4. Cardiff chemiluminescent technology underpinning global adoption of nucleic acid-based clinical diagnostic assays.
Submitting InstitutionCardiff University
Unit of AssessmentAllied Health Professions, Dentistry, Nursing and Pharmacy
Summary Impact TypeTechnological
Research Subject Area(s)
Chemical Sciences: Analytical Chemistry
Biological Sciences: Biochemistry and Cell Biology
Medical and Health Sciences: Clinical Sciences
Summary of the impact
Cardiff University research led to second-generation chemiluminescent
technology. The invention allowed for internal amplification control in
nucleic-acid based clinical diagnostic assays for infectious disease and
produced results with greater accuracy and fulfilled previously unmet
regulatory standards. Adopted by the market leader in nucleic acid
diagnostics (a sub-licensee of Cardiff University) the Cardiff technology
is used globally in more than 60 million in vitro diagnostic tests
annually. Sales of the tests approach $500 million per year and the
sub-licensee was subsequently sold for $3.8 billion.
A family of dyes with distinct chemiluminescent profiles
Cardiff University School of Medicine staff Stuart Woodhead (L/SL/Reader
1971 to 1998 inclusive and Professor 2001-2004) and Ian Weeks (Lecturer
1985-1994 inclusive, and Professor 2009 to present) investigated from 1993
the chemiluminescent (light emitting) properties of a family of
acridinium-ester (AE) dyes. Specifically, the emissions of various AE dyes
that were distinguishable in terms of their light-emitting wavelengths
(colour), quantum yield (light emitting efficiency) and significantly the
kinetics or life time of emitted light signal, i.e. "flash" or "glow") [3.1,3.2,3.3].
The work involved a series of design-synthesis-characterisation rounds
with chemical modifications to the AEs leading to novel molecules each
possessing distinct chemiluminescent properties. It evolved to a
`reduction to practice' for the simultaneous, though independent,
quantification of the light emissions of several different AE labelled
oligonucleotide probes. The work also addressed AE dye protection during
nucleic acid hybridization reactions and the effect of background signal.
This required optimisation of the critical property of AE dye protection
within a double stranded nucleic acid helix (such as is formed when an
AE-labelled oligonucleotide probe binds to its specific nucleic acid
target) as opposed to the deactivation of the AE dye (and its light
emitting properties) in non-hybridized probes.
A number of patent families arose from the research for the use and
development of modified AE dyes as molecular labels in in-vitro
chemiluminescent-based nucleic acid diagnostic assays [3.1,3.2,3.4,3.5].
In particular the research underpinned Cardiff technology allowing for the
concurrent use of two types of AEs distinguishable by kinetic emission,
i.e. one that "flashes" and another that "glows". This innovation provided
a solution to the unmet requirement for simultaneous detection and
quantification of multiple targets, i.e. multiplex analysis, in nucleic
acid hydridization assays.
The timeliness of the research was crucial since there was a burgeoning
interest in nucleic acid diagnostics on which Cardiff's technology
ultimately impacted. Namely, a vital element in highly sensitive nucleic
acid based diagnostic tests is nucleic acid amplification, a process
providing a high level of assay sensitivity but one prone to non-specific
interferences potentially yielding incorrect results, particularly when
conducted in automated systems. For clinical approval of nucleic acid
based diagnostic tests the regulatory authorities required the
amplification process in such assays to be internally controlled. The
Cardiff technology provided a novel, patentable solution to this. For
example, it allowed the amplification of the clinical nucleic acid target
(slow light emission) to be internally controlled by including a parallel
amplification reaction for a control sequence that could be independently
monitored (fast light emission) [3.2,3.4,3.5]. The technology
also allowed simultaneous detection of multiple screening or diagnostic
targets within the same sample.
Patent landscape and research partnering
The initial research in Cardiff University was part-funded by the
University and a University spin-out company (Molecular Light Technology
Ltd; MLT) in which the University held equity and Board representation,
and in which Weeks later became a director. The later developmental
research in Cardiff University was also sponsored by GEN-PROBE Inc., a San
Diego based company and one of the market leaders in nucleic acid
diagnostic test systems. GEN-PROBE was also a sub- licensee of previous
but completely unrelated Cardiff chemiluminescent `first-generation'
technology, which, though very successful, only permitted single analyte
detection and quantitation. The crucial underpinning research of the
University described here was the discovery of the `second-generation'
"flash" or "glow" chemiluminescent technology. The role of University
researchers is evidenced by the initial granting to the University of
background patents with PIs - Weeks and Woodhead named as sole inventors [3.1]
and later [3.2,3.4,3.5] joint inventors. The commercial
partner GEN-PROBE was ultimately assigned the patents for Cardiff's
second-generation technology which formed GEN-PROBE's chemiluminescent
Dual Kinetic Assay (DKA) approach, and which is used in their in-vitro
blood screening and clinical diagnostics products (see section 4).
References to the research
In light of its commercially sensitive nature the underpinning research
was initially published as patents (Weeks and/or Woodhouse as Cardiff
University leads) with any research articles relating back to this earlier
work. Note patents underlined are referred to in sections 4 and 5.
[3.2] Nelson, N.C., Woodhead, J.S., Weeks, I. and Cheikh, A.B.
Compositions and methods for the simultaneous detection and quantification
of multiple specific nucleic acid sequences. WO9613612 (filing 1995,
published 1996) also published as US5658737 (filing 1996, published
1997), US5840873 (filing 1997, published 1998), US5756709
(filing 1996, published 1998), US5827656 (filing 1996, published
[3.3] Smith, K*., Li, Z., Yang, J.J., Weeks, I. and Woodhead,
J.S. Synthesis and properties of novel chemiluminescent biological
probes: substituted 4-(2-succinimidylcarbonylethyl)phenyl 10-methylacridinium-9-carboxylate trifluoromethane sulphonate. J. Photochem.
Photobiol. (2000) 132: 181-191. *undertook contract chemistry synthesis
for Woodhead and Weeks who are inventors on earlier patents. http://dx.doi.org/10.1016/S1010-6030(00)00209-4
Details of the impact
Nature of impact
Cardiff University's research has led to the development of in-vitro
blood screening and clinical diagnostic nucleic-acid amplification assays
that have been adopted worldwide for the detection of infectious agents.
These assays have been brought to the market by GEN-PROBE Inc. (San Diego,
USA) and offer improved detection sensitivities and importantly possess
internal control for the nucleic acid amplification ensuring earlier
disease detection and a reduced incidence of incorrect results. The
distinct contribution of Cardiff research to the products is evident in
the respective `product inserts' [5.1] which cite patents
(underlined in references [3.1] and [3.2] of
Section 3 above) upon which Cardiff University staff (Weeks and/or
Woodhead) are named inventors. The Cardiff technology has led to year on
year benefits to commerce and clinical practice.
Clinical adoption and commercial outcomes
GEN-PROBE's key nucleic acid detection assays use the Dual Kinetic Assay
(DKA) technology with more than 60 million clinical tests performed
worldwide each year. In April 2012 Hologic Inc. (a global healthcare
company seeking to strengthen its diagnostic portfolio) acquired GEN-PROBE
for $3.8 billion [5.2] with GEN-PROBE realising a doubling in
its value from 2008 (NASDQ share price of $42) to the point of acquisition
(share price $83). GEN-PROBE as a wholly-owned subsidiary of Hologic
continues to market and develop nucleic acid tests. The Hologic 2012
annual report [5.2] states "Diagnostics product sales
increased 25% in fiscal 2012 compared to fiscal 2011 primarily due to
the inclusion of GEN-PROBE, which contributed $86.7 million in revenue".
Some of the tests that have seen first-user adoption in the assessment
period are described below.
Blood screening assays: GEN-PROBE's blood screening assays
are marketed by Novartis Diagnostics under the PROCLEIX® trade mark [5.3.,5.4],
with those approved for the Tigris® platform shown in Table 1; Tigris® is
a fully automated robotic instrument enabling high-throughput sample
processing and sample containment. Worldwide more than 80 million units of
blood are donated annually with screening performed for infectious agents
at major reference laboratories. The most common screens are for HIV,
hepatitis C (HCV), hepatitis B (HCB) and West Nile Virus (WNV). The
PROCLEIX assays for these indications use the Cardiff DKA technology and
importantly have been designed and approved for use on Tigris®. Since 2008
the number of Tigris® platforms in worldwide use at major blood testing
centres has doubled from 200 units (2008) to 390 units in 2011 [5.5];
typically a single Tigris® unit is sufficient for a single testing centre.
|Table 1: PROCLEIX
||HIV-1, HCV, HBV
||HIV-1, HCV, HBV
||HIV-1, HCV, HBV,
|First approval date
on Tigris® platform
|CE marked and
in March 2007
|CE marked and
FDA approved to
testing in 2008
|CE marked in
2009 and FDA
|CE marked in 2012
|Country approvals at
2013 (new approvals
6 other (2)
26 other (9)
19 other (19 inc.
|Notes: (a) CE = marked compliant for Europe
PROCLEIX ULTRIO Plus provides improved detection sensitivity to the
The PROCLEIX assays have a ca. 32% share of the global blood
screening market (i.e. 25 million assays per year) and are used
in ca. 80% (14.5 million tests) of US blood screens [5.3,5.4,5.6].
The PROCLEIX assays yield product sales to GEN-PROBE of ca. $200 million per
year [5.3,5.6] with ca. $16 million p.a. representing "at cost
sales" of the Tigris® instrumentation platform [5.6]; in 2012
the FDA approved GEN-PROBE's low-to mid-volume throughput testing platform
(Panther®) which will use the DKA-based assays. The DKA-based assays
represent a clinical advance to blood screening services. For example, in
2011 the South African National Blood Service showed PROCLEIX ULTRIO to
identify 9744 contaminated blood donation units from 3.8 million tested,
with 447 of these positive samples not otherwise detected by existing
approaches [5.7]. In adopting PROCLEIX ULTRIO the Australian
National Blood Authority (2010) showed the risk of transmission of HBV from
blood donations was decreased by 33% relative to HBsAg serology [5.8].
Clinical diagnostic assays for sexually transmitted disease (STD):
GEN-PROBE's APTIMA family of clinical diagnostic STD assays all utilise
Cardiff technology. Placement of Tigris® platforms worldwide for APTIMA
STD diagnostics has increased 66% from 150 units (2008) to 250 units in
2011 [5.5]. APTIMA Tigris® products approved 2008 onward
- APTIMA Trichomonas vaginalis (CE marked 2010 and FDA approved
2011). The only FDA-approved nucleic acid amplification test to
specifically detect this parasite, which causes trichomoniasis, the most
common curable STD (the Center for Disease Control estimates 7.4 million
cases p.a. in USA).
- APTIMA HPV (CE marked 2008 and FDA approved 2011) which detects the
presence of one or more of the high risk 14 subtypes of human
papillomavirus (HPV) associated with cervical cancers; HPV is the
etiological agent responsible for more than 99% of all cervical cancers.
- APTIMA HPV Genotype 16, 18/45 (FDA approved 2012). Detects the
presence of the very high risk HPV genotypes 16 and 18 or 45 associated
with 80% of invasive cervical cancers.
In 2012 USA healthcare professions and USA government recommended HPV
screening every 5 years in women over 30 years. In clinical studies
involving approximately 45,000 women the APTIMA HPV assay (detecting
certain mRNAs) has consistently shown similar sensitivity and better
specificity than the most frequently used DNA-based tests. Based on a
clinical evaluation of more than 10,000 US women with normal cytology
results, the APTIMA HPV assay had 24% fewer false positives than the
leading FDA-approved DNA test [5.9].
GEN-PROBE APTIMA STD kits using the Cardiff technology hold a 49% global
market share with ca. 29 million tests p.a. [5.5] and revenue
of ca. $200 million (2008) rising to over $350 million in 2011 [5.3].
In 2013 Hologic entered into a strategic alliance with Quest Diagnostics
(the world's largest clinical diagnostics testing company) who will use
APTIMA STD assays in diagnostic services [5.10].
The Cardiff research was transformative in providing a solution to the
unmet regulatory need for the internal control of nucleic acid
amplification in high sensitivity in-vitro screening and
diagnostic assays. The technology has been adopted by the market leader in
nucleic acid diagnostics in a range of clinical assays for infectious
agents. The technology has achieved worldwide reach and underpinned
significant commercial and clinical practice uptake [5.3].
Sources to corroborate the impact
[5.1] Patents underlined (references [3.1] and [3.2]
in Section 3 above) refer to FDA approved package inserts for GEN-PROBE
products PROCLEIX Ultrio, PROCLEIX Ultrio Plus, APTIMA Combo 2
Chlamydia/Gonorrhoea, APTIMA Trichomonas vaginalis, APTIMA HPV amongst
others, e.g.: p 53. http://www.fda.gov/downloads/biologicsbloodvaccines/bloodbloodproducts/approvedproducts/licensedproductsblas/blooddonorscreening/infectiousdisease/ucm092120.pdf
[5.2] Hologic 2012 annual report showing: revenue from diagnostic
products from GEN-PROBE acquisition (p 50); GEN-PROBE acquisition for $3.8
billion (p 65). http://www.sec.gov/Archives/edgar/data/859737/000119312512482840/d393079d10k.htm#tx393079_10
[5.3] Statement from former Vice-President Business Development,
GEN-PROBE Inc. Corroboration of the distinct significance of Cardiff
discoveries to the development and regulatory approval of GEN-PROBE's
amplified nucleic acid screening and clinical diagnostic assays. Marketing
of assays and income generation from the assays.
[5.4] Novartis Diagnostics PROCLEIX Assays. Shows range of assays and
links to worldwide product approvals. Impact through PROCLEIX Assays
triangulates through this to 5.1,5.3,5.5,5.6,5.7. http://www.novartisdiagnostics.com/products/procleix-assays/index.shtml
[5.5] GEN-PROBE presentation from Carl Hull, Chairman and CEO. `An
innovative growth company in molecular diagnostics' at 30th
Annual JP Morgan Healthcare Conference. January 10th, 2012.
Slide 8-placement of Tigris systems; Slide 11 - APTIMA 49% share of
[5.6] GEN-PROBE Annual Report 2011 highlighting: key products using the
DKA technology (pp 9-13); revenue from clinical (APTIMA STD) diagnostics
of $350M in 2011 (p 46); PROCLEIX assays used in 80% of USA blood tests (p
8) and yielding sales ca. $200 million per year with ca. $16 million per
annum representing "at cost sales" of the Tigris® instrumentation platform
(p 46-50). http://www.gen-probe.com/pdfs/2011%20Annual%20Report.pdf
[5.7] Success of PROCLEIX preventing HIV and hepatitis transmission from
donated blood (South African National Blood Service, 2011). http://en.prnasia.com/pr/2011/07/25/110712511.shtml
[5.8] Adoption of ULTRIO for blood screening by the Australian National
Blood Authority (p 2). http://www.transfusion.com.au/sites/default/files/MedilinkMay10FIN.pdf
[5.9] HOLOGIC/GEN-PROBE Clinical Diagnostics webpage listing clinical
findings for its assays. Inthis link the clinical diagnostic benefits of
APTIMA HPV assay. http://www.gen-probe.com/products-services/aptima-hpv-assays
[5.10] Quest Diagnostics enter into strategic alliance with HOLOGIC to
use APTIMA STD family ofproducts in its diagnostic services. http://ir.questdiagnostics.com/phoenix.zhtml?c=82068&p=irol-newsArticle_pf&id=1827821
All documents, testimony and webpages saved as PDFs are available from
the HEI on request.