SLN Colloidal Handout-portugal_2007

7/30/2019 SLN Colloidal Handout-portugal_2007

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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

[email protected]

Universidad de Sevilla, Facultad de Farmacia, November 2007

A.J. Almeida, 2007

MICROPARTICLES AND NANOPARTICLES:

THE BASIC CONCEPTS


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A.J. Almeida, 2007

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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A.J. Almeida, 2007

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


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


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



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A.J. Almeida, 2007

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



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Emulsion Polymerization


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



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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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A.J. Almeida, 2007

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.J. Almeida, 2007

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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A.J. Almeida, 2007

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


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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A.J. Almeida, 2007


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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A.J. Almeida, 2007

0

100

200

300

400

500

Distilledwater

3%Tween 80

3% NaCholate

3% Poloxamer188

Lysozymeactivity(Units/ml)

Day 1

Day10


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


-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

(%)


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



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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


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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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


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


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



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3 min 8 min 60 min

99mTc-HMPAO

99mTc-HMPAO-LN

BIODISTRIBUTION


A.J. Almeida, 2007

LYMPHATIC DISTRIBUTION OF 99mTc-HMPAO-LN



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Periaortic nodes Inguinal nodes

Axillarynodes

LYMPHATIC DISTRIBUTION


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



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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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A.J. Almeida, 2007

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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A.J. Almeida, 2007

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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A.J. Almeida, 2007

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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A.J. Almeida, 2007

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


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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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 !

Documents

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