ENCAPSULATION OF SHT-DNA, SIRNA AND POLYPEPTIDE-17 INSIDE HYBRID POLYMERICNANO- PROTEIN FOLICACID (HPNP-FA) CARRIER AS TARGETED TGF. INHIBITOR FOR HEPATOCELLULAR CARCINOMA

Background and Aims: Gene therapy is an important area in cancer treatment due to its specific selection for target. The issue of this approach is related to integrate successfully genetic materials since their size is larger than membranous pores and carriers can be degraded by endogenous nucleases. The strategy applied to solve this issue by inserting foreign DNA fragments into vector suffers from several drawbacks, since un-methylated CPG of bacterial DNA can elicit strong host immune responses. Nanotechnology has proven its ability to encapsulate DNA, siRNA, and P-17 and to functionalize nano-carrier surface to be selective for its target. Folic acid (FA) has good binding affinity to folate receptors and Bovine Serum Albumin (BSA) has special property to easily bind hepatocytes. Our aim is to create site-chemical conjugation of (FA-BSA) and use this complex to coat SHT-DNA, siRNA and P-17 attached with Polyethylene glycol (PEG).Methods: The activation of FA has been investigated by FTIR spectroscopy and spectrophotometry. Successful BSA-FA attachment was detected by X-ray diffraction and electrophoretic mobility of Agarose gel-FA bands. HPNP-FA colloidal stability was measured by photon correlation spectroscopy. SHT DNA, siRNA and P-17 entrapment was demonstrated by urinayl acetate -TEM. HPNP-FA topography was measured by AFM. Concentration of folic acid attached with HPNP carrier was quantified by using FA absorbance and Beer-Lambert Law. HPNP-FA cellular internalization was demonstrated by ELISA fluorescence spectrum Reader. The efficiency of SHT DNA, siRNA and P-17 cellular uptake was quantified by confocal microscopy. Results: Activated FA has shown good absorbance, high fluorescence intensity and revealed isolated bands compared to that one dissolved in MilliQ water. Modified carboxylic group improves affinity binding and active site of FA. FTIR and X-ray diffraction show successful conjugation of for BSA-FA. Photon correlation spectroscopy result indicates a good stability of HPNP-FA as colloidal suspension. ELISA reader of HCC micro plate confirms successful internalization of HPNP-FA. Uranyl acetate-TEM photomicrograph shows encapsulation of SHT DNA, siRNA and P-17 inside HPNP-FA layers. Confocal microscopy shows good efficiency of SHT DNA, siRNA and P-17. Conclusions: HPNP-FA demonstrated to be a specific target for HCC because high affinity binding of FA and BSA to folate receptor and cell membrane. Additionally by PEG it increases time circulation in blood stream.