Presentation1

Liposome: Novel Drug Delivery SystemLiposome: Novel Drug Delivery System

Jignesh PatelJignesh Patel M. Pharm.(sem-3)M. Pharm.(sem-3)

NPC,visnagarNPC,visnagar

Lipids, along with proteins and nucleic acids, are essential biomolecules for the structure and function of living matter

Liposomes are spherical self-closed structures, composed of curved lipid bilayers, which enclose part of the surrounding solvent into their interior. The size of a liposome ranges from some 20 nm up to severalmicrometers and they may be composed of one or several concentric membranes, each with a thickness of about 4 nm.

Liposomes possess unique properties owing to the amphiphilic character of the lipids, which make them suitable for drug delivery.

Formation of such a typical structural configuration is attributed to the amphiphilic character of phospholipids. When the letter are dispersed in excess of an aqueous phase, hydration of the polar head groups of the lipid results in a heterogeneous mixture of closed structures.

What is a liposome?What is a liposome?

What is a liposome?What is a liposome?

– Spherical vesicles with a phospholipid bilayerSpherical vesicles with a phospholipid bilayer

Hydrophilic

Hydrophobic

FundamentalFundamental properties: properties:

1. Size:1. Size: SUV: Small unilamellar vesicles 0.02-0.05 µµ LUV: Large unilamellar vesicles 0.1-0.5 µµ MLV: Multilamellar vesicles 1.0- >10 µµ

2. Fluidity:2. Fluidity:

3. Surface properties:3. Surface properties:

Size determined by methods:Size determined by methods:

Sonication: SUV Smaller than 100 nm diameterExtrusion: LUV (Size depends on the filters) 100 nm—1 µm diameterEvaporation: MUV Larger than 1 µm diameter

Materials used for preparation:Materials used for preparation:

Natural PhospholipidsNatural Phospholipids Synthetic PhospholipidsSynthetic Phospholipids SphingolipidsSphingolipids SteroidsSteroids Polymeric materialsPolymeric materials Charge lipidsCharge lipids

Phospholipids:Phospholipids:Polar Head Groups

Three carbon glycerol

Each phospholipidEach phospholipidincludesincludes aa polar polar region: region:

glycerolglycerol, , carbonylcarbonyl of of fatty acids, fatty acids, PPii, & the , & the polar head group (polar head group (XX))

2 2 non-polarnon-polar hydrocarbon hydrocarbon tails of fatty acids (tails of fatty acids (RR11, , RR22). ).

Such an Such an amphipathicamphipathic lipid lipidmay be represented as atmay be represented as atright.right.

O P O

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

X

glycerophospholipid

3D picture of a Phospholipid

PreparationPreparation of liposomes:of liposomes:

Selection of bilayer component is imp from toxicity as well as shelf life Selection of bilayer component is imp from toxicity as well as shelf life optimization point of view.optimization point of view.

Common process steps for liposome formulation:Common process steps for liposome formulation:

- Bilayer components are mixed in a volatile organic solvent or Bilayer components are mixed in a volatile organic solvent or solvent mixtures (eg. Chloroform, ether, EtOH, or combination solvent mixtures (eg. Chloroform, ether, EtOH, or combination of these)of these)

- Cholesterol upto 40-50 mol % is included to provide greater Cholesterol upto 40-50 mol % is included to provide greater stability in biological fluids.stability in biological fluids.

- A charged species may be added (5-20%) to prevent A charged species may be added (5-20%) to prevent aggregation. (natural acidic phospholipids eg PS, PG, PI, PA, aggregation. (natural acidic phospholipids eg PS, PG, PI, PA, etc)etc)

- Small amt of antioxidants such as aplha-tocopherol are included Small amt of antioxidants such as aplha-tocopherol are included when polyunsaturated natural lipids are used.when polyunsaturated natural lipids are used.

- Once a suitable solution of the lipid component is made, the Once a suitable solution of the lipid component is made, the mixture may be filtered to remove minor insoluble components mixture may be filtered to remove minor insoluble components or ultra filtered to lower pyrogens.or ultra filtered to lower pyrogens.

- The solvent is subsequently removed under conditions The solvent is subsequently removed under conditions (Pressure and Temp) that ensure no phase separations of the (Pressure and Temp) that ensure no phase separations of the component of the mixture take place.component of the mixture take place.

Preparation of lipid for hydration: Preparation of lipid for hydration:

Methods to check the Methods to check the morphology of liposomes:morphology of liposomes:

Freeze-fracture electron microscope Freeze-fracture electron microscope (FFEM) or electron microscope (EM) (FFEM) or electron microscope (EM)

Light scattering (LS)Light scattering (LS)

Methods to control vesicle Methods to control vesicle size of liposomes:size of liposomes:

CentrifugationCentrifugation Size-exclusion chromatographySize-exclusion chromatography HomogenizationHomogenization ExtrusionExtrusion

CharacterizationCharacterization of liposomes:of liposomes:

1. Physical Parameters:1. Physical Parameters: SizeSize Number of LamellaeNumber of Lamellae ChargeCharge Bilayer fluidityBilayer fluidity Encapsulated volumeEncapsulated volume Drug ReleaseDrug Release Phase behavior Phase behavior

2. Chemical Parameters:2. Chemical Parameters:

Quantitative determination of phospholipidQuantitative determination of phospholipid

Phospholipid hydrolysis Phospholipid hydrolysis

Phospholipid analysisPhospholipid analysis

Cholesterol analysisCholesterol analysis

Classes of liposomes:Classes of liposomes:Conventional Long circulating

Immuno Cationic

ModesModes of liposome/cell of liposome/cell interaction:interaction:

Adsorption Endocytosis

Fusion Lipid transfer

Uses of liposomes:Uses of liposomes:Chelation therapy for treatment of heavy metal

poisoning

Enzyme replacement

Diagnostic imaging of tumors

Study of membranes

Cosmetics

Drug Delivery

WhyWhy use liposomes in use liposomes in drug delivery?drug delivery?

Inactive: Unmodified liposomes gather in specific tissue

Active: alter liposome surface with ligand (antibodies, enzymes, protein A, sugars)

Directly to site

Physical: temperature or pH sensitive liposomes

Drug targeting

ProtectionDecrease harmful side effects

Pharmokinetics - efficacy and toxicityChanges the absorbance and biodistribution

Change where drug accumulates in the body

Protects drug

Deliver drug in desired formMultidrug resistance

Why use liposomes in Why use liposomes in drug delivery?drug delivery?

ReleaseAffect the time in which the drug is released

Prolong time -increase duration of action and decrease administration

Dependent on drug and liposome properties Liposome composition, pH and osmotic gradient, and

environment

Why use liposomes in Why use liposomes in drug delivery?drug delivery?

Liposomes help improve:Liposomes help improve:Therapeutic indexRapid metabolismUnfavorable pharmokineticsLow solubilityLack of stabilityIrritation

Custom design:Custom design:

Lipid contentSize

Surface charge

Method of preparation

CurrentCurrent liposomal drug liposomal drug preparations:preparations:

Type of Agents ExamplesAnticancer Drugs

Anti bacterial

AntiviralDNA materialEnzymes

RadionuclideFungicidesVaccines Malaria merozoite, Malaria sporozoite

Hepatitis B antigen, Rabies virus glycoprotein

Amphotericin BIn-111*, Tc-99m

Hexosaminidase A Glucocerebrosidase, Peroxidase

Duanorubicin, Doxorubicin, Epirubicin Methotrexate, Cisplatin*, CytarabinTriclosan, Clindamycin hydrochloride, Ampicillin, peperacillin, rifamicinAZTcDNA - CFTR

No decrease in effectiveness of drug against fungi

LiposomalLiposomal formulation of AmB:formulation of AmB:

Decrease in toxicity

Exact Mechanism of liposomes not understood

Cholesterol - only few %moles

Phospholipid: AmB ratio

DiffusionLipid transfer

AmB

Lipid

Problems with liposomal Problems with liposomal preparations of drugs:preparations of drugs:

Cost Fungizone $40.58 Amphotec $2334

Doxil $1200 per treatment, twice the cost of normal protocol of chemotherapy and drugs

Lack long term stability (short shelf life)

Freeze dry and pH adjustment

Low “Pay Load” - poor encapsulation

Physical and chemical instability

Polar drugs and drugs without opposite charge

Modifications

Possibility of new side effects

ProblemsProblems continuedcontinued

Efficacy

Studies with insulin show that liposomes may be an effective way to package proteins and peptides for use

Clinical Trials for several liposomal formulations More studies on the manipulation of liposomes

FutureFuture

JournalsAllen, Theresa M. "Liposomal Drug Formulations: Rationale for Development and What We Can

Expect for the Future." Drugs 56: 747-756, 1998.Allen, Theresa M. "Long-circulating (sterically stabilized) liposomes for targeted drug delivery ."

TiPs 15: 214-219, 1994.Allen, Theresa M. "Opportunities in Drug Delivery." Drugs 54 Suppl. 4: 8-14, 1997Janknegt, Robert. "Liposomal and Lipid Formulations of Amphotericin B." Clinical Pharmacokinetics.

23(4): 279-291, 1992. Kim, Anna et al. "Pharmacodynamics of insulin in polyethylene glycol-coated liposomes."

International Journal of Pharmaceutics. 180: 75-81, 1999.Quilitz, Rod. "Oncology Pharmacotherapy: The Use of Lipid Formulations of Amphotericin B in Cancer

Patients." Cancer Control.5:439-449, 1998. Ranade, Vasant V. "Drug Delivery Systems: Site-Specific Drug Delivery Using Liposomes as Carriers."

Pharmacology. 29: 685-694, 1989. Storm, Gert and Daan J.A. Crommelin. "Liposomes:quo vadi?" PSTT 1: 19-31, 1998. Taylor, KMG and JM Newton. "Liposomes as a vecicle for drug delivery." British Journal of Hospital

Medicine. 51: 55-59, 1994

WebsitesJames, John S. "Doxil Approved for KS." www.immunet. org.imminet/atn.nsf/page/a-235-03.Wasan, Ellen. "Targeted Gene Transfer." Member.tripod.com/~rrishna/lipos1.html"Introduction to Controlled Drug Delivery Systems." www5.bae.ncsu.edu/bae/reearch/blak…

k/otherprojects/drugDeliver_97/http://www. Mssm.edu/medicine/thrombosis/phosphol.html"Doxorubicin." http://tirgan.com/adria.htm"Clinical Pharmacology Online." http://www.cponline.gsm.com/scripts/fullmono/showmono. "Drugstore.com" http://www.drugstore.com/pharmacy/prices/Amphotec."Sequus' Doxil Becomes First Liposome Product Approved In U.S." www.slip.net/~mcdavis/

database/doxor_1"Liposomes." www.collabo.com/liposom0.htmPaustin, Timothy. “Cellular Membranes.”www.bact.wisc.edu/microtextbook/bacterialstructure/Membranegen.htmlwww.cbc.umn.edu/~mwd/cell_www/chapter2/membrane.html#PHOSPHOLIPIDS

BooksJones, Macolm N. and Chapman, David. Micelles, Monolayers and Biomembranes. Wiley-Liss. New York (1995).

Garrett, R. and Grisham C. Biochemistry, 2nd ed. Saunders Colleges Publishing. New York (1999). 264.

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