In Vivo Fate of Avidin-Nucleic Acid Nanoassemblies as Multifunctional Diagnostic Tools

Abstract

This study describes the formulation optimization and body-cell distribution and clearance in mice of a dually fluorescent biodegradable poly avidin nanoassembly based on the novel Avidin-Nucleic-Acid-Nano-ASsembly (ANANAS) platform as a potential advancement of classic avidin/biotin-based targeted delivery. The nanoformulation circulates freely in the bloodstream; it is slowly captured by filter organs; it is efficiently cleared within 24-48 h, and it is poorly immunogenic. The system displays more favorable properties than its parent monomeric avidin and it is a promising tool for diagnostic purposes for future translational aims, for which free circulation in the bloodstream, safety, multifunctionality and high composition definition are all necessary requirements. In addition, the assembly shows a time-dependent cell penetration capability, suggesting it may also function as a NP-dependent drug delivery tool. The ease of preparation together with the possibility to fine-tune the surface composition makes it also an ideal candidate to understand if and how nanoparticle composition affects its localization.

Detection of fluorescent organic nanoparticles by confocal laser endomicroscopy in a rat model of Barrett’s esophageal adenocarcinoma

“…At the same time, they are small enough to navigate easily in the vasculature and cross barriers through small leaky capillaries to reach tumor cells by enhanced permeability and retention effect. [7][8][9][10] NPs offer large surface area, so that surface chemistry allows their physical and biological properties to be widely modified. In particular, they may have prolonged circulation time and additional modification of NPs with various "homing" molecules increases their affinity and specificity for tumor cells.…”

Targeted nanotherapeutics in cancer

“…Notably, the emerging new genomic knowledge has revolutionized molecular medicine, 11 and targeting with antibody therapeutics 6,12 and siRNA holds great promise as a potential new class of therapeutics with an ability to treat complex tumor types 13,14 that have, thus far, been resistant to available therapies. However, the lack of identified molecular biomarkers that could fully explain epigenetic changes and the heritability of complex tumors, 15 making it very difficult for target recognition and treatment, 16 should benefit from vigorous target validation 17,18 and other efforts [19][20][21][22][23] to develop new targeted drugs.…”

Detection of a fluorescent-labeled avidin-nucleic acid nanoassembly by confocal laser endomicroscopy in the microvasculature of chronically inflamed intestinal mucosa

“…The nanoparticles are capable of circulating for at least 2 hours before being eliminated within the next 2 days through classic scavenging systems with no apparent toxicity and low immunogenicity. 17 These findings paved the way for further exploitation of this nanoparticle tool for in vivo diagnostics and delivery applications. 18 The ANANAS platform is especially interesting because it also meets the current need for convenient and reproducible preparation methods for multifunctional nanocarriers.…”

“…In this study, red 681 -labeled ANANAS nanoparticles were synthesized in collaboration with ANANAS Nanotech (Padova, Italy) according to a procedure described elsewhere. 17 In detail, a highly PEGylated ANANAS nanoparticle formulation was prepared and stabilized by mixing plasmid DNA and a complex of avidin and compound 1 at 1:0.25 molar ratio. 17,29 After purification and ultravioletvisible characterization of the nanoassembly, biotin-C 6 -red 681 (compound 3) was added to at 1:3 avidin:biotin molar ratio.…”

“…17 In detail, a highly PEGylated ANANAS nanoparticle formulation was prepared and stabilized by mixing plasmid DNA and a complex of avidin and compound 1 at 1:0.25 molar ratio. 17,29 After purification and ultravioletvisible characterization of the nanoassembly, biotin-C 6 -red 681 (compound 3) was added to at 1:3 avidin:biotin molar ratio. Finally, the mixture was purified from low molecular weight derivatives by gel permeation chromatography using a Sepharose 6-FF column on a fast protein liquid chromatography system (AKTA purifier, GE Healthcare, Little Chalfont, UK) with 10 mM phosphate, 150 mM NaCl pH 7.4 (phosphatebuffered saline) as the eluent.…”