Micro‐scale RNA Interference using Iron Oxide Nanoparticle‐modified Lentivirus

Abstract

The US Food and Drug Administration (FDA) has approved the use of virotherapy. However, significant improvements are anticipated if effective localized infection can be achieved. Herein, we demonstrate magnetic lentivirus is guided by magnetic fields. Introduction of short hairpin RNA (shRNA) into the lentivirus genome enabled micro-scale RNA interference (RNAi) therapy. Although epidermal growth factor receptor (EGFR)-targeted therapies have successfully treated individuals with non-small cell lung cancer (NSCLC) cells, clinical therapeutic outcomes have been compromised by acquired resistance to first-generation tyrosine kinase inhibitors. We assess magnetic lentivirus with NSCLC cells using an in vitro model and achieve micro-scale RNAi through EGFR silencing. This proof-of-principle study demonstrates guided and highly localized micro-scale, RNAi virotherapy.

Magnetically Guided Viral Transduction of Gene-Based Sensitization for Localized Photodynamic Therapy To Overcome Multidrug Resistance in Breast Cancer Cells

“…We further desired to identify whether the ROS generation led to cell death and, if so, if it was due to apoptosis. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was used to identify apoptotic cells. − Consistent with the observed CellROX-positive CDR MCF-7 cells infected by ironized AAV2 (Figure ), a small amount of TUNEL fluorescence in the cells treated with AAV2 or ironized AAV2 was observed (Figure A). After the addition of the magnetic field, the TUNEL staining resulted in a significant increase in fluorescence from CDR MCF-7 cells.…”

“…With data confirming the improved viral transduction of CDR MCF-7 cells, we performed light-triggered virotherapy utilizing AAV2-KR with a corresponding wavelength of 561 nm. We further chose an irradiation time of 20 min for optimized ROS generation. ,, The generation of ROS was visualized in the photoactivated KillerRed-expressed cells in CDR MCF-7 or MCF-7 cell cultures with CellROX staining. In the cells treated with the ironized AAV2-KR ± magnetic field, the ironized coating had a positive effect on the ROS generation with a pronounced improvement in CDR MCF-7 compared to virus only (Figure A) due to the enhanced cellular uptake of viral particles.…”

“…10 4 CDR MCF-7 or MCF-7 cells were seeded in each of the wells of a 96 well plate and cultured with complete medium for 12 h. The cells were then exposed to DOX at various concentrations (0.5, 1,5, 10, 25, 50, or 100 μg mL –1 ) and incubated for 24 h. The CellTiter 96 AQueous one solution cell proliferation assay system with the tetrazolium compound (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt; MTS) was used to measure the survival rate and proliferation of mammalian cells. , The optical density (OD) of formazan at 490 nm quantified the cell viability. The reduction of MTS achieved by untreated cells was set at 100%, and that of test cells was expressed as a percentage of untreated cells.…”

“…A variety of delivery concepts have emerged for innovative treatment in cancer therapy largely attempting to improve specificity for drugs and proteins but also in gene-based concepts. Specificity can be opportunistically achieved via exploiting tumor microenvironment physicochemical properties, such as pH, for example. − However, these are dependent on these properties being exhibited by a solid tumor volume. To guide delivery independent of tumor characteristics, magnetic nanoparticles (MNP) conjugated to a carrier or vector provide accelerated accumulation in target cells when directed with magnetic fields .…”

“…or RNA interference (RNAi) . In the case of PDT, irradiation with a specific light bandwidth generates reactive oxygen species leading to irreversible DNA damage and cell killing via an apoptotic pathway; , offering additional options in cancer treatment. Here, we present an improvement to this previous work that promotes viral transduction in chemotherapy drug resistant MCF-7 ( CDR MCF-7) cells under magnetic field and express the KillerRed (KR) protein in transduced cells for light-triggered virotherapy (AAV2-KR) (Figure ).…”