sln colloidal handout-portugal_2007
TRANSCRIPT
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A.J. Almeida, 2007
SOLID LIPID NANOPARTICLES AS COLLOIDAL
DRUG CARRIER SYSTEMS
Antnio J. Almeida
Research Institute for Medicines and Pharmaceutical Sciences (iMed.UL),Faculdade de Farmcia
Universidade de Lisboa
Portugal
Universidad de Sevilla, Facultad de Farmacia, November 2007
A.J. Almeida, 2007
MICROPARTICLES AND NANOPARTICLES:
THE BASIC CONCEPTS
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The recent t remendous advances in health science technology and inexorable
progress in related scient if ic innovat ions, coupled with changes in populat ion
demographics and the colouring of pol it ical agendas w i th the economics of
d isease, are al l ac ting in concer t to take the pharmaceutical industry into
excit ing and challenging new dimensions.
G. Gregoriadis (2002)
INOVATIVE DRUGS
A.J. Almeida, 2007
Inovative
Drugs
AIDS
Cancer
Ebola
Tuberculosis
Alzheimer
SARS
Economics
Demographic changes
Globalization
New drug delivery systems
Genomics
Biotechnology
Neurosciences
Gene Therapy
BioinformaticsProteomics
Metalolomics
INOVATIVE DRUGS
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PHYSICAL APPROACH
Macroscopic systems activated by physicalprocesses (capsules, granules, etc)
CHEMICAL APPROACH
Chemical modification of drugs (derivativepreparation, salts, pro-drugs, association to
polymers, etc.)
BIOCHEMICAL / BIOTECHNOLOGICAL APPROACH
Naturally occurr ing m acromolecules Cells
Colloidal polymeric or l ipid systems
DIFFERENT APPROACHES
PHARMACEUTICAL
APPROACH
(pharmaceutical formulation)
A.J. Almeida, 2007
in v it ro
in v ivo
C
Time
0 6 12 18
Time (hour)
Drugconcentration
inserum
t
GENERAL CONCEPTS IN MODIFIED DRUG RELEASE
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A.J. Almeida, 2007 Chien (1992). Novel Drug Delivery Systems
GENERAL CONCEPTS IN MODIFIED DRUG RELEASE
Convencional Release
Modified Release
Delayed Release
Prolonged Release
Controlled Release
Drug Targeting
A.J. Almeida, 2007
NEW DRUG DELIVERY SYSTEMS
SKF
Spansule
ALZA
Oros OcusertProgestasert
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Physicochemical problems
Molecular weight and size
Complex struture
Conformational stability
Solubility
Sensitivity (light, temperature, pH)
Crystallization
Molecularar interactions
Adsorp tion
Aggregat ion
Presence of pr otein impuri ties
Biological problems
Biodegradation by digestive enzymes
Short in vivo half-life
Imunogenicity
Complex metabolism
Dificulty in crossi ng mucosal barriers
No access to some compartments
Pharmacokinetic Problems
Analytical Problems
FORMULATION OF NEW DRUG MOLECULES
PROTEIN DRUGS
A.J. Almeida, 2007
9 Investigation of alternative administration routes (nasal, rectal, pulmonary, etc.);
9 Investigation of alternative prolonged or pulsed release drug delivery systems;
9 Investigation of new dosage forms that can provide pr otection against premature
degradation;
9 Invstigation of new site specific drug delivery systems.
PROTEIN DRUGS
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PARTICULATE SYSTEMS
liposomes
microemulsions
nanosuspensions
microspheres
nanoparticles
niosomes
virosomes
cubosomes
Particulate
Systems
iscoms
cochleates
nanocapsules
micelles
transfersomes
A.J. Almeida, 2007
Definition IUPAC
COLLOIDAL
The term refers to a state of subdivision, implying that the moleculesor polymolecular par ticles dispersed in a medium have at leas t in
one direction a dimension roughly between 1 nm and 1 m, or that in
a system discontinuities are found at distances of that order.
COLLOIDAL SYSTEMS
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Microparticles
Solid particles made of polymeric or solid lipid materials
Podem conter o frmaco disperso, encapsulado ou adsorvido superfcie.
Microspheres (1 m) Microcapsules (reservoir systems)
Microparticles (matrix systems)
Nanospheres (
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Biodegradation Mechanisms
Type I
Type II
A B A C
Type III
POLYMERIC MICRO- AND NANOPARTICULATE SYSTEMS
A.J. Almeida, 2007
BIODEGRADABLE IMPLANTS
POLYMERIC MICRO- AND NANOPARTICULATE SYSTEMS
Poly(lactide-co-glycolide) (PLGA)
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9 Drug release by polymer enzymatic degradation.
9 Drug release rate is contr olled by t he rate of the enzymatic reaction.
9 Predominat use in controll ed release from biodegradable polymers often used in
particulate carriers (e.g. albumine, gelatine, polyalkylcianoacrylates).
F F FF
POLYMERIC MICRO- AND NANOPARTICULATE SYSTEMS
A.J. Almeida, 2007
Main Technologic and Therapeutic Uses
Diagnostic systems
Controlled drug delivery systems
Implants Aerosols Solid dosage forms Vaccines Site-specific drug delivery
POLYMERIC MICRO- AND NANOPARTICULATE SYSTEMS
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MAIN MICRO / NANOENCAPSULATION TECHNIQUES
1. Spray coating; Pan-coating, Spray-drying
2. Droplet extrusion
3. Coacervation / Phase separation
4. Emulsion solidif ication
4.1. Solvent evaporation (simple or multiple emulsion)
4.2. Solvent extraction (simple or multiple emulsion)
4.3. Melting and solidification (simple or multiple emulsion)
5. Polymerization methods
5.1. Interfacial polymerization
5.2. Emulsion polymerization
Colloidal Systems
A.J. Almeida, 2007
A
Aqueous phase+
surfactant
Polymer in organic
solvent
Drug suspended in
polymer solution
Drug
Evaporation orExtraction
of organic solvent
Particle isolation by
filtration or centrifugation
simple [A] or multiple [B]
emulsion
w/o
emulsion
Aqueous phase
+ surfactant
Polymer in
organic solventDrug +water +
emulsifying agent
B
Solvent Evaporation/ Extraction
simple emulsion multiple emulsion
MAIN MICRO / NANOENCAPSULATION TECHNIQUES
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Emulsion Polymerization
MAIN MICRO / NANOENCAPSULATION TECHNIQUES
9 A si ngl e liq uid (or d iso lved ) reacti ng m ono mer
9 Drug disolved or suspended
9 Polymerisation started by chemical agent or radiation
9 Polymer MW and particle size dependent on:
9monomer concentration
9Initiator concentration
9Temperature
9 Emulsion stabilizer needed
9 Particles >20 nm
Ex.: PIBCA; polyacrylamide
A.J. Almeida, 2007
CaCl2
Ferreira Almeida and Almeida (2004). J Control Release.
Gelatine +
alginate + drug
Washing
Fluid-bed drying
Droplet Extrusion
MAIN MICRO / NANOENCAPSULATION TECHNIQUES
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CONTROL OF FORMULATIONS
Particle size
9Electron microscopy
9Electric sensing zone
9Laser diffraction
9Photon correlation spetroscopy (PCS)
9Optical sensing zone
Encapsulation efficiency
Drug loading
Surface charge (zeta potential)
Hydrofobicity
9Contact angle9Partition coeficient
9Hydrophobic interaction chromatography
Release profile
Sterilization
Stability
MICRO- AND NANOPARTICULATE SYSTEMS
A.J. Almeida, 2007
General drug release kinetics
Dependent on:
9 Drug location inside the particle
9 Type and amount of polymer
9 Particle size and density
9 Cross-linking
9 Physicochemical properties of drug and polymer
9 Drug MW and concentration
9 Presence of excipients
9 Release medium
DRUG RELEASE
MICRO- AND NANOPARTICULATE SYSTEMS
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PARTICULATE CARRIERS IN THE PHARMACEUTICAL MARKET
LIPOSOMES
9physical and chemical stability problems
9no cheap liposome available
9marketed products but behind expectations
POLYMERIC NANOPARTICLES
9input : 30 years of research
9no output: no products on the market
A.J. Almeida, 2007
SITE-SPECIFIC DRUG DELIVERY
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SITE-SPECIFIC DRUG DELIVERY
The magic bullet concept
The term used to describe a specific c ure
for syphilis, which would attack the
syphilis spirochaete while having no
effect whatsoever on human tissue.
Paul Ehrlich (1854-1915)
A.J. Almeida, 2007
RATIONALE FOR SITE-SPECIFIC DRUG DELIVERY
Target site
Other sites
Carrier
Drug
SpecificityTranslocation across
biological barriersProtection against inactivation
Puisieux and Roblot-Treupel (1989) STP Pharma
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Exclusive delivery to specific c ompartments (and/or di seases)
Access to previously inaccess ib le s ites (e.g . in tracel lu lar in fections)
Protection of drug and body from unwanted deposition, which could lead tounwanted reactions and metabolism, etc.
Controlled rate and modality of delivery to pharmacological receptor
Reduction in the amount of drug employed
RATIONALE FOR SITE-SPECIFIC DRUG DELIVERY
Drug safety Drug efficacy Patient compliance
A.J. Almeida, 2007
ACTIVE TA RGETING
Magnetic fieldsLynphotropic adjuvants
MPS blockingReceptor mediated processes
Immunotherapy
0,1 4
0,01
PASSIVE TARGETING
Uptake by MPS and tropism for the lysosomesmacrophagesKupffer cells
Translocation to the tissues hepatocytesbone marrowsplenocitestumors (?)
Capillar embolismlung targeting (iv) specific regions (ia)
4 7 10 100
Diametre (m)
SITE-SPECIFIC DRUG DELIVERY: DRUG TARGETING
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Utilizao de lipossomas como
transportadores de frmacos
Crommelin et al. (http://www.drugdeliverypartnerships.com/lip.html)
DRUG TARGETING
A.J. Almeida, 2007
Soluble carriers (monoclonal antibodies, dextrans, soluble s ynthetic polym ers)
Particulate carriers (liposomes, microspheres, nanoparticles, etc.)
Target-specific recognition moieties (monoclonal antibodies, carbohydrates)
Antibody-di rected enzyme/prodrug therapy
Virus-directed enzyme/prodrug therapy
MAIN DRUG TARGETING SYSTEMS
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A.J. Almeida, 2007
SOLID LIPID NANOPARTICLES
A.J. Almeida, 2007
LIPID NANOPARTICLES
9 Lipophilic colloidal delivery system (R.H. Mller, Berlin; M.R. Gasco, Turin);
9 Efficient and non-toxic drug carrier specially for li pophilic drug molecules;
9 Composed of physiological / well tolerated excipients e.g. GRAS (= similar to
emulsions and liposomes);
9 Possess solid matrix (= similar to polymeric nanoparticles);
9 Protective properties
9 Controlled release properties
9 Colloidal dimensions and controlled release behaviour enable drug protection
and administration by parenteral and non-parenteral routes.
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A.J. Almeida, 2007
LIPID NANOPARTICLES
9 Versatility
9Parenteral administration (Fundar et al, 2000)
9Brain delivery (Fundar et al, 2000)
9Ocular delivery (Cavalli et al, 2002)
9Rectal delivery (Sznitowska et al, 2001)
9Oral deli very (Garca-Fuentes et al, 2002)
9Topical delivery (Souto et al, 2004)
9 Potential vaccine delivery systems (Almeida et al, 1997)
9 Large scale production possibl e using l ines available in pharmaceutical plants (i.e.
lines for parenteral emulsions (Mlleret al, 2001)
A.J. Almeida, 2007
SLN PREPARATION
High Pressure Homogenization (R.H. Mlleret al.)
-SkyePharma PLC, Berlin, Germany
-PharmaSol GmbH, Berlin, Germany
Microemulsion Technology (M.R. Gasco)
- Vectorpharma/Eurand Spa, Turin, Italy
Solvent Evaporation
Phase Inversion Type
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A
Aqu eous ph ase
at same
temperature
Melted Lipid
Drug solution in
melted lipid
Drug
Emulsion
w/oAqu eou s ph ase
at same
temperature
Melted LipidDrug + Water
+ surfactant
B
Cooling for solidification
and particle hardening
SLN suspension
High-Pressure Homogenisation
Emulsification using Ultra-turrax
single [A] or double [B]
SLN PREPARATION BY HIGH PRESSUE HOMOGENIZATION (HPH)
HOT HOMOGENIZATION PROCESS
A.J. Almeida, 2007
HIGH PRESSURE HOMOGENIZATION (HPH)
E. Souto (2005) PhD Thesis Dept. Pharmaceutics, Biopharmaceutics and Biotechnology, Free University of Berlin
APV Gau lin LAB 40 dis continuo us
(40 g batch)
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SLN PREPARATION BY HPH
HOT HOMOGENIZATION PROCESS
Basic principles:
9equipment can be qualified and validated
9accepted by regulatory authorities in product ion li nes used for parenterals
9existing industri al production l ines for i.v. parenteral emulsions can be used
9all production lines developed for SLN are usable
A.J. Almeida, 2007
SLN PREPARATION BY HPH
APV Gaul in L AB40 c ont inu ous
(0.5-2 Kg batch )APV Gau lin LAB 60
(2-10 Kg batc h)
APV Gau lin 5.5
(150 Kg/h)
Dept. Pharmaceutics, Biopharmaceutics and Biotechnology, Free University of Berlin
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SLN PREPARATION BY HPH
COLD HOMOGENIZATION PROCESS
Melted Lipid
Drug suspension
in melted lipid
Drug
SLN suspension
High-Pressure Homogenisation
Suspension in water using Ultra-turrax
Liquid nitrogen
Micronization
Dept. Pharmaceutics, Biopharmaceutics and Biotechnology, Free University of Berlin
A.J. Almeida, 2007 Mller et al (2000). Eur J Pharm Biopharm
DRUG ENTRAPMENT INTO SLN
A m.p. drug m.p. lipid homogeneous matrix
B m.p. drug < m.p. l ipid lipid-enriched core
C m.p. drug > m.p. l ipid drug-enriched core
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PROBLEMS ASSOCIATED WITH SLN
Formation of perfect crystalline structure during storage ( modification) drugexpulsion
DRUG ENTRAPMENT INTO SLN
Souto (2007)
A.J. Almeida, 2007
9 Physical stability of aqueous solution
9 Gel formation
9 Particle aggregation
9 High water content of dispersions (70 - 95%)
9 Need to remove too much water in tablet / pellet production
9 Dosing problems (e.g. dispersion for soft gelatin capsules, lipid
particl e content max. 30%)
SLN PROCESSING DISADVANTAGES
Souto (2007)
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SLN PROCESSING DISADVANTAGES
SLN Solid Lipid Nanoparticles
9Produced from solid lipids
9Tend to form perfect crys tals drug expulsion
NLC Nanostructured Lipid Carriers
9Produced from blend of solid and liquid lipi ds
9Particles are in solid state at body temperature
9 Inhibit crystallization process by mixing spatially
very different m olecules imperfections in lattice not just mixing solid lipids but: controlled nanostructuring of lipid matrix
9 to accommodate drug
9 to control release
9 to tr igger release
Souto (2007)
A.J. Almeida, 2007
LIPID NANOPARTICLES
Solid lipid
particleOil-loaded
particle
Souto et al (2007) Pharm Tech Europe (in press)
NLC a multiple carrier
SLN: one phase (=solid lipid)
NLC: multiple O/F particle (oil nanodroplets in solid fat nanoparticles)
Advantages:
higher drug load, firm incorporation (e.g. 1% Retinol in SLN, 6% in NLC)
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LN AS A POTENTIAL PROTEIN / ANTIGEN
DELIVERY SYSTEM
A.J. Almeida, 2007
Lipid compositionHard fats (Softisan 142and WitepsolE85)Cetyl alcoholPropyleneglycol palmitostearate (Monosteol)PEG 300 mono-, di-stearate (Superpolystate)
Surfactant
Poloxamer 188Tween 80
PROTEIN NANOENCAPSULATION IN SLN
Lipid +
poloxamer 188
Aqu eous
Lysozyme
melting
Solubilisationusinga rotary evaporator
Micronisation in a
powder-mill
Pre-mix using a high-
speed homogeniser
HPH at room
temperature
Solidification in
liquid nitrogen
Almeida et al (1997). Int J Pharm
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0
100
200
300
400
500
Distilledwater
3%Tween 80
3% NaCholate
3% Poloxamer188
Lysozymeactivity(Units/ml)
Day 1
Day10
Almeida et al (1997). Int J Pharm
STABILITY OF LYSOZYME THROUGHOUT FORMULATION
MW rt 0,5h 1h 2h 3h 4h 5h
Influence of temperature (50)
MW Ct 3 4 5 6 7 8 9 10
Influence of HPH treatment
(N cycles/Pressure)
500 bar/1 cy/ 25C 1000 bar/3 cy/ 50C
A.J. Almeida, 2007
MW Water S/CA W/CA Mono Sup
NANOENCAPSULATION OF LYSOZYME IN LN
Almeida et al (1997). Int J Pharm.
0
1000
2000
3000
4000
5000
Softisan
142/Cetylalcohol
Witepsol
E85/Cetylalcohol
Monosteol Superpolystate
Lysozymeactivity
(Unit/goflipid)
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0
20
40
60
80
100
120
140
Lysozymeactivity(%)
Wit/CASoft/CA
Initial Sup LN Initial Sup LN
MW Ct 3 4 5 6 7 8
97.0 KDa66.2 KDa45.0 KDa
31.0 KDa
21.5 KDa14.4 KDa
NANOENCAPSULATION OF LYSOZYME IN LN
Almeida et al (1997). Int J Pharm
-9.80.75497.0D50 0.580.00
D90 0.930.01
0.30W E85/CA
-11.00.264412.9D50 0.600.00
D90 1.860.02
0.22S 142/CA
Zeta Potential
(mVsd)
Particle Size
PCS
(nm sd)
Particle Size
Laser diffraction
(m sd)Lys
Content
(%)
Lipid matrix
A.J. Almeida, 2007 Videira and Almeida (1998). Proceed III SPLC-CRS Conf
Lipid compositiongliceryl behenate(Compritol888 ATO)gliceryl palmitosterarate (Precirol ATO 5)
cetylAlcohol Surfactant
Tween 80
Speed rate8 000 - 10 000 rpm
Lipid
Aq.p hase
melting
heatingheating
emulsification
high-speed homogenization8 000 -10 000 rpm
solidification
DEVELOPMENT OF OVA-CONTAINING LN
Particle s ize (PCS)
100-400 nm
PI - 0,250
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SLN (nm) OVA SLN (nm)
Lipid (%) dmt=3d dm t=30d dm t=3d dm t=30d
Pr 103 115 128 153
6 Cp 171 168 196 205
W/AC 284 280 314 320
Pr 196 205 214 206
8 Cp 240 253 280 305
W/Ac 252 265 290 312
Pr 230 200 300 nd
10 Cp 373 378 390 ndW/Ac 307 289 400 nd
NANOPARTICLE CHARACTERISATION: PHYSICAL STABILITY
Videira et al. (2002). Proceed V SPLC-CRS Conf
A.J. Almeida, 2007
330
330
330
330
330
330
330
330
330
Lipid
10%
Pr
Cp
W/CA
8%
Pr
Cp
W/CA
Pr
Cp
W/CA
6%
Tween 80
2%
60% 100%
OVA-6SLN 92% EE with 2% surfactant
ENTRAPMENT EFFICIENCY
Videira et al (2002). Proceed V SPLC-CRS Conf
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Protein Integrity
97.0 KDa66.2 KDa
45.0 KDa
31.0 KDa
21.5 KDa14.4 KDa
MW LNsup 1W 2W 3W
0
10
20
30
40
50
60
70
80
0 5 10 15 20 25 30 35
Time (day)
OVAreleased(%)
FORMULATION STUDIES OF OVA OF LYSOZYME IN LN
0
10
20
30
40
0 50 100 150 200 250 300 350 400
time (min)
Conc.O
VA
(%)
Videira et al (2002). Proceed V SPLC-CRS Conf
A.J. Almeida, 2007
ADSORPTION STUDIES
Desorption profiles
OVA/lipid (%) 4 7 10
25C 90 80 82
15C 80 70 79A.E
(%)
0
20
40
60
80
100
0 250 500 750 1000 1250 1500
time(min)
Conc.
OVA(%)
25C
15C
Videira et al (2002). Proceed V SPLC-CRS Conf
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9 LN are potential protein carriers with high protein binding
9 Formulations are stable
9 Partial adsorption of protein may not be precluded
9 Production conditions seem not to damage protein integrity
9 Following a burst effect, a sustained release profile is obtained
9 Loading capacity is suitable to the usual antigen doses
LN AS A POTENTIAL PROTEIN / ANTIGEN DELIVERY SYSTEM
Videira et al (2002). Proceed V SPLC-CRS Conf
A.J. Almeida, 2007
PULMONARY DELIVERY OF SLN
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PULMONARY ADMINISTRATION
The Lung
Unique features that can facilitate systemic delivery via pulmonary administration of drugs:
Large surface rea (75 m2).
Good vascularisation.
Large capacity for solute exchange.
Ultra-thineness of the alveolar epithelium (0.1-0.5 mm). First-pass metabolism is avoided.
A.J. Almeida, 2007
PULMONARY DELIVERY USING PARTICULATE CARRIERS
Advantages
An alternative non-invasive means for both local andsystemic drug delivery using particulate carriers.
Avoids instability in blood stream and uptake by the MPS.
Allows high concentrations of drug in the lungsminimizing side toxic effects.
A potential route for drugtherapy and immunisation.
Poyner et al (1995).J Controlled Release
Free
Liposomal (0,7 m)PLGA Microencapsulated (0,7 m)
Lung
0
20
40
60
80
100
Kidney
0
20
40
60
80
6 24
Time (hour)
Tobr
amycin(%)
Blood
0
2040
60
80
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LYMPHATIC TARGETING AND LUNG CANCER THERAPY
A leading cause of death around the world, accounting for13% of newly diagnosed cancers and 30% of all cancerdeaths.
Very low survival rate (14% in 5 years; mean survival time14 months).
Metastatic disease starts at very early stages, particularly inSCLC, the majority of patients with lung cancer havingmetastatic disease at the time of diagnosis.
Metastasis spread mainly via the lymphatics to the lymphnodes, liver, brain, bones and adrenal glands.
Early diagnosis is crucial to increase survival rate and stagingis critically dependent upon the involvement of the lymph
nodes that drain the region containing the tumour.
Chemotherapy plays an important role alongside surgery andradiation therapy, but lung cancer is usually diagnosed at anincurable stage.
A.J. Almeida, 2007
IMAGING USING PARTICULATE CARRIERS
Lung perfusion imaging is based on the trapping of i.v. injectedradiolabelled albumin microspheres (10-50m) in the capillary bed
of the lung: Technetium [99m Tc] Microspheres Injection (Ph Eur).
I.v. injected radiolabelled PLGA microspheres have also been
proposed for lung imaging (Delgado et al. 2000).
Several particulate carriers have been used forlymphoscintigraphy, such as liposomes, sulfur colloid, PMMA and
PHCA nanoparticles.
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PULMONARY ADMINISTRATION OF LIPID NANOPARTICLES
To investigate LN as a potential drug carrier to the lungs and, through alveolarairways, to the lymphatic system, thus optimising concentration at the tumour site or
at distant metastasis sites.
To evaluate LN in vivo fate after pulmonary absorption upon nebulisation anddelivery to laboratory animals.
Lipid Nanoparticles
Administrationhave been studied by parenteraland non-parenteral routes.
A.J. Almeida, 2007
NANOPARTICLE PRODUCTION
Lipid composition
gliceryl behenate(Compritol888 ATO)gliceryl palmitosterarate (Precirol ATO 5)cetylAlcohol
SurfactantTween 80
Speed rate8 000 -10 000 rpm
Lipid
Aq.p hase
melting
heatingheating
emulsification
high-speed homogenization8 000 -10 000 rpm
solidification
Videira et al (2002).J Drug Targeting
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A.J. Almeida, 2007
D,L-hexamethylpropyleneamine oxime
(HMPAO)
N
NH
N
NH
OH OH
MeMe
Me
Me
Me
Me
+99mTcO4-NaCl
Radiolabelled LN
LABELLING AND ADMINISTRATION
Lipid Nanoparticles
Ultrasonic
nebulization
Labelling efficiency
Particle size
- Scintigraphy
anaesthetisedmale Wistar rats
Videira et al (2002).J Drug Targeting
A.J. Almeida, 2007
CHARACTERISATION OF 99mTc-HMPAO-LN
Beforeaerosolisation
Afteraerosolisation
D50 (nm) PI D50 (nm) PI
197.5 0.231 218.3 0.378
194.8 0.291 220.3 0.379
196.1 0.261 219.3 0.379
PARTICLE SIZE
Videira et al (2002).J Drug Targeting
ASPECT
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CHARACTERISATION OF 99mTc-HMPAO-LN
Labellingefficiency: 972%
Radiochemical Purity (TLC)
Sys I Sys II
t0
t30
99mTc-HMPAO
butanone 0.9% aq. NaCl
Sys I
t0
t30
Sys II
99mTc-HMPAO-LN
butanone 0.9% aq. NaCl
99mTc-HMPAO reduced-hydrolysed99mTc
Na 99mTcO4
Other hydrophilic species
A.J. Almeida, 2007
99mTc-HMPAO-LN99mTc-HMPAO
BIODISTRIBUTION
Videira et al (2002).J Drug Targeting
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A.J. Almeida, 2007
3 min 8 min 60 min
99mTc-HMPAO
99mTc-HMPAO-LN
BIODISTRIBUTION
Videira et al (2002).J Drug Targeting
A.J. Almeida, 2007
LYMPHATIC DISTRIBUTION OF 99mTc-HMPAO-LN
Videira et al (2002).J Drug Targeting
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Periaortic nodes Inguinal nodes
Axillarynodes
LYMPHATIC DISTRIBUTION
Videira et al (2002).J Drug Targeting
T1/2 10 minLungs
A.J. Almeida, 2007
BIODISTRIBUTION IN RATS 4h AFTER INHALATION
0
5
10
15
20
25
30
35
Lungs
Serum
Periaorticlymphnodes
Axillarylymphnodes
InguinallymphnodesLiver
Spleen
Thyroid
KidneysHeart
Brain
Smallintestine
Largeintestine
Testicles
Urine
%a
ctiv
ity/goftissue
Videira et al (2002).J Drug Targeting
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BIODISTRIBUTION OF 99mTc-HMPAO IN RATS AFTER i.v. INJECTION
Sharp etal (1986).J Nucl Med
21.51.519.80.315.80.215.41.7Intestines
8.61.010.40.410.20.413.91.9Liver
14.60.39.50.13.20.30.40.4Urine
8.60.39.40.210.51.111.00.4Blood
2.80.33.10.13.10.23.70.8Lungs
6.41.26.60.46.30.26.51.1Kidneys
% Dose in OrganTissue
2.00.21.80.22.10.22.10.1Brain
60 min30 min10 min2 min
A.J. Almeida, 2007
RADIOACTIVITY IN RAT LUNGS AFTER ENDOTRACHEAL ADMINISTRATION
(n=4; meansd)
0
20
40
60
80
100
120
3 8 15 30 60
Time (min)
%
activity/g
%A
ctivity/g
Videira et al (2006).J Microencapsulation
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ALVEOLAR CLEARANCE
Uptake of 1.1 m fluor escent polystyrene particles administered i.n. to miceEyles et al (2001). Vaccine
24 h15 min
Alv eolar clearance of 400 nm fl uorescent pol yst yrene part icl es admini stered i.n . to m iceByersdorfer et al (1995).J Immunol
6 h
Dendritic cells
24 h
Peribronchial LN
A.J. Almeida, 2007 Valentine and Kennedy (2001) In: Principles and Methods of Toxicology
PARTICLE UPTAKE AT THE LUNGS
LN
BALT
Blood circulation
Lymphatics
Alveolus
Interstitium
Pleura
M
Possible Mechanisms
Dissolution Macrophage phagocytosis Direct passage Uptake by vascular system Movement into the lymphatic system Axonal translocation to CNS
Elimination
Mucociliaryescalator Exhaled air
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TREATMENT OF BREAST CANCER LUNG METASTASIS
non-treated
LN-paclitaxel aero (last 15 days)
Taxol i.v. (last 15 days)
LN aero control
LN-paclitaxel aero (first 15 days)
Taxol i.v. (first 15 days)
LN iv control 100
100
25
100
100
75
100
% mice metastised pergroup
PRELIMINARY STUDY
Videira (2007). PhD Thesis
A.J. Almeida, 2007
LIPID NANOPARTICLES AS TOPICAL
DELIVERY SYSTEMS
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LIPID NANOPARTICLES AS TOPICAL DELIVERY SYSTEMS
BACKGROUND
Topical application of LN have been used with promising results either for therapeutic orcosmetic purposes (J enninget al. 2000; Mller et al. 2002).
SLN have shown some protective activity on skin surface, such as a UV-blockingpotential (Wissing et al. 2003).
SLN may be formulated in creams, gels,sprays.
SLN allow modulated drug release.
A.J. Almeida, 2007
MECHANISMS OF UV PROTECTION
Background:
Topical application of SLN have been used for therapeutic or cosmeticpurposes.
SLN act as an efficient particulate UV blocker (scattering effect) SLN andmolecular sunscreens have synergistic effect.
Reduced release of sunscreens from SLN when compared to emulsi ons
Reduced side effects
SLN may be formulated in creams, gels, sprays, allowing modul ated drugrelease.
Is it so ?
UV
Particulate sunscreen
Jenning et al. (2000); Mller et al. (2002); Wissing et al. (2003)
Molecular sunscreen
heath, ligh t
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z Lipid composition
gliceryl behenate(Compritol888 ATO) gliceryl palmitosterarate (Precirol ATO 5)
z Surfactant
Tween 80
z Speed rate
8 000 - 10 000 rpm
z Model Drugs
Octylmethoxycinamate (OMC)
Emulsification
Lipid
Aq.phase
meltingmelting
heatingheating
solidification
Homogenization
NANOPARTICLE PRODUCTION
A.J. Almeida, 2007
Formulation Time(months)
PI
D50 D90 D95
1 147,0 174,3 206,6 0,516
C (empty) 12 80,4 320,4 320,4 0,546
16 102,2 201,8 201,8 0,529
1 16,5 83,6 105,3 0,541
C-OMC 12 33,9 41,4 41,4 0,443
16 14,9 49,5 49,5 0,418
1 221,6 247,3 247,3 0,541
P (empty) 6 269,4 269,4 269,4 0,176
16 289,1 289,1 289,1 0,614
1 144,7 203,3 241,0 0,204
P-OMC 6 59,3 139,8 186 0,589
16 87,2 202,4 268 0,586
Particle size (nm)
NANOPARTICLE CHARACTERISATION: PHYSICAL STABILITY
Toscano et al. (2004) Eur J Pharm Sci, 23 (Suppl. 1)
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OMC RELEASE STUDIES
Membrane: Silicone
Receiving medium: 2% albumin in phosphate buffer (pH=7.4)
0
50
100
150
200
250
300
350
0,0 0,5 1,0 1,5 2,0 2,5 3,0
SQRT time (h)
Amountreleased(g/cm
2)
NL PR/OMC NL CP/OMC
Toscano et al. (2004) Eur Conf Drug Delivery Pharm Technol
A.J. Almeida, 2007
0
3
6
9
12
15
0,0 3,0 6,0 9,0 12,0
Time (h)
OMCamountinreceptorphase
(ug/cm2)
NL CP/OMC NL PR/OMC
IN VITRO PERCUTANEOUS ABSORPTION OF OMC
Membrane: Human skin
Receiving medium: 2% albumin in phosphate buffer (pH=7.4)
Toscano et al. (2004) Eur Conf Drug Delivery Pharm Technol
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IN VIVO STUDIES
16 female volunteers (24 to 54 years old; phototypes II and III 10-day application trial on the antero-external surface of both legs
3 different areas studied corresponding:
LN-OMC formulation
LN blank formulation
non-treated area (1 negative control per leg).
Experimental challenge: controlled exposure of volunteers to a UV radiation source(Sunny HB-406 Solarium, Phillips)
Parameter measured: skin colorimetry (erythema and melanisation indexes), TEWLand elasticity.
Fonte50 cm de
Radiao
A.J. Almeida, 2007
Erythema
0123
456789
10
0 3 5 7 10
Time (days)
Er
ythema(AU)
LN
Neg Cont
LN-OMC
Neg Cont
Toscano et al. (2004) Eur J Pharm Sci
56
5860
62
64
66
68
70
0 3 5 7 10
Time (days)
Mel
anisation(AU
LN
Neg cont
LN-OMC
Neg Cont
Melanisation
IN VIVO RESULTS
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Cutanova
Dr. Rimpler GmbH/Germany
Nanobase
Yamanouchi/Poland
MARKETED PRODUCTS
A.J. Almeida, 2007
PRODUCTION OF LIPID MICRO- OR NANOPARTICLES
USING SUPERCRITICAL FLUIDS
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AIM
Production of drug-lipid micro- and nanoparticles by a PGSS (particles fromgas saturated solutions) process, as an alternative method.
MAIN ADVANTAGES
9 Avoid the use of solvents
9 Particles are obtained as a dry powders, instead of suspensions
9 Mild pressure and temperature conditions
LIPID MICROPARTICLES USING SUPERCRITICAL FLUIDS
A.J. Almeida, 2007
LIPID MICROPARTICLES USING SUPERCRITICAL FLUIDS
PGSS System
(A) CO2 supply cylinder(B) Gas compressor(C) CO2 storage cylinder(D) Mixing cell(E) Collecting chamber
Lipid composition
Tripalmitin
Temperature
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LIPID MICROPARTICLES USING SUPERCRITICAL FLUIDS
0
20
40
60
80
100
0 6 12 18 24
Time (hours)
%Re
leased
T F L c o m m e r c i al
160 bar
180 bar
0,100,46
2,2
10
46120
140
160
180
0
2
4
6
8
10
12
14
Percentage
ofparticles
Diameter(m) Pressure
(bar)
Rodrigues et al (2004).J Supercrit Fluid
A.J. Almeida, 2007
LYSOZYME-CONTAINING LN PREPARED USING SUPERCRITICAL CO2
Rodrigues et al (2007). submitted
Lysozyme Biological Activity
Lysozyme particles produced fromsolutions with an ethanol mole fractionof C =0.28 showed no significant lossof activity.
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LN show suitable properties to become one of the most versatile nanoparticulatecarriers for drug and protein delivery.
Several drug incorporation techniques are applicable, including protein adsorption.
Inhalation may be an effective route to deliver LN, representing an alternative to theintraperitonial route for targeting colloidal carriers to the lymphatics.
LN present lymphatic tropism, thus providing the possibility of using radiolabelled LNas a lymphoscintigraphic agent, and allowing the direct delivery of cytotoxic drugs tolung cancer in limited or extensive stage.
The limited amount of particle retention in lungs may allow the use of LN as asustained release formulation in chronic lung therapy.
LN show an effective control release of lipophilic drugs, whici is suitable forpercutaneous absorption, with a minimum permeation thus making LN a safe vehiclefor sunscreen agents.
CONCLUSIONS (I)
A.J. Almeida, 2007
CONCLUSIONS (II)
In vivo results do not confirm early reports, i.e. LN on their own no not improvesignificantly TEWL and elasticity (results not shown).
Melanisation index was the only parameter that showed a trend, demonstrating thatLN present some protective capacity against UV radiation in vivo.
SCF techniques can be successfully applied to the formulation of solid lipid micro-
and nanoparticles with controlled-release characteristics.
Using a modified SCF technique, it is possible to produce LN containing intact andbiologically active protein molecules.
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MODERN LISBON
Thank you for your attention !