BMBF Go-Bio project: From single tissue culture to multi-organ-chips ...

BMBF Go‐Bio project: From single tissue culture to multi‐organ‐chipsFrom single tissue culture to multi organ chips

OUTLINE

the global substance testing dilemma

“human‐on‐a‐chip” – concepts

OUTLINE

project history

our multi‐organ‐chip approach scientific background

Summary & outlook

scientific background biological challenges & technical hurdles status quo

Dr. Uwe MarxHead of “Multi‐Organ Chip programInstitute of Biotechnology,

Summary & outlook

in sili‐tro ‐ synergies!?

AXLR8 satellite meeting / Berlin 22.05.2011

gy,Technische Universität BerlinGermanyuwe.marx@tu‐berlin.de

The dilemma of predictive substance testing

human cell culture test systems

animal models

Dynamic single organbioreactors

Static culture systems

„human on a chip“

&“virtual human”

Complexityphylogenic Proximity

systemic but not human

human

systemic but not humanAND not humane human but not systemic

Berlin 22.05.11 Uwe Marx AXRL8 satellite 2

Living systems on a chip – state of the art

Nature March 31st 2011: Technology feature: A living system on a chip. Vol 471 pp. 661‐665

miniaturized single‐organ‐chips

Huh D. et al Science 328, (2010) Guenther A. et al Lab Chip 10, (2010)

Hepregen

Hµrel

CellASIC

systemic multi‐organ‐chip concepts

Michael Shuler, Cornell University, U.S.A.S J H l L b Chi 11 389 392 (2011)Sung J.H. et al Lab Chip 11, 389‐392 (2011)Sung J.H., Kam C.; Shuler, M.L. Lab Chip 10 446‐455 (2010)Mahler G..J. et al. Biotechnol Bioeng. 104, 193‐205 (2009)

Kiichi Sato, University of Tokyo, JapanImura Y. et al Anal. Chem. 82, 9983‐9988 (2010)

Shuichi Takayama, University of Michigan, U.S.A.

Uwe Marx, Technische Universität Berlin, GermanySonntag F. et al , J. Biotechnol 148, 70‐75 (2010)

AXRL8 satellite 3Berlin 22.05.11 Uwe Marx

History …

… of the GoBio project, targeting for a systemic solution for long term substance testing ultimately predictive to human substance exposure

2007 U. Marx : Why is human tissue culture important to drug discovery? In: Drug Testing In Vitro – Breakthroughs and Trends in Cell Culture Technology, Eds. Marx and Sandig; Wiley‐VCH Verlag GmbH&Co.KgaA.

20112008 2009 2010

DM T‐chipA Eadsorption

01.03.10

Partnership:TU Berlin, BCRT BerlinFraunhofer IWS DresdenFraunhofer IGB StuttgartGESIM GmbH Dresden

Phase 1: ~ €3 Mio/3 years, distributionmetabolism

toxicityPhase 2: spinoff financing

adsorption

excretion

Partnership:

BMBF GO-BIO(multi-organ-chip)

GESIM GmbH Dresden

Routes i.v.. dermal oral

FP7 Health 2010

pTU Berlin, GermanyUniversitè de Geneve, SwitzerlandProBioGen GermanyInnsbruck Medical University, AustriaFraunhofer IGB IWS ITEM Germany

inhalation


Alternatives … Fraunhofer IGB , IWS, ITEM, Germany

Track record in dynamic bioreactor development

Human bone marrow

1991‐1994 Marx et al Nature's UK Product Review 569; 34, 1993

leukemia research

Human micro capillaries

1997‐2001 trans‐vascular leukocyte traffic tumor invasion mechanisms

Human artificial lymph node

2001‐2007 tumor invasion mechanisms

immunogenicity testing

Giese Ch et al. J. Artificial Organs (10):803‐808. 2006 and J. Biotechnology; 148, S. 38‐45; 2010

immune reactivity patternHuman artificial lymph node immunogenicity testing

Modular mini bioreactor

2006‐2008

high content tissue toxicity testing

Ski i

2007‐2009

ki l f i

Modular mini‐bioreactor high content tissue toxicity testing

Skin regeneration system skin transplant manufacturing

id lf bl th h i i i t ifi i hit t

Long term homeostasis at 100% viability?

Berlin 22.05.11 Uwe Marx 5AXRL8 satellite

organoid self assembly through angiogenesis into organ specific micro‐architectures

The crucial role of dynamic blood circulation

nutrient and oxygen transport through blood plasma and red blood cells

blood tissue barrier and neo angiogenesis through endothelial cells

interconnection of organs to create an organism

blood‐tissue barrier and neo angiogenesis through endothelial cells

tissue repair and immune response through white blood cells

kidney vasculaturekidney vasculature


Natures unique organ building blocks …

bibio-molecule cell

organs human

organism organoids

… are of µl‐scalei t lti l i li d ll t

7AXRL8 satellite Berlin 22.05.11 Uwe Marx

… consist multiple specialized cell types

Size, architecture & fluid dynamics matter!

A paradigm of stringent correlation between architecture, dynamic microenvironment and

skin segment withhair follicles

liver lobulus

, yfunctionality applies to all levels of biological

existence on earth


hair follicles

Biological challenges & technical hurdles

long term performancecapillary blood circulation

organoids of different functionality

performance

0 organoids of different functionality

micro‐pump28 days

control unit90 days

standardized chip platform1 year

OECD guidelines forrepeated dose oral toxicitytesting of chemicals: TG407 (28-days, rodents); TG408 and TG 409 (90 da s


408 and TG 409 (90-days,rodents and non-rodents)

Standardized chip fabrication platform establishedLayer Material Function

4 PDMS(2000 µm)

Channels and reservoirs

3 Glass (200 µm)

Closing layer with holes

2 Sili Ch l2 Silicon (400 µm)

Channels, organ growth segments & stem cell niches

1 Glass Sensors and heater

Sonntag et al Design and prototyping of a chip‐based multi‐micro‐organoid culture system for substance testing, predictive to human (substance) exposure J. Biotechnology 148, 70‐75, 2010

(200 µm)

Rapid prototyping of any sophisticated designThermal oxidation of Si‐Wafers

the size of a standard microscopic slide

organ culture spaces accessible for live tissue imaging

2 months

Anodic bounding

Wafer cutting and structuring PDMS‐layer

Laser structuring of silicon and glass organ culture spaces accessible for live tissue imaging

single use biocompatible materials

adequate micro‐systems fabrication procedures


Micro‐pump and control unit in place

peristaltic micro pump

t l it

Control of up to 8 pneumatic actors Free configurable e.g. 1 pump + 5 valves L 0 1 b Hi h 1 2 bcontrol unit Low‐pressure: 0 .. ‐1 bar; High‐pressure: 1 .. 2 bar Control via buttons and lamps Remote‐control via serial interface Platform independent PC control software(e g WINDOWS LINUX MAC)


(e.g. WINDOWS, LINUX, MAC)

Three pillars of the tissue engineering program

capillary blood circulationskin organoid researchchip basedblood circuit investigative dynamic

liver sinusoid model

investigative dynamic skin culture chip

liver organoid research

liver sinusoid model(technical perfusion)

p(technical perfusion)

vascularized liver and skin equivalent

epidermis / dermis equivalents Including hair follicles

O2

and skin equivalent

knowledge knowledge

Lee P.J. et al (2007)Biotechnol. Bioeng . 97,


Investigative dynamic skin culture chiphair follicle and skin self assembly in static culture

i i

hair follicle

in vitroneopapillae

in vivo

Lindner et al.; J. Biotech 152:108 (2011) pan‐cytokeratin trychohyalin

epidermis/dermis14,3 mm2 transwell

continuous medium perfusion(valves and reservoirs )

transwell insert for skin tissue culture

peristaltic micro pump

Dynamic instead of static human skin equivalent culture should support hair follicle ingrowths into a


Dynamic instead of static human skin equivalent culture should support hair follicle ingrowths into aepidermis‐dermis equivalent.

Investigative dynamic liver sinusoid model

Designed omocer® photo resist inserts in placepO2

Peri‐portal:60 mmHg

Peri‐venous:25 mmHg

500 µm

~400 hepatocytes

Zone IIIZone I

50 µm

glycolysislipogenesiscyt p450 baseddrug detoxification

ß‐oxydationgluconeogenesis

cholesterol synthesis

25

Disse space bile segregation stellate cell function d fl

zonal albumin secretion drug metabolism (CYP3A4) bile synthesis (CYP7A1)


adequate flow rates … static 7day culture of human fibroblast(2‐Photon microscope Z‐stack, 10µm steps)

bile synthesis (CYP7A1)

Summary

capillary blood circulation (under investigation)capillary blood circulation

organoids of different functionality

(under investigation)

vascularized liver and skin ‐multi‐organ chip (upcoming)vascularized liver and skin multi organ chip (upcoming)

investigative dynamic skin bioreactor investigative dynamic liver sinusoid bioractor (under investigation)

(in place)

micro‐pump (in place)

control unit (in place)

(i l )standardized chip platform (in place)


“In sili‐tro” synergies !?“virtual human”

in silico„human on a chip“

in vitro

be ‐ chip bio ‐ chip

knowledge basedData quality bio‐fit ‐ based

degree of emulationPrediction isrestricted to:

known mechanismsand their combinations

TGN 1412 – anti CD28 super‐agonistoverrides the basic immune control


mechanisms

Acknowledgement

Fraunhofer IWS, DresdenFrank SonntagNiels SchillingRoland Lauster

Institute of Biotechnology

R k H l d

Niels SchillingUdo Klotzbach

A ik J i kReyk HorlandGerd LindnerIlka WagnerEva Materne Funding

Annika JaenickeAgnes SchumacherLuzie Reiners‐SchrammDesiree Geßner

Katharina CierpkaKatharina SchimekSilke HoffmannS B i k

FundingBMBF: GO‐Bio

Karolina TykwinskaDevasena KanthiJennifer BinderShirin FatehiSven Brincker

Alexandra LorenzMatthias Gruchow

Shirin FatehiMark Rosowsky


Announcement


BMBF Go-Bio project: From single tissue culture to multi-organ-chips ...

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