Jornada BIOINTEGRASAUDE 2013

SCHOOL OF PHARMACY

NANO-DRUG DELIVERY María José Alonso

http://webspersoais.usc.es/mariaj.alonso

PHARMAprocess- Innovation Forum in Pharmaceutical Process, Barcelona, Oct 2015

http://webspersoais.usc.es/mariaj.alonso

1960---

J. Folkman

P. Speiser

A. Bangham

http://www.fasebj.org/content/vol19/issue13/images/large/z380110532480001.jpeg

OUR WORK

More than 40 Nano-drug delivery products

OUR EXPERIENCE

FORMULATION OF COMPLEX MOLECULES

Peptides: sCT, Insulin, Ag… Proteins: IFN, FGF, Ag…

DNA, siRNA, miRNA

NANOTECHNOLOGIES NANOCOMPOSITIONS DESIGN Polyesters, Polysaccharides Polypeptides, Proteins Lipids

Oral Delivery Nasal Delivery Ocular Delivery

Parenteral Delivery

OVERCOMING BIOLOGICAL BARRIERS

1992- NANOBIOPHARMACEUTICALS

OUR EXPERIENCE AT THE USC

The rational design of delivery systems for complex molecules

MATERIALS: Polysaccharides, polypeptides, surfactants, oils

100 nm

THE KEY FEATURES: Small size: 50-300 nm Multiple layers Functionalized surface Negative/positive surface charge Hydrophobic/hydrophilic core

One or more drugs Protective agents Controlling agents Targeting agents

TOOL-BOX FLEXIBILITY

KEY ACHIVEMENTS

LATEST TECHNOLOGIES

EASY SCALABLE TECHNOLOGIES - Solvent displacement

- Self-emulsification

MULTI-LAYER POLYMER NANOCAPSULES

BIOMATERIALS: Polyglutamic acid Polyasparagine Polyarginine Hyaluronic acid…

THE KEY: POLYMER/SURFACTANT/OIL Self-assembling Ionic/ hydrophobic…forces

SELF-EMULSIFICATION TECHNIQUE

Ocular

Nasal

Intestinal Skin

Systemic barriers

Targeting

Cellular barriers

The goal: helping drugs overcoming barriers and reaching their targets

OCULAR DRUD DELIVERY SURFEye, NABBA Dry eye, corneal healing

CURRENT APPLICATIONS

ORAL PEPTIDE DELIVERY TRANS-INT-eu

NASAL VACCINE/PEPTIDE DELIVERY- NIH

CANCER NICHE, NANOFAR, WORLDWIDE Immunotherapies

TOPICAL APPLICATION: Psoriasis

Diabetes

LYMPHATIC TARGETING: THERAPEUTIC VACCINES Auto-immuno diseases

Multiple esclerosis Diabetes

CARDIO- DRUG DELIVERY Cardiac ischemia

CURRENT APPLICATIONS

Rat nasal mucosa

KEY ACHIVEMENTS: APPLICATIONS

NASAL/ORAL DRUG/GENE/ANTIGEN DELIVERY

Drugs: Insulin sCalcitonin Antigens: Tetanus Toxoid, DT Hepatitis B, HIV

Tobío M. et al. Pharm. Res., 15, 270 (1998)

Enhancement of systemic absorption of complex drugs

Needle-free vaccine delivery systems Thermostable vaccines

NASAL VACCINE DELIVERY

http://www.gatesfoundation.org/

Protection Presentation Multi-adjuvant

MULTIFUNCTIONAL POLYMER NANOCAPSULES

Antigen (capsomer/protein/peptide) (encapsulation/adsorption/ chemical linking)

Polymer/ Immunomodulator (poly(I:C)

Oily core (Imiquimod)

S. Vicente et al. JCR, 2014

…12 PCS peptides A cocktail of 12 nanopackaged peptides

J. C. Pinto, Collaboration with University of Manitoba

NASAL NANOPACKAGED SIV PEPTIDE ANTIGENS

IN

S SIISIV challenges

Collaboration with University of Manitoba

IN

55

65

75

85

95

105

-30 0 30 60 90 120 150 180 210 240 270 300

Pla

sma

gluc

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(% o

f ini

tial l

evel

)

Time (min)

*

* *

Insulin CS NP 1 CS NP 2

Teijeiro-Osorio et al. Biomacromolecules 10, 243 (2009)

CHITOSAN-BASED NANOPARTICLES FOR NASAL INSULIN DELIVERY

50

60

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100

110

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Insulin solution

NP

LNP1

LNP2

LNP3

24

CHITOSAN-BASED NANOPARTICLES FOR ORAL INSULIN DELIVERY

ACADEMIC PARTNERS

BIG PHARMAs: Sanofi

SMIs/CRO: Veneto Sero Sci Sigmoid

Nanotechnologists (4)

Barriers biology (4)

Tox/Immunology (3)

Chemical biology (1)

Peptides Preclinical evaluation Nanocarriers design, development and assessment of formulations

www.TRANS-INT.EU

Mucus layer

Agregation Pancreatine/salts

mucoadhesion

Mucopermeation

Nanoparticle

Nanocapsule

Micelle

Easy to scale-up, mild technologies!!!

More than 1,000 prototypes

Mechanistic issues

PK/PD analysis

4 peptides

12

FINAL DOSAGE FORM

MULTI-TARGET ONCOLOGICAL THERAPIES

QUIMIOTHERAPY

Passive/Active targeting: THE TARGET CELLs

IMMUNOTHERAPY

Cytotoxic drugs mAb

siRNA/miRNA

Immuno-modulators

IMMUNOGENIC CELL DEAD IMMUNOACTIVATION

TAMs

MDSCs

Cancer Cells

CELL DEAD

THE « LYMPHOTARG » EUROPEAN CONSORTIUM

Bioactive molecules: Cytotoxic drugs: Plitidepsin Docetaxel Curcumine Gemcitabine mAb Immunomodulators: Imiquimod Poly(I:C) Polynucleotides: miRNA/siRNA

THE « NICHE » EUROPEAN CONSORTIUM

THE WORLDWIDE CANCER RESEARCH CONSORTIUM

MULTI-LAYER NANOCAPSULES: MULTITARGET ONCOLOGICAL THERAPIES

PK analysis: Nanocapsules improve Docetaxel tumour accumulation, compared to Taxotere®,

(Orthotopic lung cancer model)

39 times higher drug accumulation than Taxotere®

Nanocapsules increases docetaxel lymphatics accumulation, compared to Taxotere®

(Orthotopic lung cancer model)

LUNG

Control TXT targeted NCs0

5000

10000

15000

20000

25000

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µg p

rote

in

MEDIASTINAL NODES

C o n t r o l T X T t a r g e t e d N0

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10000

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µg p

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Docetaxel-loaded Targeted HA Nanocapsules: the only treatment capable of reducing metastasis

37 days 47 days 47 days 37 days 47 days 47 days

BLUE: DAPI RED: FALOIDIN GREEN: MONOCLONAL ANTIBODY

Internalization of mAb-loaded nanoparticles

Collaboration with Ana Cadete, Dolores Torres and Gema Moreno

Listening to industry

Listening to

clinicians

- Advances in material sciences and engineering

- Advances in pharmaceutical technology

- Advances in cell and molecular biology

THE FUTURE

Jornada BIOINTEGRASAUDE 2013

SCHOOL OF PHARMACY

Nanobiopharmaceuticals Lab

M.J. Alonso

“Bill & Melinda Gates Foundation”, National Institutes of Health, European Commission, Ministerio de Economía y Competividad, Xunta de Galicia

Nanobiopharmaceuticals Lab M.J. Alonso

THERAPEUTIC POTENTIAL OF PEPTIDE DRUGS AND ANTIGENS

Hay M. et al.: Nat. Biotech. 32, 40-51, 2014

Large molecules

Small

10 times higher success of large molecules!

Preclinical Phase I Phase II Phase III

2%

10%

4%

15%

10%

25%

50%

60%

Small molecules

Large molecules

() Insulin solution () CS-GM nanoparticles

0 4 8 12 16 20 2440

60

80

100

120

Serum

Gluc

ose le

vels (

% of

basal

)

Time (hours)

*

* *

* *

*

*

* *

* *

* * *

* *

Maria Alonso et al. (non published results)

ORAL INSULIN DELIVERY

KEY ACHIVEMENTS: APPLICATIONS

1. To facilitate the penetration of complex drugs across the cornea 2. To target and deliver drugs to the back of the eye Peptides, proteins, nucleic acids

Cornea transfected for 5 days

OCULAR DRUG/GENE DELIVERY

Colaboration with C. Pastor, M. Calonge, J. Merayo

Epithelium surface 5 µm depth 10 µm depth

CS/SBE-β-CD nanoparticles

z-section

y-sections

Calu-3 differentiated cells:

Do the nanoparticles enter epithelia?

NASAL PEPTIDE DELIVERY

Study of CS Nanocapsules in Caco-2 cells

ORAL PEPTIDE/PROTEIN DELIVERY

Do the nanoparticles enter epithelia? -

VS- P1 36

Venice – September 8th 2011

VS- P1 37

Intercostal space left lung

1x106 células A549-luc injected in SCID mice

Orthotopic methastatic lung cancer model

Experimental groups: - 5 control mice without treatment - 5 mice iv. Taxotere® 10 mg/kg - 5 mice i.v. Taxotere® 20 mg/kg - 5 mice i.v. NCs PGA-PEG DCX 10 mg/kg - 5 mice i.v. NCs PGA-PEG DCX 20 mg/kg

Antitumor efficacy in orthotopic methastatic lung cancer model

E. Borrajo. colaboration with A. Vidal and M. Garcia-Fuentes

time/days

(Orthotopic A549-tumor bearing mice)

Antitumor efficacy in methastatic orthotopic lung cancer model

E. Borrajo. collaboration with A. Vidal and M. Garcia-Fuentes

Docetaxel-loaded PGA-PEG Nanocapsules are less toxic than Taxotere®

(SCID non tumor-bearing mice)

Mann Whitney test: ** p< 0.01

E. Borrajo. colaboration with A. Vidal and M. Garcia-Fuentes

Docetaxel-loaded PGA-PEG Nanocapsules inhibit tumor growth as compared to Taxotere®

Mann Whitney test: * p< 0.05, ** p< 0.01

Measurement of luciferase activity

Docetaxel-loaded PGA-PEG Nanocapsules inhibit metastasis as compared to Taxotere®

Mann Whitney test: * p< 0.05, ** p< 0.01

Docetaxel-loaded Nanocapsules (NCs) increases lymphatics accumulation, as compared to Taxotere® (TXT), in orthotopic lung cancer model

Docetaxel-loaded Nanocapsules (NCs) exhibit the same accumulation as Taxotere® (TXT) in heart, kidney, spleen and liver. No detectable levels were found in brain tissue.

24h

5

50

500

5000

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ug D

CX/

mL

plas

ma

time/h

pharmacokinetics

TXT

HA NCs

targeted HA NCs

t1/2 (h) AUC 0-inf obs ((mg/L).h) Cl_obs (L/kg.h) MRT 0-inf_obs

(h) VdB (L/kg)

TXT 5,40±1,27 2576,15 ± 346,71 0,0039±0,00053 2,89±0,2420 0,0113±0,00060

HA NCs 5,75±0,55 4400,45 ± 1110,96 0,0024±0,00062 4,25±0,5017 0,0099±0,00137

Targeted HA NCs 8,43±0,91 3162,72 ± 707,1 0,0033±0,00075 6,33±0,4195 0,0209±0,00619

Docetaxel-loaded Nanocapsules (NCs9 shows increased blood circulation time, as compared to Taxotere® (TCT), in orthotopic lung cancer model

Pharmacokinetic parameters using a non compartimental model

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