This document discusses methods for clinical testing, specifically 3D cell culture and organ-on-chip technologies. It notes that animal testing is time-consuming, costly, and often does not predict human outcomes. Organ-on-chip technologies use microfabrication and microfluidics to create microenvironments that better simulate human physiology and organs. This allows for testing of drugs and toxins using human cells in a way that may replace animal models. Examples discussed include a lung-on-a-chip to study pulmonary edema and a proposed "body on a chip" with 3D printed miniature organs to improve drug development and reduce costs.

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5. Methods for Clinical Testing

7. Animal Testing

8. Introduction
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Drug Testing in animal models is time consuming, costly and often does not predict the adverse effects in
humans and also 60% of animal models are not able to predict the toxicity .(Hamelton et al 2011)
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3D Culture models have recently garnered great attention .
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They promote levels of cell differentiation not possible in conventional 2D Culture systems.
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3D Culture systems is a large microfabrication technologies from the microchip industry and microfluidics.
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It approaches to create cell culture microenvironments
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This technology supports both tissue differentiation and recapitulate the tissue-tissue
interfaces, spatiotemporal micro gradients and mechanical microenvironments of living organs.
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This organs on chips permit the study of human physiology in an organ specific context, enable novel in
vitro disease models, and could potentially serve as replacements of animals used in drug development and
toxin testing

10. Failure to predict human drug
toxicity
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50% of Drug Candidates are failed in clinical trails due to the toxicity.
Post marketing with draw or limited use of drugs to adverse drug effects
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Drug metabolism as a key determinant of species-species differences in
drug toxicology.
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The development of safe and effective drugs is currently hampered by
the poor predictive power of existing preclinical animals that often lead
to failure of drug compounds and late development after they enter in to
the human clinical trials.
Huh et al 2011

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Miniature human organs made by 3D printing could create a "body on a chip" that enables better
drug testing. That futuristic idea has become a new bioprinting project.
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The 2-inch "body on a chip" would represent a realistic testing ground for understanding how the
human body might react to dangerous diseases, chemical warfare agents and new drugs intended
to defend against biological or chemical attacks.
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Such technology could speed up drug development by replacing less-ideal animal testing or the
simpler testing done on human cells in petri dishes — and perhaps save millions or even billions
of dollars from being wasted on dead-end drug candidates that fail in human clinical trials.
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Microscale engineering technologies are combined with cultured living human cells to create
microfluidic devices that replicate the physiological and mechanical microenvironment of whole
living organs.

15. 3D Cell Culture
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It is defined as the culture of living cells within the microfabricated devices having 3D
structures that mimic tissue and organ specific microarchitechture.
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Cell cultures in 3D matrix gels are referred to as 3D ECM gel cultures, 3D gel cultures or
conventional 3D cultures.
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Microengineering techniques such as Photolithography, replica modeling and microcontact
printing are well suited to create structures with defined shapes and positions on the
micrometer scale .
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That can be used to position cells and tissues control cell shape and function, and create
highly structured 3D culture microenvironments.

17.  An Organ-on-a-Chip (OC) is a multi-channel 3-D microfluidic cell culture chip that simulates the
activities, mechanics and physiological response of entire organs and organ systems.
 It constitutes the subject matter of significant biomedical engineering research, more precisely in bioMEMS.
 The convergence of Lab-on-Chips (LOCs) and cell biology has permitted the study of human physiology in
an organ-specific context, introducing a novel model of in vitro multicellular human organisms.
 One day, they will perhaps abolish the need for animals in drug development and toxin testing.
 Nevertheless, building valid artificial organs requires not only a precise cellular manipulation, but a detailed
understanding of the human body’s fundamental intricate response to any event.
 A common concern with Organs-on-Chips lies in the isolation of organs during testing. “If you don’t use as
close to the total physiological system that you can, you’re likely to run into troubles” says William
Haseltine, founder of Rockville, MD. Microfabrication, microelectronics and microfluidics offer the
prospect of modeling sophisticated in vitro physiological responses under accurately simulated conditions.

19. Microfluidics: The use of microfabrication techniques from the
IC industry to fabricate channels, chambers, reactors, and
active components on the size scale of the width of a human
hair or smaller
Credit: Dr. Karen Cheung, UBC ECE

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Sample savings – nL of enzyme, not mL
Faster analyses – can heat, cool small volumes
quickly
Integration – combine lots of steps onto a single
device
Novel physics – diffusion, surface tension, and
surface effects dominate
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This can actually lead to faster reactions!

21. • Three PDMS layers are aligned and irreversibly
bonded to form two sets of three parallel
microchannels separated by a 10-mm-thick
PDMS membrane containing an array of
through-holes with an effective diameter of 10
mm. Scale bar, 200 mm.
• After permanent bonding, PDMS etchant is
flowed through the side channels. Selective
etching of the membrane layers in these channels
produces two large side chambers to which
vacuum is applied to cause mechanical
stretching. Scale bar, 200 mm.
• Images of an actual lung- on-a-chip microfluidic
device viewed from above.
PDMS:Poly dimethylsiloxane
ECM : Fibronectin, collagen

22. Cating
PDMS
Membrane
Etching the
membrane
with TBAF
& NMP
Apply
Hydrostatic
Pressure
&Vacume
Bound
irreversibly
with the two
layers
Run the
etchant
solution
Pre
polymered
the layers
Photolithograph
y of
microchannels
Coat with the binding layer
and incubate at 65 c
overnight
Upper chamber
is Alveolar
chamber
Lower chamber
Blood flow

25. Biologically inspired design of a human breathing lung-on-a-chip microdevice.
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The microfabricated lung mimic device uses
compart- mentalized PDMS microchannels to
form an alveolar-capillary barrier on a
thin, porous, flexible PDMS membrane coated
with ECM.
• The device recreates physiological breathing
movements by applying vacuum to the side
chambers and causing mechanical stretching of
the PDMS membrane forming the alveolarcapillary barrier.
• During inhalation in the living
lung, contraction of the diaphragm causes a
reduction in intrapleural pressure (Pip), leading
to distension of the alveoli and physical
stretching of the alveolar-capillary interface.

26. .
IL-2 therapy is associated with vascular leakage that causes excessive fluid accumulation
(edema) and fibrin deposition in the alveolar air spaces.

27.  Endothelial exposure to IL-2 (1000 U/ml) causes liquid in the lower microvascular channel to leak
into the alveolar chamber (days 1 to 3) and eventually fill the entire air space (day 4).
• During IL-2 treatment, prothrombin (100 mg/ml) and fluorescently labeled fibrinogen (2 mg/ml) introduced
into the microvascular channel form fluorescent fibrin clots (white) over the course of 4 days.
• A fluorescence confocal microscopic image
shows that the fibrin deposits (red) in (D) are found on the upper surface of the alveolar epithelium
(green). (F) The clots in (D) and (E) are highly fibrous networks, as visualized at high image.

31. Different Organs on a Chip

33. Organ
Rationale
Cell Lines
Characteristics
Liver
Marrow
(Hematopoietic)
p450 activity
sensitive to chemo
dose limiting toxicity
Hep-G2/C3A
MEG-01
Tumor
(Sensitive)
Tumor (MDR)
(Resistant)
initial tumor primary
type
resistant tumors can
MES-SA
hepatoma
megakaryoblast line
attachment/suspension
inducible attachment
uteran sarcoma
sensitive to doxorubicin
variant
selected for resistance
to doxorubicin
MES-SA/DX-5

34. Application to Study Multidrug Resistance Suppressors
Other Tissues/ Debubbler
Device on peristaltic pump
in incubator
Sensitive Tumor
Cells (MES-SA)
Liver Cells (HepG2/C3-A)
Resistant Tumor Cells
(MES-SA/DX-5)
Bone Marrow Blood
Cells (MEG-01)
All cells labeled with cell tracker green before experiment

40. Thank You