脂质体教学讲解课件.ppt

1、Burgess,June 28,20011REGULATORY SCIENCE OF LIPOSOME DRUG PRODUCTSDiane J.Burgess,Ph.D.Professor of Pharmaceutics University of Connecticut Office of Testing and Research CDER,FDABurgess,June 28,20012Outline What are liposomes?What are they used for?What drugs?Why liposomes?Liposome formulation Lipos

2、ome characterization Safety concerns Performance concerns In vitro release testing stabilityBurgess,June 28,20013Outline Continued Purpose of in vitro release tests?Design of in vitro release test Accelerated/stress tests Method variables affecting release Methods under development In vivo factors a

3、ffecting release In vivo data and models?IVIVC?Research proposal LIPOSOMESLiposomes are colloidal,lipid vesicles consisting of one or more self-assembled lipid bilayers enclosing a similar number of aqueous compartments.Lipids,such as lecithin(diacylphosphatidylcholine),are amphiphilic molecules.Due

4、 to the bulky nonpolar part of the molecule they do not pack into spherical micelles in aqueous phase but rather self-assemble into bilayers which tend to self-close at low concentrations into spherical structures.LIPOSOMES Contd.Liposomes can be subcategorized into:Small unilamellar vesicles(SUV),2

5、5 to 100 nm in size that consist of a single lipid bilayer Large unilamellar vesicles(LUV),100 to 400 nm in size that consist of a single lipid bilayer Multilamellar vesicles(MLV),200 nm to several microns,that consist of two or more concentric bilayers Vesicles above 1 m are known as giant vesicles

6、.Burgess,June 28,20016LiposomesLocalized and rate controlled delivery:Improved therapeutic response Achieve appropriate tissue or blood levelsReduced adverse reactions Less drug administered Targeted drug releaseLower dosing frequency Improved patient compliance Simpler dosing regimens Lower cost pe

7、r doseUtilization of otherwise un-useable compounds Poorly soluble drugsBurgess,June 28,20017Drug Candidate SelectionKnown therapeutics with clear toxicity and pharmacokinetic profiles Potent compounds Not“Narrow Therapeutic Index”drugs Problems associated with the current dosage forms First pass ef

8、fects or poor absorption Gastric irritation Rapid clearanceMedical need for improved deliveryDrugs compatible with manufacturing conditionsBurgess,June 28,20018.APPROVED LIPOSOME PRODUCTS:Doxil Daunorubicin1995 Daunoxome Daunorubicin1996 Ambisome Amphotericin B1997 DepocytCytarabine1999APPROVED LIPI

9、D COMPLEX PRODUCTS:AmbelcetAmphotericin B1995 AmphotecAmphotericin B1997Burgess,June 28,20019SELECTION OF DELIVERY SYSTEMLiposomes targeted delivery.They can deliver agents directly into cells.Routes:i.v.,s.c.,i.m.,topical,pulmonaryMicrospheres-can provide continuous drug delivery over periods of mo

10、nths to years.Systemic and localized.i.m.,s.c.,oral,pulmonaryEmulsions-can be used to make highly water insoluble compounds bioavailable.i.v.,oral,topicalBurgess,June 28,200110LIPOSOME FORMULATIONLIPOSOMESLiposomal composition determines the properties(e.g.surface charge,rigidity and steric interact

11、ions)and the in vitro and in vivo performance.Both water soluble and water insoluble drugs may be encapsulatedProcessing methods affect particle size,percentage drug entrapment,stability and release ratesBurgess,June 28,200111LIPOSOME FORMULATION Processing methods:Extrusion,ultrasonication and micr

12、ofluidization for hydrophobic drugs and Reversed phase and freeze-thaw for hydrophilic drugs.Burgess,June 28,200112Liposomes:Factors Affecting Performance Release Rate and StabilityPhase transition temperature(Tg)effects membrane changes from ordered solid to disordered fluid and is dependent on the

13、 length and degree of saturation of the hydrocarbon chains.Cholesterol-disordering of the ordered phase and ordering of the disordered phase eventually leading to an elimination of the phase transition.High stability and low leakageSurface charge and steric interaction:RES targeting/avoiding RES upt

14、akeBurgess,June 28,200113Types of Liposomes Conventional Liposomes-Prepared form natural neutral and anionic lipids and have nonspecific interactions with their environment-Relatively unstable,have low carrying capacities,and tend to be“leaky”to entrapped drug substances-May literally fall apart on

15、contact with plasma,particularly those of high fluidity,-Choleterol is often added to increase plasma stabilityBurgess,June 28,200114Types of Liposomes Non-conventional Liposomes-Small sized(100 nm),surface modified to overcome some of the short comings of conventional liposomes-Modified to reduce n

16、egative charge,decrease fluidity and cause steric hinderance to phagocytosis-Properties altered(e.g.by incorporation of cholesterol)-Polymerized liposomes more stable and less“leaky”-Polyetheylene glycol,“pegylated”liposomes,avoid uptake by the mononuclear phagocytic cellsBurgess,June 28,200115TYPES

17、 OF LIPOSOMES Target specific ligands,such as antibodies,immunoglobulins,lectins and oligosaccharides attached to the surface to actively target to specific sites in the body Targeting via particle size Liposomes prepared with cationic and fusogenic lipids are currently being utilized in gene therap

18、y to deliver DNA into target cellsBurgess,June 28,200116TYPES OF LIPOSOMES Highly reactive liposomes-readily undergo phase transition in particular situation sensitive to pH,ions,heat and light For example,pH-sensitive liposomes can undergo phase transition in acidic conditions resulting in increase

19、d membrane fluidity and loss of encapsulated materialsBurgess,June 28,200117CRITICAL FACTORS IN LIPOSOME PREPARATIONJParticle size Method of manufactureLipid types Phase transition temperaturePolymerizationInterfacial chargeSteric stabilizationSterilizationBurgess,June 28,200118Liposomes:Factors Aff

20、ecting Performance Liposome preparations can be stored:frozen,in liquid form and as a freeze dried powder.Reconstitution of liposomes may affect particle size and size distribution.Burgess,June 28,200119SAFETY CONCERNS:LIPOSOME FORMULATION Lipid toxicity(RBC lysis)Type and concentration%Lyso-lipids

21、Presence of protein and lipoprotein for natural lipids Residual solvent Overload of RES Particle size (tail above 1 um)-Blockage of capillaries Size affects RES uptake and tissue targeting Stability:shelf-live and in vivo Dose dumping(via protein binding)SterilityBurgess,June 28,200120LIPOSOME CHARA

22、CTERIZATION StabilIty Drug Lipids Liposome Phase transition temperature Percent drug loading Percent free drug Drug release rate/stability Particle size Morphology(lamellarity)SterilityBurgess,June 28,200121STERILITYTerminal sterilization?Aseptic processingMust consider both internal and external st

23、erility Burgess,June 28,200122STABILITY Active Inactives(especially the lipids)Liposome as a whole need Any change in particle size can affect targeting,RES uptake,safety and efficacy.In vivo stability of whole liposome is particularly important for targeted liposomes,since they should remain stable

24、 in the plasma without loss of contents until uptake at the target site.Burgess,June 28,200123LIPOSOME DESTABILIZATIONProtein bindingMembrane fusionBurgess,June 28,200124Drug Release from LiposomesRelease profiles are application dependent.Targeted liposomes should remain intact until delivery at si

25、te Other(short term CR and solubilization)release during appropriate time scale.Release controlled by Fluidity/stability(lipids/co-lipids)Condition sensitivity of lipids Size MLV or a SUV Physicochemical properties of drug Drug/lipid interactionBurgess,June 28,200125In Vitro Drug Release Apparatus?M

26、edia?Sampling methods?Testing intervals?Total percent release?No standard method at presentBurgess,June 28,200126Liposome Performance In Vitro Release and Stability Separation of liposomes from dissolution media complicates testing Current USP methods designed for oral and transdermal routes In vitr

27、o tests need to take into account the expected in vivo performance of liposomes Burgess,June 28,200127Liposome Performance In Vitro Release and Stability Release test for a targeted liposome would need to show 1)liposome is stable until uptake at the site 2)liposome releases drug at the site(based o

28、n the mechanism of release in vivo).Release test for an immediate release liposome would need to show Drug is released immediately in conditions mimicking human plasma.Burgess,June 28,200128Current Methods of In Vitro Testing of Liposome Systems Membrane Diffusion Technique Sample and Separate Techn

29、ique In Situ Technique Continuous Flow TechniqueBurgess,June 28,200129Development of In Vitro Release and Stability Methods for Liposomes Purpose:methods to be used in setting regulatory specifications for these products for quality control(QC)purposes to differentiate between“good”and“bad”batches.T

30、ests design will vary depending on the intended in vivo performance of liposomes Burgess,June 28,200130Purpose of In Vitro Release Test?Quality control and safety evaluation Batch to batch Manufacturing process changes Substantiation of label claims Evaluation of potential dose dumping Assessment of

31、 in vivo stability“Real time”vs accelerated/stress test In vitro-in vivo correlationBurgess,June 28,200131Design of In Vitro Release Method Select media and apparatus to achieve reproducible results Attempt to overcome limitations of existing methods Miniaturize methods Prepare formulation variants

32、with different in vivo performance Test formulation variants in vitro and in vivo Modify in vitro test if not discriminatory Determine in vivo factors that effect release Modify in vitro methods to obtain IVIV relationshipBurgess,June 28,200132Accelerated In Vitro Release Methods These tests should

33、be predictive of“real time”in vitro tests Drug release mechanism should not be altered Accelerated test should not simply dissolve the liposomeBurgess,June 28,200133Media and Methods that can affect Release Solvents pH Temperature Agitation Enzymes Cell culture Sink conditions Volume Sampling interv

34、alBurgess,June 28,200134In Vivo Factors Affecting Drug ReleaseBurgess,June 28,200135In Vivo Factors Delivery System Independent(Type I)Delivery System Dependent(Type II)Barriers to drug diffusion:fluid viscosity,tissue barriers(e.g.connective tissue)Drug partitioning at the site Available volume at

35、the site Motion at Site Enzymatic degradation of delivery system Protein adsorption Phagocytosis Inflammatory responseBurgess,June 28,200136In Vivo Data Systemic delivery,then plasma levels may be suitable Localized delivery,plasma levels will be low and unrepresentative.Requires tissue levels Use a

36、nimal models in method development Use BiomarkersBurgess,June 28,200137In Vivo Data Use animal model to help design in vitro test Establish relationship between in vitro data and animal in vivo data Establish a relationship between animal in vivo data and human PK,biomarkers,PD responseDevelop relationship between in vitro data Human data Burgess,June 28,200138