Liposomal Gene Delivery Systems.....pptx

The topic is :- Liposomal gene delivery system.
This topic is enough for B.Pharm. and M.Pharm. study purposes and notes.
As we know liposomes are small, spherical vesicles, are made from phospholipid bilayers and cholesterol.
Due to their small size, it is used to incorporate the desired DNA or genetic proteins and deliver them to their targeted organ or tissue.
This slide share tells us about how the DNA and liposome form a specific structure called lipofection, which is ultimately responsible for gene delivery to its particular site or organ, hence known as a targeted gene delivery system.
The Principle of formation of lipofection complex and the process how it will enters into the cell..

Classification of lipids on the basis of their charge present on the outer surface of the lipids and the charge present at the surface of DNA and how they react to each other to form a complex lipoplex, which is responsible for gene delivery.

1. Liposome Drug Delivery System UTTARANCHAL UNIVERSITY MADE BY PRAMESH PANWAR

2. Liposome :- Discovered in the year of 1960 by British hematologist Dr Alec D. Bangham . Liposome are the small, spherical vesicle that are formed/ made from phospholipids bilayer and cholesterol. Size 10-100nm Liposome have been explored as a delivery system for DNA as early as in 1979. The technique which is used to inject the genetic material into a cell by means of liposome is known as Lipofections or liposome transfection or lipoplex. Lipoplex act as a Non Viral type vector.

3. Principle:- Lipids have cationic charge (+ve) + DNA (-ve) charge A structure or complex is formed Liposome/Lipofection The complex is then enter into the cell through endocytosis. Once the cell enters inside the cell it diffuse through cytoplasm and enter the nucleus for gene expression Eg,. Animal cell, plant cells, Bacteria, yest, protoplast are susceptible to lipofection method. Liposome DNA Lipofection Complexation

4. Enhanced permeation and Retention effect

5. CLASSIFICATION OF LIPIDS.

6. Cationic Lipids:- Cationic lipids are those having +ve charge, which associate with the –vely charged DNA molecule by electrostatic interactions forming a stable complex and are used for the transfer of nucleic acid. Cationic head group appear to be better suited for DNA delivery due to attraction to the –vely charged phosphate group in DNA molecule aggregate the facilitation of the binding and delivery of the DNA. Its idea is to Neutralize charge of plasmid with positively charged lipids to capture plasmid more efficiently and to deliver DNA into cells. The Basic structure of cationic lipids mimics the chemical and physical attribution of biological lipids Here lipoplex is formed by combining the cationic lipids, neutral lipids and genetic material. CATIONIC LIPIDS

7. TYPES OF CATIONIC LIPIDS

8. DOTMA DOTMA :- Dioleoyl-3-trimethylamonium propane It is a synthesized cationic lipid, was discovered by “felgner et al” Structure :- 2 Unsaturated oleoyl acid chain 3 Carbon skeleton of a glycerol These two are connected with a ether bond and A quaternary amine as a cationic head group attached to the skeleton. Procedure:- Mix Cationic lipid + Neutral lipid to cells + DNA Formation of an aggregation compound i.e., Lipoplex

9. b) DOTAP :- Synthesized by Leventis and silivius 1990. The structure of DOTAP is similar to the DOTMA but the difference b/w DOTAP and DOTMA are that ester bond linked the chains to the backbone rather than ether bond. Liposomal sensitivity is 25-30% DOTAP is completely protonated at PH (7.4) so, that more energy is required to separate the DNA from the lipoplex for successful transfection. Thus, for DOTAP to be more effective in gene delivery it should be combined with a helper lipid In this, high temperature and long incubation time have been used to create lipoplex that exhibit resistance to serum interaction. DOTAP

10. DC -CHOL Gao and Huang in 1991 It contain cholesterol moiety attached by an ester bond to a hydrolyrable dimethyl ethylene diamine. Cholesterol – chosen – for its biocompatibility and stability. B) Multivalent Cationic Lipids:- DOSPA:- Its structure is similar to the DOTMA except for a spermine group bound via peptide bond to a hydrophobic chain. This cationic lipid, used with a Neutral helper lipid DOPE at a 3:1 ratio which is commerically available as the transfection regent Lipofectamine. b) DOGS :- I t is similar to the DOSPA However, chains are saturated and linked to the head group through a peptide bond and lack quat. Amine. Commercially named = Transfectam Its gene expression level = more than to fold as compared with others.

11. NEUTRAL LIPIDS Neutral lipids :- Most liposomal formulations used for gene delivery consist of a combination of charged lipids and neutral helper lipids It used DOPE and DOPC Neutral lipid stabilize the complex in serum and reduce toxicity. AN-IONIC LIPID A n-ionic lipid:- In general gene delivery by anionic lipids is not very efficient/ The –vely charged head group prevents efficient DNA compactions due to Repulsive electrostatic forces that occurs b/w the phosphate backbone of DNA and the anionic Head group of the lipids. Commonly used lipids in this category are phospholipids ex., phosphatidic acid.

12. Little toxicity, compaction with body fluids, complicated and time consuming process. Sometime, the DNA molecule get entrapped within the aqueous interior of these liposome. Then divalent cations, are used (Ca2+ , Mg2+ ) which can Neutralized the mutual electrostatic Repulsion. They are termed as Ph-Sensitive due to destabilization at low ph. Due to Reduced toxicity and interference from serum protein, ph-sensitive liposome are considered as potential gene therapy vehicles than the cationic liposome's.

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