Electroporation based Technologies and Treatments

Damijan MiklavčičUniversity of LjubljanaFaculty of Electrical EngineeringLjubljana, SLOVENIA

“Cells can be funny. Try to grow them with a slightly wrong recipe, and they turn over and die. But hit them with an electric field strong enough to knock over a horse, and they do enough things to justify international meetings, to fill a sizeable book, and to lead one to speak of an entirely new technology for cell manipulation.”

Adrian Parsegianfrom the foreword to the

Electroporation and Electrofusion in Cell Biology

Edited by E. Neumann, A. S. Sowers and C. A. Jordan

Electroporation based Technologies and Treatments

• Molecular cell biology research• Protein insertion into cell membrane• Cell fusion• Gene expression silencing by siRNA• Electrochemotherapy• Genetherapy based on electro genetransfer• Transdermal drug delivery • Tissue ablation • Biotechnology• Water and liquid food sterilisation

timeElectrodes

Electric pulse generator

Tumour

Chemotherapeutic surrounds the cells

Increased membrane permeability allows access to the cytosol

Pores reseal

Injecting chemotherapeutic

Electric pulse application

Electrochemotherapy = = chemotherapy + electroporation

Skin metastasis of squamous cell carcinoma of the supraglottis

Skin metastasis of malignant melanoma

Before treatment One year after ECT

EU 5th FP: Quality of Life

Cliniporator QLK3-1999-00484 (2000-2003)– Electro-medical device for ECT

ESOPE QLK3- 2002-02003 (2003-2004)– European Standard Operating Procedures for

ECT

Final outcome of ESOPE study

4,7%10,5%11,1%

73,7%

0

10

20

30

40

50

60

70

80

90

CR PR NC PD

%

171 NODULES

Objective Response Rate: 85%No Response: 15%

GeneTherapy of diseases with altered angiogenesis:

Psoriasis and Malignant Melanoma

A B C 100 ms 1500 ms 3 min

E

A B C

E

Electroporation of cells: Experiments and modelling for better understanding

A B C

Cell (electro)fusion

• Monoclonal antibody production

• Cancer immunotherapy• Receptor insertion into

membrane• Tissue regeneration• Research (dissease

models)

Dielectrophoresis

Pozitivna dielektroforeza Negativna dielektroforeza

V homogenem električnem polju ni dielektroforeze

Biochip and results of experiments

• CHO cellls• Medium conducivity 15 mS/cm • negativve dielectrofporesis• frequency 500 Hz• amplitude 4 V

32(1 ) (1 2 ), 1 /

(2 ) (1 )m i

p mm i

d R

• Molecular cell biology research• Electrochemotherapy• Electrogenetherapy• Gene expression silencing by siRNA• Transdermal drug delivery • Protein insertion into cell membrane• Cell fusion• Biotechnology• Water and liquid food sterilisation• Tissue ablation

Electroporation based Technologies and Treatments

LBK expertise and resources:

Cell biology and chemistry labLipid bilayer setup for preparation

of planar lipid membranes.

• chamber for lipid bilayers• microscope with cold-light illumination for lipid bilayer observation (SZF 30, Olympus, Germany)

LBK expertise and resources:

Cell biology lab

Prototype of an electroporator developed in LBK connected to digital oscilloscope LeCroy 9310C.

Inverted phase contrast microscope Olympus CK40 for routine work with cell cultures & lipid vesicles.

LBK expertise and resources:

Cell biology lab

System for dynamic imaging of cells and lipid vesicles.

• Zeiss Axiovert 200 epiflurescence inverted microscope (Zeiss, Germany)• high resolution cooled CCD camera (VisiCam1280) • sensitive high-speed cooled CCD camera (BOOST 2000, Diagnostic Instruments, GB)

Other equipment available:• Spectrofluorometer (Jasco, Japan) • Microplate reader (Tecan, Switzerland)• Field jump system for fast monitoring optical changes in lipid vesicles • Computer facilities• Software: Mathematica, Matlab with different toolboxes; • Numerical modeling software: MSC/EMAS, Maxwell, COMSOL

Multiphysics• NIRO2-X2: near infrared spectrophotometer for measurement of oxygenation and

blood perfusion changes in tissue (Keele University, U.K.)• OxyLite 2000: 2-channel instrument for luminescence-based fiber-optic oximetry in

tissue; OxyFlo 2000: 2-channel instrument for laser Doppler flowmetry in tissue; OxyData 2000: data acquisition unit for OxyLite and OxyFlo instruments (Oxford Optronix, Oxford, U.K.)

Equipment available and used in other institutions:• flow cytometer (FACSort Becton Dickinson, Mountain View, CA) Medical faculty,

University of Ljubljana• Confocal microscope (Leica TCS SP5, Germany)

List of other equipment

SLOVENIAN:• Institute of Oncology, Ljubljana• Institute of Rehabilitation, Ljubljana• Medical Faculty, University of Ljubljana• Institute Josef Stefan, Ljubljana

INTERNATIONAL:• Institut Gustave-Roussy (Villejuif, France) • Igea S.r.l. (Modena, Italy) • Universität Bielefeld (Bielefeld, Germany) • Faculty of Electrical and Computer Engineering (Zagreb,

Croatia)• Institut de Pharmacologie et de Biologie Structurale

(Toulouse, France) • Université catholique de Louvain (Brussels, Belgium) • Orthologic Corp. (Tempe, Arizona, USA) • Sewanee University of the South (Sewanee, Tennessee,

USA) • Johns Hopkins University (Baltimore, Maryland, USA)

Collaborating Institutions

Laboratory for bio cybernetics

Peter, Katja, Marko, Jakob, Tadej, David, Matej K., Tomaž, Matej R., Denis,Blaž, Damijan, Ivan, Janez, Mojca, Stanislav, Gorazd, Vilko, Selma, Čarli,

Nataša, Maša, Alenka, Barbara (mising from the photo: Anže, JernejaTina, Duša)

http://lbk.fe.uni-lj.si

Thank you !

damijan.miklavcic@fe.uni-lj.si

http://lbk.fe.uni-lj.si