Liposome

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Liposome

  1. 1. Liposomes 1
  2. 2. OUTLINE• WHAT ARE LIPOSOMES?• BASIC LIPOSOME STRUCTURE.• WHY USE LIPOSOMES IN DRUG DELIVERY?• ADVANTAGES OF LIPOSOMES.• STRUCTURAL COMPONENTS OF  LIPOSOMES.• CLASSIFICATION OF LIPOSOMES.• PREPARATION OF LIPOSOMES. 2
  3. 3. Liposomes (= vesicles) Liposomes are concentric bilayered vesicles in which an aqueous core is entirely enclosed by a membranous lipid bilayer mainly composed of natural or synthetic phospholipids. Liposomes were first produced in England in 1961 by Alec D. Bangham, who was studying phospholipids and blood clotting. The size of a liposome ranges from some 20 nm up to several micrometers. 3
  4. 4.  The lipid moecules are ususally phospholipids- amphipathic moieties with a hydrophilic head group and two hydrophobic tails. On addition of excess water, such lipidic moieties spontaneously originate to give the most thermodynamically stable conformation. In which polor head groups face outwords into the aqueous medium, and the lipidic chains turns inwords to avoid the water phase, giving rise to double layer or bilayer
  5. 5. HydrophobicHydrophiliccavity 5
  6. 6. Basic liposome structure 6
  7. 7. What is a lamella?– A Lamella is a flat plate like structure that appearsduring the formation of liposomes. The phospholipidsbilayer first exists as a lamella before getting converted into spheres.– Several lamella of phospholipids bilayers are stacked one on top of the other during formation of liposomes  to form a multilamellar structure.  7
  8. 8. Unilamellar vesicles  Multilamellar vesicles        8
  9. 9. Structural Components of Liposomes• THE MAIN COMPONENTS OF LIPOSOMES ARE :-    1.  Phospholipids2.  Cholesterol 9
  10. 10. Phospholipids • Phospholipids are the major structural compone nts of biological membranes such as  the  cell membrane. Phosphoglycerides Two Types Of Two Types Of Phospholipids(Along With Phospholipids(Along With Their Hydrolysis Products) Their Hydrolysis Products) Sphingolipids 10
  11. 11. Phosphatidylcholine• Most common phospholipids used is     phosphatidylcholine (PC).•   Phosphatidylcholine is an amphipathic    molecule in which exists:-    – a hydrophilic polar head group,        phosphocholine.    – a glycerol bridge.    – a pair of hydrophobic acyl hydrocarbon        chains. 11
  12. 12. Generally phospholipids are represented as follows:- follows: 12
  13. 13. Phospholipids Polar Head Groups Three carbon glycerol 13
  14. 14. 14
  15. 15. ROLE OF CHOLESTEROL IN BILAYER FORMATION• Cholesterol by itself does not form bilayer structure.• Cholesterol act as fluidity buffer• After intercalation with phospholipid molecules alter the freedom of motion of carbon molecules in the acyl chain• Restricts the transformations of trans to gauche conformations• Cholesterol incorporation increases the separation between choline head group & eliminates normal electrostatic & hydrogen bonding interactions
  16. 16. Some other commonly used  phospholipids Naturally occurring phospholipids:      –  PC : Phosphatidylcholine      –  PE : Phosphatidylethanolamine      –  PS : Phosphatidylserine Synthetic phospholipids:      –  DOPC : Dioleoylphosphatidylcholine      –  DSPC : Distearoylphosphatidylcholine 16
  17. 17. Why Use Liposomes in Drug  Delivery? Drug Targetingnactive:  Unmodified liposomes gather in specific tissue  reticuloendothelial system Active:  alter liposome surface with ligand  (antibodies,  enzymes, protein A, sugars) Physical: temperature or pH sensitive liposomes  Directly to site 17
  18. 18. Why Use Liposomes in  Drug Delivery? Pharmacokinetics - efficacy and toxicityChanges the absorbance and biodistribution  Deliver drug in desired form Multidrug resistance ProtectionDecrease harmful side effects Change where drug accumulates in the body Protects drug 18
  19. 19. Why Use Liposomes in  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 19
  20. 20. ADVANTAGES OF LIPOSOMES• Provides selective passive targeting to tumor       tissues.•    Increased efficacy and therapeutic index.•    Increased stability of encapsulated drug.•    Reduction in toxicity of the encapsulated agent.•    Site avoidance effect (avoids non-target tissues).•    Improved pharmacokinetic effects (reduced        elimination increased circulation life times).•   Flexibility to couple with site specific ligands to      achieve active targetting.   20
  21. 21. DISADVANTAGES PHYSICAL/ CHEMICAL STABILITY VERY HIGH PRODUTION COST DRUG LEAKEGE/ ENTRAPMENT/ DRUG FUSIONSTERILIZATION  SHORT  BIOLOGICAL ACTIVITY / t ½ OXIDATION OF BILAYER …LIPIDS AND LOW  SOLUBILITY RATE OF RELEASE  and ALTERED BIODISTRIBUTION  LOW THEARAPEUTIC INDEX and DOSE EFFECTIVENESS OVERCOMING RESISTANCE EXTENCIVE CLINICAL  AND LABORATORY  RESEARCH TO      ACERTAIN LONG CERCULATING LIPOSOMESREPEATED IV ADMINISTRATION PROBLEMES
  22. 22. CLASSIFICATION OF LIPOSOMES• Liposomes are classified on the basis of      – Structural parameters     – Method of preparation     – Composition and applications 22
  23. 23. 1. Based on structural parameters Based on structural parameters MLV OLV  MVV UV Unilamellar Multilamellar  oligolamellar  Multivesicular  Vesicles (all      Large  vesicles vesicles size ranges)  vesicles (>0.1-1.0 um) (> 1.0 um) (>0.5 um) SUV Small Unilamellar  MUV Medium Unilamellar  Vesicles  Vesicles 20-100nm LUV Large Unilamellar GUV Giant Unilamellar Vesicles Vesicles >1um >100nm 23
  24. 24. 24
  25. 25. 25
  26. 26. 3. Based on composition & application1- Conventional.2- Stealth.(PEG, increase  blood circulation time and  decrease phagocytic  attack).3- Cationic.(lipoplex)4- Targeted.(antibody) 26
  27. 27. PREPARATION OF LIPOSOMES Methods of liposome  preparation Active or remote loading: certain types of compounds with  Passive loading: ionisable groups  Involves loading of  the   and those with both  entrapped  manufacturing procedureagents before or during the   lipid and water solubility can be manufacturing procedure introduced into the liposomes  after the  formation of the intact vesicles27
  28. 28. Methods of liposome preparationPassive loading techniques Active loading techniquesMechanical dispersion Solvent dispersion Detergent removal methods methods methods Detergent(Cholate,Lipid film hydration by  Ethanol injection    Alkyl glycoside,     hand shaking non-hand Ether injection    Triton X-100) removal   shaking and freeze drying Double emulsion            Micro emulsification    from mixed micelles by   vesiclesSonication Stable plurilamellar  DialysisFrench pressure cell Vesicles Column Membrane extrusion Reverse phase                  chromatographyDried reconstituted                 evaporation vesicles Dilution  vesicles Reconstituted sendaiFreeze thawed liposomes    virus enveloped     vesicles 28
  29. 29. BASIC METHOD OF FORMULATION OF LIPOSOMES SOLVANT eg.CHCl3 LIPID e.g.. LICITHIN DISPERSION FORMATIONN Separate liposomes from supernatatant by Encapsulated solute centrifugation, gel filtration/ sonication or FILM OF LIPID dialysis or by addition of OCCURS AT SIDES buffers + drug. OF VESSELS
  30. 30. Method Vesicles Mechanical methodsVortex or hand shaking of phospholipid dispersions MLVExtrusion through polycarbonate filters at low or medium  OLV, LUVpressureExtrusion through a French press cell “Microfluidizer”  Mainly SUVtechniqueHigh-pressure homogenization Mainly SUVUltrasonic irritation SUV of minimal sizeBubbling of gas BSV Methods based on replacement of organic solvent(s) by aqueous mediaRemoval of organic solvent(s) MLV, OLV, SUVUse of water-immiscible solvents: ether and petroleum MLV, OLV, LUVEthanol injection method LUVEther infusion (solvent vaporization) LUV, OLV, MLVReverse-phase evaporation Methods based on detergent removalGel exclusion chromatography SUV“Slow” dialysis LUV, OLV, MLVFast dilution LUV, OLVOther related techniques MLV, OLV, LUV, SUV
  31. 31. PASSIVE LOADING TECHNIQUES • Mechanical Dispersion method • Solvent Dispersion method • Detergent Solubilisation method
  32. 32. MECHANICAL DISPERSION METHODS
  33. 33. LIPID HYDRATION METHODThe mechanical energy required for swelling of lipids and dispersionof casted lipid film is imparted by manual agitation (hand shakingtechnique)The % encapsulation efficiency as high as 30% is achieved due toloss of water soluble component during swelling and entrapped only10-15%. On other hand lipid soluble drug encapsulated 100%
  34. 34. 1. Hand‐shaken multilamellar vesicles2. Non‐shaking vesicles3. Pro‐liposomes4. Freeze drying   :–AFTER THESE METHODS, OTHER PROCESSING METHODS ARE USED TO MODIFY THESE TYPE OF VESICLES THAT ARE PRODUCED SUCH AS:• Micro emulsification liposomes (MEL)• Sonicated unilamellar vesicles (SUVs)• French Pressure Cell Liposomes• Membrane extrusion liposomes• Dried‐reconstituted vesicles (DRVs)• Freeze thaw Sonication (FTS)• pH induced vesiculation• Calcium induced fusion    :–These methods are known as “the mechanical treatment of MLVs” or  “Processing of  lipids by physical means 34
  35. 35. Hand shaken multilamellar vesicles• Simplest and most widely used method of physical dispersion• Basic method involves – Dissolution of the lipid mixture and charge components in         chloroform:methanol solvent – Evaporation of the solvent in a rotary evaporator or by hand      shaking to form a film shaking to form a – Further drying of the film by attaching the flask to the       manifold of the lyophilizer. – Casted film is then dispersed in an aqueous medium. – Upon hydration, lipid swell and peel off the wall of the flask     and vesiculate forming multilamellar vesicles (MLVs)  35
  36. 36. PROCESS IN MORE DETAIL – STEP 1:• Lipid mixture of different phospholipids and charge components in  chloroform:methanol solvent mixture (2:1 v/v) is prepared first and  then introduced into a round bottom flask with a ground glass neck.• This flask is then attached to a rotary evaporator and rotated at 60  rpm.• The organic solvents are evaporated at about 30 degrees Celsius or  above the transition temperature of the lipids used.• The rotation is continued for 15 mins after the dry residue first  appears.• The evaporator is isolated from the vacuum source by closing the  tap.• The nitrogen is introduced into the evaporator and the pressure at  the cylinder head is gradually raised till there is no difference  between inside and outside the flask between inside and outside the  flask.• The flask is then removed from the evaporator and fixed on to the  manifold of lyophilizer to remove residual solvents. 36
  37. 37. STEP 2‐ Hydration of lipid layer:• After releasing the vacuum and removal from the lyophilizer,  the flask is flushed with nitrogen.• 5ml of saline phosphate buffer (containing solute to be  entrapped) is added.• The flask is attached to the evaporator again (flushed with N2)  and rotated at room temperature and pressure at the same  speed or below 60 rpm.• The flask is left rotating for 30 minutes or until all lipid has  been removed from the wall of the flask and has given  homogenous milky‐white suspension free of visible particles.• The suspension is allowed to stand for a further 2 hours at  room temperature or at a temperature above the transition  temperature of the lipid in order to complete the swelling  process to give MLVs. 37
  38. 38. Hand shaken method in general   38
  39. 39. Non‐shaking vesicles• The procedure differs from hand shaken method in that it uses a stream  of nitrogen to provide agitation rather than the rotationary movements.• Solution of lipid in chloroform:methanol mixture is spread over the flat  bottom conical flask.• The solution is evaporated at room temperature by flow of nitrogen  through the flask without disturbing the solution.• After drying, water saturated nitrogen is passed through the flask until  the opacity of the dried lipid film disappears (15-20mins).• After hydration, lipid is swelled by addition of bulk fluid. The flask is  inclined to one side, 10‐20 ml of 0.2M sucrose in distilled water  (degassed) is introduced down the side of the flask, and the flask is  slowly returned to upright orientation.• The fluid is allowed to run gently over the lipid layer on the bottom of the  flask.• The flask is flushed with nitrogen sealed and allowed to stand for 2 hrs  at 37 degrees Celsius. Take care not to disturb the flask in any way.• After swelling, the vesicles are harvested by swirling the contents of the  flask gently , to yield a milky‐suspension. 39
  40. 40. PROLIPOSOMES To increase the surface area of dried lipid film and to facilitate continuous hydration and lipid is dried over the finally divided particulate support i.e.- NaCl, Sorbitol, or other polysaccharides. These dried lipid coated particulates are called as proliposomes Proliposomes form dispersion of MLVs on addition of water, where support is rapidly dissolved and lipid film hydrate to form MLVs Methods overcome the stability problem and entrapment efficiency doesn’t matter when formation of stable liposome.
  41. 41. Freeze drying•   Another method of dispersing the lipid in a finely divided  form, prior to addition of aqueous media, is to freeze dry  the lipid dissolved in a suitable organic solvent.• The solvent choice depends on the freeze point which  needs to above the temperature of the condenser  lyophilizers. Tertiary butanol is considered to be most  ideal solvent.• After obtaining the dry lipid which is an expanded foam  like structure, water or saline can be added with rapid  mixing above the phase transition temperature to give  MLVs. 41
  42. 42. SONICATION METHODPROBE  SONICATOR: is employed fordispersions, which require high energy in asmall volume (e.g., high concentration oflipids, or a viscous aqueous phase)Disadvantage- lipid degradation due to highenergy and sonication tips release titaniumparticles into liposome dispersionBATH  SONICATOR:  The bath is moresuitable for large volumes of diluted lipids.Method: Placing a test tube containing thedispersion in a bath sonicator and sonicatingfor 5-10min(1,00,000g) which yield aslightly hazy transparent solution. Usingcentrifugation to yield a clear SUVdispersion
  43. 43. FRENCH PRESSURE CELL LIPOSOMESThis techniques yields rather “uni oroligo lamellar liposomes” of intermediatesize of 30-80 nm in diameter depending onthe applied pressure.Dispersion of MLVs can be converted toSUVs by passage through a small orificeunder high pressure.MLV dispersion are placed in the Frenchpressure cell and extruded at about20,000psi at 450C By multiple extrusioni.e.., 4.5 passed about 95% of MLVs getconverted into SUVs which can bedetermined by size exclusionchromatography.
  44. 44. MICRO EMULSIFICATION LIPOSOMES(MEL) “Micro Fluidizer” is used to prepare small MLVs from Concentrated lipid dispersion The lipids can introduced into fluidizers, either as a dispersion of large MLVs or as a slurry of unhydrated lipids in organic medium. Microfluidizer pumps the fluid at very high pressure(10,000psi, 600-700 bar) through a 5um orifice. Then it is forced along defined micro channels, which direct two streams of fluid to collide together at right angles at a very high velocity, thereby affecting an efficient transfercanenergy. The fluid collected of be recycled through the pump and interaction chamber until vesicles of the spherical dimension are obtained. After a single pass, the size of vesicles is reduced to a size 0.1 and 0.2um in diameter.
  45. 45. VESICLES PREPARED BY EXTRUSION TECHNIQUES (VETs)It is used to process LUVs as well as MLVs.Liposomes prepared by this tech. are called as membrane filter extrusion liposomesThe 30% capture volume can be obtained using high lipid conc. The trapped volume in this process is 1-2 litre /mole of lipids It is due to their ease of production, readily selectable vesicle diameter, batch to batch reproducibility & freedom from solvent or surfactant contamination is possible
  46. 46. FREEZE THAW SONICATION METHOD (FTS)The method is based onfreezing of a unilamellardispersion & then thawing atroom temp for 15 min.Thus the process ruptures &refuses SUVs during which thesolute equilibrates betweeninside & outside & liposomesthemselves fuse & increase insize.Entrapment volume can beupto 30% of the total vol. ofdispersion. Sucrose, divalentmetal ions & high ionicstrength salt solutions can notbe entrapped efficiently
  47. 47. SOLVENT DISPERSION METHODSNote:- Organic solvent miscible with aqueous phase
  48. 48. SOLVENT DISPERSION METHODS FOR PASSIVE LOADING
  49. 49. Ethanol injection:-• An ethanol solution of lipids is injected rapidly through a fine needle into an excess of saline or other aq. medium• This method has low risk of degradation of sensitive lipids• The vesicles of 100 nm size may be obtained by varying the conc. Of lipid in ethanol or by changing the rate of injection of ethanol solution in preheated aqu. solution.• Limitation-solubility of lipids in ethanol & vol. of ethanol that can be introduced into medium(7.5%v/v max)• Difficulty to remove residual ethanol from phospholipid membrane Ether injection:-• Involves mixing of organic phase into aqu. Phase at the temp. of vaporizing the organic solvent• It has low encapsulation efficiency
  50. 50. REVERSE PHASE EVAPORATION METHOD
  51. 51. DETERGENT SOLUBILISATIOIN METHODSNote:- Liposome size and shape depend on chemical nature of detergent, concentration and other lipid involved
  52. 52. Detergent depletion method• The phospholipids are brought into intimate contact  with the aqueous phase via detergents which  associate with phospholipid molecules• The structures formed are called as micelles• The conc. of detergent at which micelles are formed is  called as CMC• The detergent methods are not very efficient in %  entrapment values• The methods employed for removal of detergent  include dialysis, column chromatography & use of  biobeads
  53. 53. Dialysis MethodDetergent commonly use for this purpose exhibit resonably high CMC (10 to 20 mM) so that their removal is facilitated A commercial version of the dialysis system is available under the tradename LIPOREPColumn ChromatographyPhospholipid inthe form of either sonicated vesicle or as a dry film, at a molar ratio of 2:1 with deoxycholate form unilamellar vesicles of 100nm on removal of deoxycholate by column chromatography  54
  54. 54. ACTIVE LOADING TECHNIQUES AFTER DRYING IN PROCESS• Weak amphipathic bases accumulate in the aqueous phase of lipid vesicles in response to a difference in pH between the inside and outside of the liposomes (pHin & pHout) FILM/CAKE OF LIPID IS FROM• Two steps process generates this pH imbalance and active (remote) loading.• Vesicles are prapared in low pH solution, thus generating low pH within the liposomal interiors, followed by addition of the base to STACKS OF LIPID extraliposomal medium. BILAYER FORM• Basic compounds, carrying amino groups are relatively lipophipic at high pH and hydrophilic at low pH.• In two chambered aqueous system separated by membrane liposomes, accumulation occurs at the low pH side, under SWELLING IN FLUID dynamic equilibrium conditions.• Thus the unprotonated form of basic drug can diffuse through the bilayer SHEET IS SELF CLOSE• The exchange of external medium by gel chromatography with neutral solution LOADING OF DRUG• Weak base doxorubicine, adriamycin and vincristine which co- ON pH- GRADIENT TECHNIQUE exist in aqueous solutions in neutral and charged forms have been sucessfully loaded into preformed liposomes via the pH gradient FORMATION OF BILAYER method. (LIPOSOMES) IF DRUG
  55. 55. 56
  56. 56. 57
  57. 57. 58
  58. 58.  Reverse phase evaporation vesicles partial  bilayer In rotar y evaporator close to each other At this stage- the monolayers come close to each other 59
  59. 59. Ethanol/Ether injection method 60
  60. 60. DRYING• An important step involved in the preparation of  liposomes is the drying of the lipid.• Large volume of organic solution of lipids is most    easily dried in a rotary evaporator fitted with a dried a  rotary evaporator fitted with cooling coil and a  thermostatically controlled water bath.• Rapid evaporation of solvent is carried out by gentle  warming (20‐40 degrees) under reduced pressure (400  700 mm Hg)(400‐700 mm Hg)• Rapid rotation of the solvent containing flask increases  the surface area for evaporation.  61
  61. 61. • In  cases  where  sufficient  vacuum  is  not  attainable  or  if  the  concentration  of  lipids  is  particularly  high,  it  may  be  difficult to remove the last traces of chloroform from the  lipid film.    Therefore, it is recommended as a matter of routine that  after  rotary  evaporation,  some  further  means  is  employed to bring the residue to complete dryness. Attachment of the flask to the manifold of lyophilizer, and  overnight exposure to high vacuum is a good method. 62
  62. 62. I] Physical Dispersion or Mechanical Dispersion Methods• Aqueous volume enclosed using this method is usually  5‐10%, which is very small proportion of the total volume  used for swelling.• Therefore large quantity of water‐soluble compounds are  wasted during swelling.• On the other hand, lipid soluble compounds can be  encapsulated to 100% efficacy, provided they are not  present in quantities that are greater than the structural  component of the membrane. 63
  63. 63. How LUVs Are Generated From The Suspension?• The suspension is centrifuged at 12,000g for 10  min. in a bench centrifuge at room temperature.• The layer of multilamellar vesicles floating on the  surface is removed. To the remaining fluid an  equal volume of iso‐osmolar glucose is added  and centrifuged again at 12,000g. Large  unilamellar vesicles form a soft pellet which can  be resuspended in any required medium of  appropriate osmolarity. 64
  64. 64. Pro‐Liposomes• Method devised to increase the surface of dry lipid while  keeping the low aqueous volume.• In this method, the lipids are dried down to a finely divided  particulate support, such as powdered sodium particulate  support, such as powdered chloride, or sorbitol or other  polysaccharide – to give pro‐liposomes.• The lipids are swelled upon adding water to dried lipid coated  powder (pro‐liposomes), where the support rapidly dissolves  to give a suspension of MLVs in aqueous solution.• The size of the carrier influences the size and heterogeneity  of the liposomes. 65
  65. 65. • This method overcomes the problems encountered when  storing liposomes themselves in either liquid, dry or frozen  form, and is ideally suited for preparations where the material  to be entrapped incorporates into lipid membrane.• In cases where 100% entrapment of aqueous component is  not essential this method is also of value .• For preparing pro‐liposome a special equipment i.e. Buchi  rotary evaporator ‘R’ with water cooled condenser coil and a  stainless steel covered thermocouple connected to a digital  thermometer is required.• The end of the glass solvent inlet tube is modified to ato fine  point, so that the solvent is introduced into the flask as a fine  spray. 66
  66. 66. BUCHI  Rotary Evaporator R type 67
  67. 67. Method Of Preparation of Proliposomes• The lipid solution in chloroform (60mg/ml) is prepared and sorbitol powder is  introduced into 100ml flask.• The flask is then fitted into the evaporator and rotated slowly so that the  powder tumbles gently off the walls to ensure good mixing and the solvent is  evaporated.• The flask is lowered into a water bath at 50‐55 degrees Celsius when a  good vacuum is developed.• An aliquot of 5ml of lipid solution is introduced into the flask via the solvent  inlet tube.• The solvent is absorbed completely by the powder and the temperature of  the bed is monitored.• As evaporation proceeds the temperature will decrease.• A second aliquot is introduced slowly when the temperature begins to rise  again.• The temperature is allowed to rise to 30 degrees Celsius the vacuum is  released and the drying process is completed by connecting the flask  containing the powder to lyophilizer, and leaving it evacuated overnight at  room temperature.• The powder is transferred into a 10ml glass vial containing 600mg solid  each (100mg lipid and 500mg sorbitol support) flushed with nitrogen, and  68 sealed well and stored.
  68. 68. Processing of the lipids hydratedby physical means, or the mechanical treatment of MLVs• Micro emulsification liposomes (MEL)• Sonicated unilamellar vesicles (SUVs)• French Pressure Cell Liposomes• Membrane extrusion liposomes• Dried‐reconstituted vesicles (DRVs)• Freeze thaw Sonication (FTS)• pH induced vesiculation• Calcium induced fusion 69
  69. 69. Micro emulsification liposomes (MEL)   70
  70. 70. Sonicated unilamellar vesicles (SUVs)     71
  71. 71. French Pressure Cell Liposomes     72
  72. 72. Membrane Extrusion Liposomes 73
  73. 73. Liposomes Help ImproveTherapeutic indexRapid metabolismUnfavorable pharmokinetics Low solubility Lack of stability Irritation 74
  74. 74. Current liposomal drug  preparations Type of Agents ExamplesAnticancer Drugs Duanorubicin, Doxorubicin*, Epirubicin Methotrexate, Cisplatin*, CytarabinAnti bacterial Triclosan, Clindamycin hydrochloride, Ampicillin, peperacillin, rifamicinAntiviralDNA material AZTEnzymes cDNA - CFTR*Radionuclide Hexosaminidase AFungicides Glucocerebrosidase, PeroxidaseVaccines In-111*, Tc-99m Amphotericin B* Malaria merozoite, Malaria sporozoite Hepatitis B antigen, Rabies virus glycoprotein 75 *Currently in Clinical Trials or Approved for Clinical Use
  75. 75. CFTRGene Therapy Deliver cDNA of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) to epithelial tissue of respiratory system Cationic liposome Fuse to cell membrane and incorporate cDNA into cell Clinical trials - no significant change in symptoms Now trying adeno associated virus 76
  76. 76. Doxil Chemotherapy drug doxorubin Anemia, damage to veins and tissue at injection, decrease platelet and WBC count, toxic toTreats Kaposi’s sarcoma lesions or cancer tumorsModifications of liposome “stealth” keeps doxorubin in blood for 50 hours instead of 20 minutes concentrates at KS lesions and tumors *Just approved by FDA* 77
  77. 77. Amphotericin BSystemic fungal infections in immune compromised patientsAmB - kills ergosterol-containing fungal cells, also kills cholesterol-containing human cellsSide effects: nephrotoxicity, chills, and feversFungizone - AmB with deoxycholate 78
  78. 78. Liposomal Formulation of AmB Phospholipid:AmB ratio AmB Cholesterol - only few %moles LipidExact Mechanism of liposomes not understood Diffusion Lipid transferDecrease in toxicity 79No decrease in effectiveness of drug against fungi
  79. 79. Problems with Liposomal Preparations of Drugs $$$$   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 Physical and chemical instability  Freeze dry and pH adjustment Low “Pay Load” - poor encapsulation Low “Pay Load” - poor encapsula Polar drugs and drugs without opposite charge Modifications Possibility of new side effects Doxil “hand and foot syndrome” 80
  80. 80. FutureStudies with insulin show that liposomes may be an effective way to package proteins and peptides for useClinical Trials for several liposomal formulationsMore studies on the manipulation of liposomes 81
  81. 81. THANK YOU 82

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