Design and Synthesis of Piperazine-Based Compounds Conjugated to Humanized Ferritin as Delivery System of siRNA in Cancer Cells

Abstract

Gene expression regulation by small interfering RNA (siRNA) holds promise in treating a wide range of diseases through selective gene silencing. However, successful clinical application of nucleic acid-based therapy requires novel delivery options. Herein, to achieve efficient delivery of negatively charged siRNA duplexes, the internal cavity of “humanized” chimeric Archaeal ferritin (HumAfFt) was specifically decorated with novel cationic piperazine-based compounds (PAs). By coupling these rigid-rod-like amines with thiol-reactive reagents, chemoselective conjugation was efficiently afforded on topologically selected cysteine residues properly located inside HumAfFt. The capability of PAs-HumAfFt to host and deliver siRNA molecules through human transferrin receptor (TfR1), overexpressed in many cancer cells, was explored. These systems allowed siRNA delivery into HeLa, HepG2, and MCF-7 cancer cells with improved silencing effect on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene expression with respect to traditional transfection methodologies and provided a promising TfR1-targeting system for multifunctional siRNA delivery to therapeutic applications.

Simultaneous quantification of oligo-nucleic acids and a ferritin nanocage by size-exclusion chromatography hyphenated to inductively coupled plasma mass spectrometry for developing drug delivery systems

“…Recently, some studies, on the applications consisting of NADs utilizing the natural character of ferritin have been reported in the eld of cancer chemotherapy. 20,23 In addition, features of site-specic transport of ferritin have also been reported in experiments using mouse models. 24,25 In order to efficiently develop medicines consisting of a complex of NADs and protein-based DDS carriers (such as ferritin), a proper analytical methodology is necessary to evaluate the quality of the candidates.…”

“…The conventional methodology based on absorptiometry, which quanties both the NAD and protein-based DDS carriers separately, is cumbersome and unreliable. 22,23 In the case that both NAD-and protein-based DDS carriers are contained in a single analytical sample for measurement, quantied values will be uncertain because their absorption spectra must be overlapped. 26 Therefore, in most cases, a dissociation process to isolate the NAD from the carrier is needed.…”

Advances in Biomimetic Nanoparticles for Targeted Cancer Therapy and Diagnosis

“…Small interfering RNAs (siRNAs) have long been shown to be able to selectively silence gene expression, especially in cancer therapy [ 46 ]. Pediconi et al [ 47 ] synthesised a piperazine-based compound conjugated to chimeric archaeal ferritin loaded with siRNA, called siRNA-Pas-HumAfFt nanoparticles. Due to the nature of the negatively charged siRNA, the incorporation of piperazine-based compounds into chimeric archaeal ferritin was conducted to increase the positive charges which would promote spontaneous binding with siRNA molecules.…”

Bioengineered Protein Nanocage by Small Heat Shock Proteins Delivering mTERT siRNA for Enhanced Colorectal Cancer Suppression

“…26,27 The protein nanocages have shown some advantages as drug nanocarriers, including monodisperse size, strictly ordered nanostructure, good biocompatibility and biodegradability, convenient functional interface (interior encapsulation and exterior display), and easy modifications by the genetic or chemical method. 28 Therefore, several proteins have been chosen to design a siRNA delivery platform to tumor cells, such as ferritin, 29 thermosome, 30 small heat shock proteins (Hsp), 31 and virus capsid proteins, 32 suggesting high efficiency for siRNA delivery.…”

“…Nanotechnology has opened a new era for siRNA delivery to tumor tissues. , Nanocarriers could protect siRNA from digestion and improve delivery efficiency to cancer cells due to the enhanced permeability and retention (EPR) effect in tumor tissues. , Among various nanocarriers, protein nanocages attract considerable interest as an ideal platform because of their unique structural properties. , The protein nanocages have shown some advantages as drug nanocarriers, including monodisperse size, strictly ordered nanostructure, good biocompatibility and biodegradability, convenient functional interface (interior encapsulation and exterior display), and easy modifications by the genetic or chemical method . Therefore, several proteins have been chosen to design a siRNA delivery platform to tumor cells, such as ferritin, thermosome, small heat shock proteins (Hsp), and virus capsid proteins, suggesting high efficiency for siRNA delivery.…”