose gel electrophoresis with ethidium bromide staining.
Results: Initial DLS results indicated the zeta potentials of GQDs, MiRGDs, and the complexes to be -23 mV, +6 mV, and +11 mV, respectively. Preliminary cell culture experiments were conducted on Y79 human retinoblastoma cells maintained in RPMI 1640 medium with 10% FBS and 1% penicillin/streptomycin under standard incubation conditions.
Conclusion: Future experiments will focus on optimizing the nano-complex treatment's dosage and timing through MTT assays, followed by molecular analyses including apoptosis, flow cytometry and real-time PCR.
