1. Microfluidics
and Lab-on-a-Chip
for biomedical
applications
Chapter 6 : Cancer diagnostics & monitoring.
By Stanislas
CNRS
Université de Lyon, FRANCE
Stansan International Group

2. CONTEN
T
Chapter 1: Introduction.
Chapter 2 : Basic principles of Microfluidics.
Chapter 3 : Basis of molecular biology and analytical tools.
Chapter 4 : Micromanufacturing.
Chapter 5 : Lab-on-a-Chip & applications.
Chapter 6 : Cancer diagnostics and monitoring.

3. ABOUT
CANCER
A sinister aspect of the changes in a cell are
the transformations that occur with
accumulated genetic mistakes. Critical
"gateway genes" that control fundamental
metabolic and control pathways become
mutated and the cell becomes unstable and
starts unregulated growth. This is a highly
complex process which is called CANCER.
Most cancer deaths are not
caused by the growth of the
primary tumor, but result from
its
invasive
spread
to
secondary sites and the
subsequent
formation
of
metastasis.

4. ABOUT
2.9 million new casesCANCER than 1.7 million cancer deaths
of cancer and more
in Europe in 2004 (WHO report)
lung cancer was the commonest form of cancer diagnosed (13.2%) and of
cancer death (20%)
colorectal cancer was almost equally common (13%), it represented a
smaller proportion of deaths (11.9%).
among women, breast cancer was by far the most common, representing
27.4% of all female cases and it was also the biggest killer with nearly
130,000 deaths – 17.4% of the total.
the ageing of the European population will cause these numbers to
continue to increases by +50% in 2020.
it's mandatory to introduce completely new technologies for early
diagnosis, therapy and follow up of the cancer, which become a major
public health problem in both industrialised and in developing countries.

5. What are cancer markers ?
A cancer marker can be defined as a molecule, a process
or a substance, which is altered quantitatively or
qualitatively in pre-cancerous or cancerous conditions.
Thus, cancer markers can be a specific DNA mutation,
mRNA, protein or process (apoptosis, angiogenesis,
proliferation, etc.) measured by an appropriate assay.
The types of specimen in which cancer markers can be
detected may be of various nature: tissue, blood
(plasma/serum), saliva, urine…
Simple and nearly-non-invasive
diagnostics/ & monitoring

6. MOLECULAR DIAGNOSTICS
DNA chips have been extensively used for research
applications in academia and in industrial laboratories.
A big progress in bioinformatics is still needed in order
to be able to explore in a reliable way the DNA data in the
field of cancer.
It is expected that many DNA chip for cancer diagnostic
and monitoring will be developed in the next few years.
One of challenges for the Biochip industry is also the
integration of microarrays with microfluidics, in order to
achieve Microsystems, which include the extraction of
the genetic material (e.g. DNA or RNA from white blood
cells or from rare circulating tumor cells), purification
and amplification of the extracted material and, finally,
the analysis of this material with DNA chips.

7. Categories of markers and
they clinical utility
diagnostic cancer marker is a marker that will aid in
detection of malignant disease in a patient; a diagnostic
marker should exhibit both high levels of diagnostic
sensitivity and specificity;
prognostic marker gives the clinician a tool for estimating
the risk of disease recurrence and/or cancer-related
death;
predictive cancer marker will foretell how the patient is
going to respond to a given therapy;
monitoring markers are used during follow-up of patients
who do or do not receive anti-cancer therapy ; detection
of recurrence or remission.

8. Basic Diagnostic Test
Interpretation
Test positive
Test negative
Disease present
True positives (TP)
False negative (FN)
Disease absent
False positives (FP)
True negatives (TN)
Sensitivity is the proportion of patients with disease who test positive.
SE = TP / (TP+FN)
“False Negative Ratio” : FN / (FN + TP) = 1 - SE
Specificity is the proportion of patients without disease who test negative.
SP = TN / (TN + FP) >>> FP = TN / (1/SP - 1)
“False Positive Ratio” : FP / (TN + FP) = 1 - SP
Predictive value of a positive test is the proportion of patients with positive
tests who have disease. It measures how well the test rules in disease.
PVP = TP / (TP+FP)
Predictive value of a negative test is the proportion of patients with negative
tests who do not have disease. It measures how well the test rules out disease.
PVN = TN / (TN+FN)
back

9. IN REVIEWING LABORATORY TEST
RESULTS:
No single test is 100% accurate for specificity, sensitivity or
predictive value.
Any particular laboratory result may be inaccurate, for a variety of
reasons.
The degree of abnormality and the significance of the abnormality
are proportional. That is, marginal variations likely have less
significance than wide variations.
Multiple test abnormalities are more likely significant that single
test abnormalities.
Test results, when possible, should be compared over time,
preferably with results from the same laboratory. When possible,
comparison with the patient’s test results prior to illness may be
helpful.

10. C h r o n o lo g y o f g e n e tic
a n d m o le c u la r le s io n s .
N o rm a l
H y p e r p la s ia
M é t a p la s ia
D y s p la s ia
IS C
C a r c in o m a
3p9p17pA n e u p lo id y
P 5 3 m ut
U P A -S T 3 +
(E p ith ) R a s m ut
T é lo m e r a s e +
F H IT
RbP ro té a s e s u P A , S T 3
(S tro m a )
Le progrès rapide de la Génomique et de la Protéomique
conduit à la compréhension des bases moléculaires des
cancers et à la découvertes des nouveaux marqueurs.
Il est important de suivre la concentration de ces marqueurs.
La technologie Lab-on-a-Chip permettra le développement
de méthodes d’analyse rapides, robustes et fiables pour les
cabinets de médecin.

11. Exemple du suivi d’un marqueur cancéreux au
cours d’une chimiothérapie
Si un traitement est efficace,
on observe une baisse de la
concentration de marqueur.
Récession de la
tumeur
Graphique de la chute d’un marqueur
(HCG) au cours d’une chimiothérapie
d’une tumeur testiculaire.
Le grand intérêt des analyseurs proposés réside dans le fait que le
patient pourra bénéficier d'analyses beaucoup plus fréquentes
permettant de détecter à temps une possible évolution néfaste.

12. Need of frequent testing
A high level of HCG in the blood
indicates that a cancer of the
placenta called gestational
trophoblastic neoplasia (GTN) may
be present. This cancer continues to
produce HCG. Some testicular and
ovarian cancers resemble GTN
because they both arise from
reproductive cells called germ cells.
These cancers also make HCG and
this marker is used in their diagnosis
and in monitoring their response to
therapy. In the example below, note
almost day-by-day changes in the
concentration of the HCG marker.
Unfortunately, using standard
diagnostic techniques, nobody today
can be followed-up so closely. An
enormous advantage of the
instruments which could be
developed using Lab-on-a-Chip
Thanks to Lab-on-a-Chip,
patients could be monitored
frequently, which will allow to adjust
at time the treatment or, after the
treatment, to detect at time possible
recurrent cancer. However, up to
now there is no such devices.

13. Modern oncology must be based on
systematic and frequent quantitative
analysis of tumor markers which can :
help in diagnosis
be used as indicators of the clinical response of the
cancer to various treatment modalities, in order to
personalise or to adjust at time this treatment.
after the treatment, the patient must be followed up very
closely in order to detect at time a possible recurrent
cancer.
Unfortunately, using standard analytical techniques,
nobody today can be followed-up as closely as
necessary.

15. Full names of
markers cited
in the Table1
Details :
http://www.spendloveresearch.org/Re
searchPages/tumor&cancertypes.htm

16. Example of Cancer Markers

17. Approach
Separation of bio-molecules (proteins and/or DNA fragments) by electrochromatography carried out in multiple microfluidic channels; this separation is
coupled with nucleic acid hybridization reaction (DNA) or immunological reactions
(proteins) in liquid phase or using appropriate ligands bound to the separation
matrix.
Integration of new nano-structured materials into the microfluidic channels for
micro/nano filtering or as new type of matrix for Improved separation techniques of
bio-molecules - porous Si and nano-structured polymers.
Optical integration in the Lab-on-a-Chip, which will allow a dramatic reduction of
the dimension and price of the control unit.
Integrated optics in the Lab-on-a-Chip for the redistribution of the excitation light
and collection of the fluorescence signal - spectacular improvement of the
performances, multiple separation columns…
heterogeneous integration of various materials (Silicon, glass, polymers)
combining various functions : integrated optics, integrated microelectronics,
microfluidics - micro-nano components for advanced biological functions etc..
packaging issues for future bath fabrications of such devices on large
heterogeneous substrates involving silicon/glass/plastic wafers.
Operation with a drop of blood

18. The complete system will make use of disposable chip
sensors with a size of a credit card and a control unit
with a size of a book.
“heart of the system”
Power
supply
USB
USB
Finger print protected smart
card for data storage
The developed instruments will be autonomous and simple and
compatible with straightforward use (drop of blood). The
developed instruments must have sufficient autonomy and
simplicity for point-of-care operation (e.g. in physician
cabinets).

19. Cancer Prevention and Monitoring
Card

20. Principe 1 :
Détermination de la concentration des marqueurs
cancéreux, qui seront des protéines marqueurs,
présents dans un liquide physiologique facilement
accessible (sang, sérum…).
Approche 1 :
Séparation des diverses protéines de la solution par
électro--chromatographie dans un système
microfluidique Lab-on-a-Chip multicanal couplé à
des réactions immunologiques spécifiques
(protéines).

21. Principe 2
:
Intégration des Bio-puces à l’intérieur des
canaux micro-fluidique.
Variantes technologiques de la réalisation.
Bio-puces situées à l’intersection d’un
canal micro- fluidique avec un guide
optique intégré dans les Lab-on-a-Chip.
Calibration
-Optique
- Micro-fluidique
- Biologique

22. Approche 2
:
Détection de marqueurs biologiques, tels que ;
-ADN, protéines….dans le domaine de la santé, défense,
agro-alimentaire, contrôle d’environnement…
Dispositif micro-fluidique :
-miniaturisation, très petite quantité de la solution à
analyser…
-sensibilité accrue
Intégration : optique + micro-fluidique :
-miniaturisation du système extérieur de contrôle
-sensibilité accrue.
Détection multiparamétrique :
-meilleure interprétation des résultats.
-Point important : intégration Bio-puce & Lab-on-a-Chip

23. Functional Blocks
The detection of protein cancer markers will be obtained here by
the integration on the Chip of the following blocks :
The most general block diagram of the Chip for the detection
of molecular cancer markers is the following :

24. Minimally-Invasive Systems
for blood sampling and drug
delivery
Silicon Micro-Needle

25. Examples of plasma extraction
techniques

26. Enrichment and purification
of low abundance proteins
Par example by affinity chromatography :

27. Example of a simple geometry
of the electrophoretic separation
coupled with immunoreaction

28. CONTROL UNIT
Conception de l'architecture générale et choix des composants de
l'unité de contrôle.
Développement de la première maquette d'un analyseur composé
d’un capteur à utilisation unique interchangeable (comportant le
Lab-on-a-Chip) et d'une unité de contrôle de la dimension
approximative d'un livre.
Schéma général
de l’unité de
contrôle.

29. CONTROL
UNIT

30. CONTROL
UNIT
Photo de l’unité de contrôle en test.
Exemple d’acquisition réalisée au laboratoire.
Notons que le développement de l’unité de contrôle entre
parfaitement
dans
la
thématique
d’enseignement
d’électronique à l’Ecole Centrale de Lyon. Nous avons pu
ainsi intégrer la participation des élèves-ingénieurs dans
ce développement dans les cadres des « Projets

31. Simplified Example of 3D
integration of various materials
andPolymer « service » stage which
functions
contains electrical and vacuum
interconnections with the control unit,
related micro-channels, reservoirs for
chemical products and waste.
Glass stage for electr-chromatographic
separation of bio-molecules containing
integrated optics and nano-structured
materials in the separation channel.
Silicon stage for PCR, dielectrophoresis,
integrated photodetectors, integrated
electronic components (memory, DSP…)
Polymer stage which contains micro/nano filters (serum extraction),
interface to the external word (sample and chemical inlets) ; it plays
also a role of the packaging layer.

32. Alternative telecommunication
technologies (terrestrial, wireless &
mobile, satellite communications...)
Protocols, security,
encryption,
authentication,
privacy, etc...
Home of a patient,
of a doctor or a
small analytical Lab
Consider application
requirements, limitations and
tradeoffs
Medical Center
Ensure integration with existing
healthcare information
infrastructure (HII) and
consider compatibility with
next generation clinical
information systems

33. All the best for the
future

34. THANK YOU FOR YOUR ATTENTION
Any question ?