Microfluidic preparation of niosomes nanoparticles for intracellular delivery of anti c-myc siRNA for treatment of breast and melanoma cancers

RNA interference (RNAi) is a post-transcriptional gene regulatory mechanism that involves the degradation of a target messenger RNA (mRNA) through the incorporation of short interfering RNAs (siRNA).

The discovery of the RNAi mechanism made a significant impact in the field of drug delivery as this mechanism can be used to inhibit any protein of interest in the target cells.

The design of siRNA that target specific protein can be done easily based on the sequence of the amino acids in the target protein. However, unmodified, naked siRNA is unstable to penetrate the cell membrane.

Therefore, the application of siRNA based therapeutics is limited by the development of a delivery system to deliver therapeutic siRNA to the cytoplasm of the target cells and inhibit specific protein that is vital for the cellular growth.

In previous work, we have investigated the use of niosomes nanoparticles prepared by microfluidic mixing for siRNA delivery into melanoma and breast cancer cells.

Through the use of negative and fluorescently labelled siRNA, we have proved that the siRNA was successfully uptake by the cells through the use of fluorescent microscope and flow cytometry.

Moreover, we have used siRNA targeting green fluorescent protein (GFP) in a breast cancer cell line that are producing GFP and our niosomes preparation was successfully able to inhibit the production of GFP by delivering the anti-GFP siRNA to the cytoplasm of the GFP producing cancer cells. However, none of our previous work was using an active siRNA to target and inhibit the growth of the cancer cells.

Therefore, the aim of this project is to investigate the use of our successful niosomes nanoparticles prepared by microfluidic mixing for the delivery of an active siRNA into the cancer cells both in vitro and in vivo. C-myc is one of the vital proteins that is vital for the growth of the cancer cells.

If we were able to inhibit the c-myc production in the cancer cells, then the growth of the cancer cells will be stopped.

Our aim is to encapsulate anti c-myc siRNA into our niosomes nanoparticles and investigate the level of c-myc inhibition in both breast and melanoma cancer cells.

These in vitro assays will be done to evaluate those niosomes loaded with anti c-myc siRNA as a potential anti-cancer nanomedicine. This involve the characterization of siRNA-loaded niosomes.

Particle size and surface charge of the prepared niosomes will be measured by dynamic light scattering, particle morphology will be examined by transmission electron microscopy.

After encapsulating the anti c-myc siRNA into the niosomes, the cells will be treansfected with this formulation and the level of c-myc in the transfected cells will be evaluated using Western Blot (WB) and real time polymerase chain reaction (PCR) techniques.

In virto cellular growth will be monitored after treating the cells with the target anti c-myc siRNA and the mechanism of cellular death will be investigated.