Selective Capture and Purification of MicroRNAs and Intracellular Proteins through Antisense-vectorized Magnetic Nanobeads

Abstract

MicroRNAs (miRNAs) are small non-coding nucleotides playing a crucial role in posttranscriptional expression and regulation of target genes in nearly all kinds of cells. In this study, we demonstrate a reliable and efficient capture and purification of miRNAs and intracellular proteins using magnetic nanoparticles functionalized with antisense oligonucleotides. For this purpose, a tumor suppressor miRNA (miR-198), deregulated in several human cancer types, was chosen as the model oligonucleotide. Magnetite nanoparticles carrying the complementary sequence of miR-198 (miR-198 antisense) on their surface were delivered into cells and subsequently used for the extracellular transport of miRNA and proteins. The successful capture of miR-198 was demonstrated by isolating RNA from magnetic nanoparticles followed by real-time PCR quantification. Our experimental data showed that antisense-coated particles captured 5-fold higher amounts of miR-198 when compared to the control nanoparticles. Moreover, several proteins that could play a significant role in miR-198 biogenesis were found attached to miR-198 conjugated nanoparticles and analyzed by mass spectrometry. Our findings demonstrate that a purpose-driven vectorization of magnetic nanobeads with target-specific recognition ligands is highly efficient in selectively transporting miRNA and disease-relevant proteins out of cells and could become a reliable and useful tool for future diagnostic, therapeutic and analytical applications.

Magnetic Gold Nanoparticles with Idealized Coating for Enhanced Point‐Of‐Care Sensing

“…While Fe-O bonds are visible in all spectra at 568 cm −1 , MNP exhibited a broad shoulder between 700 to 1100 cm −1 which can be attributed to C-H bending and C-N stretching of PEI. [28,29] Additionally, N-H and O-H bending at 1655 and 3500 cm −1 supports the presence of PEI and adsorbed water molecules on the MNP surface. Upon addition of citrate-capped AuNPs, additional bands appeared for MGNPs-PEI at 1080 cm −1 indicative of C-O bonds of citric acid.…”

Gene Therapy in Cancer Treatment: Why Go Nano?

“…Delivery of TNAs, such as genes, oligonucleotides, miRNAs or siRNAs to cancer cells has allowed to tackle cancer via the silencing oncogenes or restoring the expression of tumor suppressor genes [8][9][10][11][12][13][14][15][16][17][18][19]. Most of these approaches (e.g., antisense therapy, RNA interference (RNAi), gene editing) aim at gene alteration/modulation [16][17][18][19][76][77][78][79][80][81] -see Figure 3. The immunization gene therapies, particularly chimeric antigen receptor (CAR) in T cells (CAR-T cells) based therapies, stand-out since they represent the higher number of therapeutic strategies in clinical practice.…”

Non-coding RNAs and their bioengineering applications for neurological diseases

“…Such proof-of-concept studies have been performed in a relatively homogeneous system without the interference of blood, cellular factors, complex tissue architecture etc., and thus it should be noted that the number of studies performed in natural body fluids is highly limited. The ability of concentrating circulating miRNAs on the MNPs by surface conjugation or functionalization approaches has resulted in a number of highly efficient miRNA sensing devices [ 212 , 213 ]. Cationic polyurethance (PU) short branch polyethylene imine (PEI) nanocomplexes have been developed as vehicles for miR-145 delivery to inhibit brain tumor by targeting genes such as Sox4 and Oct4, in vitro [ 214 ].…”