Enzyme Electrodes and Glucose Sensing for Diabetes

Enzyme Electrodes and Glucose
Sensing for Diabetes
Anthony P.F. Turner
Biosensors and Bioelectronics Centre, Linköping University,
Sweden
[email protected]
www.ifm.liu.se/biosensors
TFYA62, 24 February 2015
Enzyme Electrodes and Glucose Sensing
for Diabetes
2
•
Enzyme electrodes
•
Diabetes diagnostics
•
In vitro glucose sensors – the “finger stick” test
•
The advent of mass production
•
Continuous glucose monitoring
•
Minimally and non-invasive monitoring
•
Some ethical considerations
Example of a conventional enzyme
electrode
Key bioelectrochemical reactions
D-glucose + H2O + O2
Anode
H2O2
D-glucose + 2 Medox+
(NH2)2CO + 2H2O +
gluconic acid + 2 Medred
HCO3- + 2NH4+
ADH
+
NAD
NADH
Anode
gluconic acid + H2O2
2H+ + O2 + 2e-
GOx
Urease
+
H
C2H5OH +
GOx
2NH3 + 2H+
C2H5O + NADH
NAD+ + 2e- + H+
Clinically Important Enzyme Electrodes
Electrode
Enzymes
Amperometric
Oxygen electrode, hydrogen peroxide
detection at platinum or carbon electrodes
or mediated amperometry
Oxidases e.g. Glucose oxidase (GOx),
Lactate oxidase, Galactose oxidase,
Pyruvate oxidase, L-Amino Acid oxidase,
Alcohol oxidase. Oxalate oxidase,
Cholesterol oxidase, Xanthine oxidase,
Uricase.
Platinum, carbon, chemically-modified,
mediated amperometric electrodes
Dehydrogenases e.g. Alcohol
dehydrogenase, Glucose dehydrogenase
(NAD and PQQ), Lactate dehydrogenase
Potentiometric
Ammonia Gas-Sensing Potentiometric
Electrode, Iridium Metal Oxide
semiconductor probe
Creatinase, Adenosine deaminase
pH Electrode, Filed-effect Transistor (FET)
Penicillinase, Urease,
Acetylcholinesterase, GOx
Carbon Dioxide Gas Sensor
Uricase, inhibition of dihydrofolate
reductase, salicylate hydroxylase
Diabetes Diagnostics:
A Special Case
Newman, J.D. and Turner, A.P.F. (2005). Home blood glucose biosensors:
a commercial perspective. Biosensors and Bioelectronics 20, 2435-2453.
Diabetes Prevalence
• Diabetes is an immense and growing public health issue:
• Fastest growing chronic disease in the World; 50% increase expected by 2035 (IDF
2014)
• 4.9 m deaths from diabetes in 2014, projected to rise by >50% in the next 10 years
(WHO, 2014)
• Afflicts around 5% of the world’s population; 387 m diabetics worldwide; 46%
undiagnosed (IDF 2014)
• 52m people in Europe or 8.1% of the population have diabetes and their healthcare
costs are at least double that of non-diabetics; 11% of adult healthcare costs relate to
treatment of diabetes (IDF 2014).
• In the USA, 9.3% of all citizens and 25.9% of Senior Citizens afflicted (NDSR 2014)
• Asia now has the world's two largest diabetic populations (20-79 years) – China
(98.4) & India (65.1 m) cases (WHO 2013)
• There is no known reliable cure!
Diabetes – Influential Studies - DCCT
• Intensive therapy (including frequent monitoring of glucose) can reduce
the risk of complications by 60%
• Intensive therapy increases the risk of hypoglycaemia
• All diabetics should benefit in the longer term by improved monitoring
and control of blood glucose
• Diabetes Control and Complications Trial. New England Journal of
Medicine, 329(14),1993 http://diabetes.niddk.nih.gov/dm/pubs/control/
The Diabetes Health Care Space
BGM Growth rates
2011/12 = 2.2%
2012/13 = 3.0%
2013/14 = 1.1%
14
18.0%
12.4
16.0%
12
10.6
14.0%
10
9.1
12.0%
7.9
8
10.0%
6.9
6.2
6
4.4
4
8.0%
5.6
4.7
5.0
6.0%
3.9
4.0%
2
2.0%
0.0%
0
2000
MedMarket Diligence, LLC; Report #D510
2001
2002
2003
2004
2005
Gluco se M o nito ring
2006
2007
2008
2009
Gro wth
Frost & Sullivan
2010
Growth and distribution of diabetes patients
A brief chronology of home testing for glucose
Hermann von Fehling (1849)
Urine testing using, for example, Clinitest Reagent Tablets (1941)
followed by visually read paper test strips for urine (1956)
Visually read paper strip for blood glucose (1964)
Instrument to measure paper strip by reflectance of light (1969)
First electrochemical home blood glucose monitor (1987)
First self-use continuous glucose monitor (2005)
Ames Reflectance Meter
Tom Clemens work led to the Ames
Reflectance Meter. Ames was a
division of Miles and is now part of
Bayer. Work started in 1966, four
years after Clark’s description of the
glucose biosensor, but development of
the reflectometer was much faster. A
U.S. patent (no. 3,604,815) was
granted on September 14, 1971,
about two years after it went on the
market. The original Meter was
expensive, large and heavy, (approx 1
Kg) and required a prescription.
Despite this, it was a success and
eventually led to the Eyetone, then to
the Ames Glucometer and eventually
to the great variety of other products. 12
Yellow Springs Instrument Company Inc (YSI)
The original YSI serumglucose biosensor for
diabetes
clinics 1975
Glucose
Biosensor
1975
YSI, Ohio 1987
Enzyme Electrode Reactions
GoX: Glucose + O2 = Gluconic acid + H2O2
+700mV:
H2O2 = O2 + 2H+ + e-
14
From analogue to digital
•
15
Oxford Instruments electrochemical work station with
chart recorder (circa 1980) via programmable
multichannel electrochemical analyser (1982) to penshaped instrument with disposable electrode (1987)
Biosensors: $13b Market
Share Market leaders in Glucose Biosensor Sales
2014
The ”Big Four” continue to
hold nearly 90% (88.7%) of
the market
% Sales
Nipro,
Terumo
Arkray
etc
Abbott
FreeStyle
Lite
Roche Accu-Check
Aviva Nano
Roche Accu-Check
Aviva Nano
Lifescan OneTouch
Ultra
Bayer Contour
Roche Diagnostics
Lifescan
Bayer
Abbott
2nd tier
Mediated Enzyme Electrode
Glucose oxidase or PQQ Glucose Dehydrogenase
Cass, A.E.G., Davis, G., Francis, G.D., Hill, H.A.O., Aston, W.J., Higgins, I.J., Plotkin, E.V., Scott, L.D.L. and Turner, A.P.F.
(1984) Ferrocene-mediated enzyme electrode for amperometric determination of glucose. Analytical Chemistry 56, 667-671.
17
Manufacturing by screen-printing
Biotest Medical Corp, Hong Kong
See also: https://www.youtube.com/watch?v=72IgXDMOE60 Liberty Medical video – manufacture of laser etched strips
Key Electrode Designs
Classical top-fill design
Ag/AgCl reference/ counter
electrode
to meter
SAMPLE
Working electrode:
Carbon, mediator, enzyme,
binder (e.g PEO:
polyethylene oxide) &
surfactant
Dielectric
(insulator)
CONTACTS
Conducting tracks: Silver &
Carbon ink
Substrate: e.g. Mylar™ Polyethylene
terephthalate (PET)
Capillary-fill Biosensors 1996 et sequa
Unilever, UK 1987
Kyoto Daiichi, Japan
Kyoto Daiichi,
Japan (& made
for Menarini,
Italy and Bayer
circa 1996)
1995
Key Electrode Designs
Capillary-fill design
Ag/AgCl reference/ counter
electrode
Soluble mediator
Spacer
CONTACTS
to meter
SAMPLE
Working electrode:
Carbon, enzyme, binder
(e.g PEO: polyethylene
oxide) & surfactant
Dielectric
(insulator)
Conducting tracks: Silver &
Carbon ink
Substrate: e.g. Mylar™ Polyethylene
terephthalate (PET)
Key Electrode Designs
Automation and error correction
Auto on
Sample detect
+
Fill detect
= haematocrit compensation via fill rate
Current Paradigm of Blood
Glucose Monitoring
Dispose of materials
Read test strip
Deposit blood drop
on to test strip &
insert strip
Prick finger or
arm
Load lancet into
launcher and
reassemble launcher
1-2 Minutes
Short Break
The Move to Integration
Ascensia® AUTODISC®
loaded the meter with 10
tests at a time (2003-2012)
Hypoguard ReliOn
100 test strips
In disposable meter
Accu-Check Compact –
Preloaded drum of 17 strips
Accu.Check Mobile (2012)
”Strip-free” testing (50 tests)
6 lancets hidden in drum
Beuer GL50 (2013)
3 in 1:
Lancing device
Meter
USB transfer
Towards the fully-printable instrument
26
Turner, A.P.F. (2013). Biosensors: sense and sensibility. Chemical Society Reviews 42 (8), 3184-3196.
Integrated biosensor platform
Components
● Sensors
● Display
● Printed Inter-connects
& resistors
● Battery
● Chip for measurements
LMP91000
● Chip for communication
PIC24F16KA101
● Push Button
Turner, A.P.F., Beni, V., Gifford, R., Norberg, P., Arven, P., Nilsson, D., Åhlin, J., Nordlinder, S. and
Gustafsson, G. (2014). Printed Paper- and Plastic-based Electrochemical Instruments for Biosensors.
24th Anniversary World Congress on Biosensors – Biosensors 2014, 27-30 May 2014, Melbourne,
Australia. Elsevier.
Printed layers
1. Pedot layer for the display
2. Electrolyte layer for the display
3. Carbon layer
4. Silver layer
5. Chip mounting
6. Sensor mounting
7. Battery mounting
8. Graphical over print
Turner, A.P.F., Beni, V., Gifford, R., Norberg, P., Arven, P., Nilsson, D., Åhlin, J., Nordlinder, S. and
Gustafsson, G. (2014). Printed Paper- and Plastic-based Electrochemical Instruments for Biosensors.
24th Anniversary World Congress on Biosensors – Biosensors 2014, 27-30 May 2014, Melbourne,
Australia. Elsevier.
The first demonstration
Turner, A.P.F., Beni, V., Gifford, R., Norberg, P., Arven, P., Nilsson, D., Åhlin, J., Nordlinder, S. and Gustafsson, G. (2014). Printed
Paper- and Plastic-based Electrochemical Instruments for Biosensors. 24th Anniversary World Congress on Biosensors –
Biosensors
2014, 27-30 May 2014, Melbourne, Australia. Elsevier.
29
Smart Mobile Biosensors
2012 – Telcare, 1st FDA-approved
wireless-capable glucose meter, no
Bluetooth or cable. BG results to an
online database, where they can be
accessed via password-protected
website or iPhone app.
2012 - LifeScan’s OneTouch VerioIQ— automatically
alerts to unusual patterns of high or low readings approved
by FDA. MSRP US$69.99; uses OneTouch Verio Gold Test
Strips
The Case for Continuous Monitoring
The Origins of Continuous Glucose Monitoring
(CGM)
Shichiri et al. (1982) subcutaneous
enzyme electrode with peroxide-based
detection
Biostator-GCIIS (Circa 1981)
Miles Laboratories in Elkhart,
glucose-controlled insulin infusion
system
The Arrival of CGM
Medtronic
Guardian
Dexcom
STS
Abbott Freestyle
Navigator
Meter Kit
Sensors/m
FDA
approval
$1,339
$350 (10x3day)
Aug 2005
$800
$240 (4x7day)
March 2006
$960-1,040
$360-390 (6x5 day)
March 2008 (CE June 07)
Reading
Frequency
1 per 5min (2h run in)
1 per 5min (2h)
1 per min (10h run in)
Reading must be checked by finger-stick method before adjusting insulin
Sensor Augmented Pump
•
•
•
•
Real-time continuous glucose monitoring
and the insulin pump were combined into
the Sensor-Augmented Pump system
(Medtronic Diabetes, Northridge, CA) and
launched in 2007.
Pilot studies demonstrated improvements in
mean glycaemia in users of this technology.
The FDA still required that a finger-stick
blood sample be taken before acting on the
result from a continuous sensor to
administer insulin and the technically
exciting possibility of hooking up a
continuous sensor to a commerciallyavailable automated insulin infusion pump is
not permitted.
In 2012, some degree of automation was
approved (first in Europe and then USA),
allowing the Medtronic device to be used to
shut off insulin if the blood sugar drops too
low, thus reducing the risk of
hypoglycaemia.
Abbott FreeStyle® Libre
•
•
•
•
•
•
•
•
•
3 Sept 2014 — Abbott received CE Mark
Available in seven European countries from Autumn 2014
No finger pricks for calibration and checking
Worn on the back of the upper arm for up to 14 days
Measures glucose every minute in interstitial fluid
Needle (5mm long, 0.4mm wide) inserted under the skin using an applicator and
held with adhesive pad
1 Hour to equilibrate then scan sensor to get a glucose result in <1 sec
Reader holds up to 90 days of data
Each scan provides a current glucose reading, 8-hour history and the direction
glucose is heading
Nb. Previous CGM product
suffered from water ingress
into replceable battery
compartment, QA issues and
a 10 h warm up
35
www.youtube.com/watch?v=0cXwO9YBJxE
Futrex Inc 1992: Non-invasive glucose
monitoring using NIR
The U.S. Securities and Exchange
Commission charged Futrex with fraud,
claiming that the Dream Beam never worked.
36
Non-invasive Monitoring
A selection from >95 companies identified in 2007. Bold = projected in clinical trials by 2009
Glucotrack: ultrasound +
COMPANY
TECHNOLOGY
SITE
BioTex Inc, TX, USA
Near-infrared
Skin
Sensys Medical (Sensys GTS), AZ, USA
Near-infrared
Skin
Mid-infrared/microfluid
Skin
Light Touch Medical Inc, PA, USA
Raman spectroscopy
Finger
Integrity applications (GlucoTrack),
Israel
Photoacoustic spectroscopy
Ear lobe
VeraLight Inc (Scout DS), NM, USA
Fluorescence spectroscopy
Skin
Lein applied diagnostics, UK
Optical
Eye
Glucolight Corp (Sentris -100), PA, USA
Optical coherence tomography
Skin
Echo Therapeutics (Symphony tCGS, MA,
USA
Sonophoresis
Skin
Calisto Medical (Glucoband), TX, USA
Bio-Electromagnetic Resonance
Wrist
AiMedics (HypoMon), Australia
Electro-physiological
Chest skin
Biosign technologies (UFIT TEN-20),
Canada
Electro-physiological
Wrist
Cnoga Inc. (SoftTouch), Israel
Optical (cell colour distribution)
Skin
EyeSense, Germany
Bio-chemical/fluorescence
Eye (ISF)
VivoMedical, CA, USA
Sweat analysis
Skin
thermal and electromagnetic Cascade Metrix Inc, IN, USA
conductivity
GlucoLight
HypoMon:
4 electrodes;
electrophys
changes
Cnoga
“The
science
fiction you
were
speaking
about is
reality “
Non-Invasive Systems
”Tattoo” Sensor
Pendragon Pendra (2003) Impedance
CE approved in May 2003 and briefly
available on the Dutch direct-toconsumer market. A post-marketing
reliability study was performed in six
type 1 diabetes patients. Mean absolute
difference between Pendra glucose
values and values obtained through selfmonitoring of blood glucose was 52%
and a Clarke error grid showed 4.3% of
the Pendra readings in the potentially
dangerous zone E. Pendragon went
bankrupt.
Bandodkar, A. J., Jia, W.,
Yardımcı, C., Wang, X., Ramirez,
J., & Wang, J. (2015). TattooBased Noninvasive Glucose
Monitoring: A Proof-of-Concept
Study. Analytical chemistry (in
press).
Low current reverse iontophoresis
for 10 min. ” tattoo picked up the
spike in glucose levels after a
meal” and was comfortable to
wear.
Cygnus Glucowatch Biographer
(2002) – Reverse iontophoresis
Cygnus Inc. in Redwood City,
California, went out of business and
stopped manufacturing its meters. It
sold its assets to Animas Corp
(which makes insulin pumps and is
now owned by J&J) in 2004 for
US$10 million.
Contact lens & tear sensors
Smart Holograms (2006)
Chu, M.K. et al. (2011)
Talanta 83, 960-965
Fraunhofer IMS (2012)
Microfoft & Univ. Washington (2012)
LiU (2013)
Google (2014)
Novoisense
(2014)
The Evolution of Home Blood Glucose
Monitoring
1969
Past
The original
Miles
Glucometer
1987
2005
2014
Evolution of Blood Glucose Monitor products
Present
MediSense
Mediated
sensor
CGM
Medtronic
Guardian
Non-invasive
monitoring?
(Google
research)
2015
Future
Integrated systems?
Newman, J.D. and Turner, A.P.F. (2008) Historical perspective of biosensor and biochip development. In: Handbook of Biosensors
and Biochips (Eds R. Marks, D. Cullen, I. Karube, C. Lowe and H. Weetall) John Wiley & Sons. ISBN 978-0-470-01905-4
Adverse Events - Deaths
1992-2009: 100 deaths associated with glucose meters reported
Unknown cause (34)
Meter malfunction (11)
False High Results (11)
Diabetic Ketoacidosis (8)
Maltose/non-glucose interference (13)
Use on Critically Ill Patient (6)
False Low results (6)
Possible Medication Interference (5)
Renal patient (2)
Dehydration (1)
Hyperosmolar Hyperglycemia (1)
Source:
C.C. Harper
(FDA)
Feeding tube –glucose (1)
Neonatal death (1)
Ethical dilemmas
1)
Finally, a company has succeeded in building an “artificial (bionic) pancreas”, but it is expensive
and there are only enough funds to give it to half the people who need it. How do you choose who to give it
to?
2)
You have invented a new biosensor that can measure a key intermediate in the formation of
collagen. In partnership with a pharmaceutical company, you have determined two immediate applications.
Your diagnostic can stop skin aging and address a multi-billion dollar market or it can cure a rare and
depilating inherited disease affecting the cartilage and causing suffers to be confined to a wheel chair in
their early 20’s. You have raised US$100 million investment and can afford to develop only one application
through to clinical practice. Which do you choose, why and what are the possible consequences of your
choice?
3)
You have invented an entirely new biosensor that can detect a key biomarker that can
determine the most effective treatment for colorectal cancer. However, the device requires a considerable
amount of money to develop it into a useful format. Do you patent it or publish it? Explain the reasons for
your decision and detail the effects it will have on the likely availability of the device and the treatment to
patients.
4)
Biosensors are increasingly interfaced to mobile telecommunications devices such as tablets
and mobile phones. This has direct benefits to the user, but could also provide invaluable information to
national health services and researchers (so called “big data”). However, companies could also benefit from
this information to develop new products and insurance companies could use the data to distribute
healthcare costs “more fairly”. Such information could also prove valuable in criminal forensics or to
intelligence services. Who owns this data and who should be granted access to it?
Ethical dilemmas
5)
You are a gynaecologist/obstetrician and you have a new, inexpensive DNA
biosensor that can safely determine sequences associated with a wide range of diseases. How
do you advise patients who are pregnant or considering pregnancy and are at risk for giving birth
to affected children as well as gynaecology patients who, for example, may have or be
predisposed to certain types of cancer? You should be aware that genetic information has the
potential to lead to discrimination in the workplace and to affect an individual's insurability
adversely. How do you deal with this?
6)
How long should you prolong life with bioelectronic devices and what factors should
you take into consideration?
7)
If people have wealth, is it right for them to purchase extra healthcare, like over-thecounter biosensors, that may prolong their life and may not be available to poorer people in the
same or other countries?
8)
I have a biosensor that can tell you when you are going to die. Do you want to know?
What else would you like to know?
Conclusions
Conclusions
Mediated amperometric glucose biosensors continue to dominate the home
diabetes diagnostics market
•
Peroxide based amperometric glucose biosensors predominate in the
decentralised and in vivo markets
•
The sector is typified by companies pursuing high levels of integration and
sophisticated data treatment, displays and transmission
•
Implantable sensors are in the market and home-use automated systems
coupled to insulin infusion have been announced
•
Minimally invasive (non-invasive) techniques are undergoing a resurgence of
interest stimulated my the “digital health” market and high profile players such as
Google and IBM
•
New technology continues to challenge us to adopt appropriate ethical
frameworks for regulation and control
•
Web Sites
https://www.youtube.com/watch?v=72IgXDMOE60 Liberty Medical video –
manufacture of laser etched strips
www.mendosa.com/articles_testingGlucose.htm General glucose testing
www.ysilifesciences.com/ Clinical chemistry analysers
www.minimed.com/products/guardian/ Continuous subcutaneous
http://echotx.com/ Minimally invasive example
The main commercial meters:
www.accu-chek.com.au/au/products/metersystems/advantage.html
www.bayerdiabetes.com/sections/ourproducts/meters/breeze2
www.onetouch.com/home
www.abbottdiabetescare.com/index.htm
45
www.ifm.liu.se/biosensors
6.451
2013
46
Turner, A.P.F. (2013)
Biosensors: sense and sensibility. Chemical Society Reviews 42 (8), 3184-3196.
www.ifm.liu.se/biosensors