Doxorubicin-Incorporated Nanotherapeutic Delivery System Based on Gelatin-Coated Gold Nanoparticles: Formulation, Drug Release, and Multimodal Imaging of Cellular Internalization

Abstract

In this work, we developed a new pH- and temperature-responsive nanochemotherapeutic system based on Doxorubicin (DOX) noncovalently bound to biosynthesized gelatin-coated gold nanoparticles (DOX-AuNPs@gelatin). The real-time release profile of DOX was evaluated at different pH values (7.4, 5.3, and 4.6) and temperatures (22-45 °C) in aqueous solutions, and its therapeutic performance was examined in vitro against MCF-7 breast cancer cells. TEM, dark-field scattering, and wide-field fluorescence microscopy indicated the effective uptake of nanochemotherapeutics with the subsequent release and progressive accumulation of DOX in cell nuclei. MTT assays clearly showed the effectiveness of the treatment by inhibiting the growth of MCF-7 breast cancer cells for a loaded drug concentration of 5 μg/mL. The most informative data about the dynamic release and localization were provided by scanning confocal microscopy using time-resolved fluorescence and surface-enhanced Raman scattering (SERS) techniques. In particular, fluorescence-lifetime imaging (FLIM) recorded under 485 nm pulsed diode laser excitation revealed the bimodal distribution of DOX in cells. The shorter fluorescence lifetime of DOX localized in nuclei (1.52 ns) than in the cytoplasm (2.4 ns) is consistent with the cytotoxic mechanism induced by DOX-DNA intercalation. Remarkably, the few DOX molecules captured between nanoparticles ("electromagnetic hotspots") after most drug is released act as SERS reporters for the localization of plasmonic nanocarriers in MCF-7 cells. The high drug loading capacity and effective drug release under pH control combined with the advantage of multimodal visualization inside cells clearly indicate the high potential of our DOX-AuNPs@gelatin delivery system for implementation in nanomedicine.

One‐Pot Synthesis of DOX@Covalent Organic Framework with Enhanced Chemotherapeutic Efficacy

“…The TAPB‐DMTP‐COF was synthesized by a condensation reaction between 1,3,5‐tris(4‐aminophenyl)benzene (TAPB) and 2,5‐dimethoxyterephthaldehyde (DMTP). DOX is a widely used anti‐cancer drug that kills tumor cells effectively and is easily characterized by UV/Vis spectroscopy . In an effort to encapsulate DOX into TAPB‐DMTP‐COF efficiently, DOX and DMTP were first mixed and stirred for 1 h, and then TAPB was added to complete the Schiff base reaction, which finally formed DOX@COF …”

“…DOX is aw idely used anti-cancer drug that kills tumor cells effectively and is easily characterized by UV/Vis spectroscopy. [24][25][26][27] In an effort to encapsulate DOX into TAPB-DMTP-COF efficiently,D OX and DMTP weref irst mixed and stirred for 1h,a nd then TAPB was added to complete the Schiff base reaction, which finally formed DOX@COF. [28,29] To prepare highly monodispersed particles with highc rystallinity,aseries of experiments were conducted.…”

Protein based therapeutic delivery agents: Contemporary developments and challenges

“…Astilean et al recently presented a nanochemotherapeutic system of gold nanoparticles coated with DOX-bound gelatin (AuNPs@gelatin) that was responsive to pH and temperature [87]. The design highlighted the functionality of imaging by combining fluorescence imaging techniques with confocal Raman microscopy, which efficiently monitored intracellular release and progressive accumulation of DOX [87]. In particular, the study utilized fluorescence lifetime imaging to confirm that the cytotoxic mechanism was induced by DOX-DNA interaction [87].…”

“…The design highlighted the functionality of imaging by combining fluorescence imaging techniques with confocal Raman microscopy, which efficiently monitored intracellular release and progressive accumulation of DOX [87]. In particular, the study utilized fluorescence lifetime imaging to confirm that the cytotoxic mechanism was induced by DOX-DNA interaction [87]. Despite that the complex did not necessarily show improvement in anticancer activity compared to free DOX in MCF-7 cells [87], such a strategy should be encouraged and further developed to gain insight into the mechanisms of delivery systems.…”

“…In particular, the study utilized fluorescence lifetime imaging to confirm that the cytotoxic mechanism was induced by DOX-DNA interaction [87]. Despite that the complex did not necessarily show improvement in anticancer activity compared to free DOX in MCF-7 cells [87], such a strategy should be encouraged and further developed to gain insight into the mechanisms of delivery systems.…”