Bottlenecks in Integrated Biomarkers Development

By Dr. Pierre Braquet, Chief Scientific Advisor, Eurofins Medinet

According to the FDA pharmacogenomics guidance, biomarkers can be classified as: (i) Exploratory, (ii) Probable Valid and (iii) Valid. For an Exploratory biomarker, a significant gap exists between scientific proposals and consensus, and consequently it cannot be accepted both for drug development purposes and for review by regulatory Agencies.  The difference between a Probable Valid biomarker and a Valid biomarker only stays in the broad consensus in the medical or scientific community about the physiologic, toxicologic, pharmacologic, or clinical significance of the results. Therefore, although huge numbers of exploratory biomarkers have been identified, few have been validated. The limited number of validated biomarker is not surprising given the technical, financial, regulatory, and social challenges linked to the discovery, development, validation, and integration of biomarker assays into clinical practice.

Bottlenecks in the Development Process

The principal bottleneck in biomarker development stands in the validation process.

Indeed, in the discovery step high-output technologies for “omics”, including powerful software, are available as biochips, 2D-gel electrophoresis, LC-MS MS (linear ion traps, MALDI-TOF-TOF, FT-ICR-MS). In contrast, validation and interpretation in a clinical context are based only on low-output methodologies/technologies.

In addition, biomarkers have diverse molecular structures ranging from simple electrolytes and metabolites to large proteins; a single biomarker can (i) exist in several forms including different amino acid residues, (ii) be present in various protein-bound states (e.g. PSA, testosterone…): as a result, it is generally not possible to validate a biomarker analytical methodology according to strict GLP procedures, at least until the clinical phase of drug development.

Finally, the validation of biomarkers continues to be settled by debate and consensus processes that take years and years. There is an urgent need for an accelerated approval of biomarkers in drug R&D by drug Agencies.

Early Biomarker Development – Asking the hard questions

There many questions that must be answered in the early steps of biomarker development:

• Do the exploratory biomarkers identified have the potential to become probable valid biomarkers?
• Do they have a link to the disease state?
• Are they specific to the disease?
• Are fluctuations related to a biological and clinical significance?
• Is a circadian rhythm expected for these exploratory biomarkers?
• Have these exploratory biomarkers the potential to be useful targets for drugs?
• Can they be modulated with the drug therapy?

In addition, technical issues must be solved:

• Are they easily accessible for sampling (matrix problem)?
• Is the stability compatible with transportation conditions in clinical trials (logistical problems)?
• Are the results reproducible?

Key Considerations

In order to solve the bottleneck problem, consider the following:

• it is mandatory to do a first selection of potential candidates based on the answers to the above questions.
• Better screening technologies should be used in order to broaden the bottleneck.
• It is important to translate the data of preclinical development into clinical trials.

Planning for the Future

Because of the present complexity of the situation, several years will be necessary before seeing new biomarkers used in clinical trials. Nevertheless, some practical measures can already be taken: 

• To select an easily accessible matrix such as plasma, urine, whole blood or saliva; since biomarker concentration is generally lower in the plasma than in the site of production, and if the assay method is sensitive enough, it is possible to select the site production for the determination of biomarkers (e.g. BAL, sputum in asthma studies for IL-5, RANTES)
• Since analyses are centralized in clinical trials, the conditions of transportation for the potential biomarkers as well as their stability over the time in different temperature conditions must be perfectly defined
• In order to accelerate the process for validation of Probable Valid biomarker, multiplex platforms (e.g. Luminex beads, cytokine arrays) which allow the determination of multiple analytes in a single reaction, must be used. This allows the definition of different profiles of patients permitting a better targeting of drugs.

As examples, some panels which could be used in clinical trials of new anti-atherosclerotic drugs will be presented:

• Biomarkers of vascular inflammation [including cytokines (IL-6, IL-10, IL-12, IL-18), cell adhesion molecules (sCD40, ICAM, VCAM)];
• Biomarkers of vascular remodeling [including metalloproteinases (e.g. MMP-9), markers of calcification (e.g. osteopontin)];
• Markers of vascular infection (e.g. Chlamydia pneumoniae).

About the Author

Dr. Pierre Braquet is a D Pharm, DSc in organic chemistry (computerization of conformers in asymmetric synthesis of amino acids). After a post-doctoral work at the Collège de France in Paris he was assigned to the St Anne Armed Forces Hospital in Toulon where he set up the first computerized management system for pharmaceutical services. In 1974 Dr. Pierre Braquet was appointed Assistant Head of IT Services at the Defense Ministry then Head of the same department in 1976. He then received an engineer degree in operational research. In 1979 he entered the pharmaceutical industry as Assistant to the President of FOURNIER Laboratories in Dijon, then in 1980 he joined MERCK SHARP & DOHME (MSD) in Paris, first as Project Director, then as Director of Development Programs. During his time at MSD he provided in 1979 the first demonstration of the free radical trapping effects of flavonoids including extracts of bilberries, grapes and blackcurrant. In 1982, Dr. Pierre Braquet joined the IPSEN BEAUFOUR Group as Research Director. In 1983 he identified a tetrahydrofuropyridin compound “cicletanine” as an antihypertensive drug able to induce prostacyclin secretion. The drug has been marketed as Tenstaten® two years later. In 1984 Dr. Pierre Braquet was promoted general manager in charge of R&D. At this time he discovered the first antagonists of Platelet Activating Factor (PAF) from an extract of the Chinese tree Ginkgo biloba. Dr. Braquet discovery triggered a worldwide effort in PAF antagonist research by pharmaceutical companies. Following this discovery, Dr. Braquet received major scientific awards such as the Prix Galien for pharmaceutical research in 1988, the EBRA Prize from the Royal Society of London the same year, the Order of Scientific Merit from Brazil in 1989, the silver medal of the City of Paris, and several other awards in Europe and America. Dr. Pierre Braquet also directed research works related to the development of the sustained release forms of somatulin, an octapeptide analog of somatostatin and of an analog of GnRH, triptorelin. Triptorelin has been marketed in the late 80’s and is used in man to treat metastatic prostate cancer and in woman to treat endometriosis and uterine fibroma (Decapeptyl®). Other research programs have been undertaken between 1985 and 1995, among them Dr. Pierre Braquet and his group conducted major research on Endothelium Dependent Relaxing Factor (EDRF, nitric oxide), on atrial natriuretic factor and on endothelin. His scientific work has resulted in more than 50 patents and about 850 publications. Some of his research works have been cited in Current Contents as "hot papers" e.g. (i) Perspectives in Platelet Activating factor Research, Pharmacological Reviews 39: 97-145, 1987, (ii) Auguet, Delaflotte, Chabrier, Pirotzky, Clostre, Braquet, Biochem,. Biophys. Res. Commun. 156: 189-192, 1988. Dr. Pierre Braquet and his group have been ranked world number 6 "top scientific producer of 1991" for their scientific production (Sciences, Vol 259, January 8, 1993).  In September 2001, Dr. Pierre Braquet was selected by the American Society for Information Science & Technology as one of the most quoted authors in Pharmacology during the past two decades. Dr. Pierre Braquet was promoted Chevalier of the “Ordre National du Mérite” by the French Ministry of Research and became a correspondent member of the Royal Academy of Sciences of Spain. Dr Pierre Braquet is visiting professor in several universities of Americas, Europe and Asia.  In 1991, Drs. Pierre and Monique Braquet founded “Bio-Inova”, a company offering central laboratory services dedicated to clinical trials. After a rapid growth, the company established laboratories in North America (Canada), Turkey (Ankara) and set operations in China. In 2003 and in order to be present on the US market, Bio-Inova merged with Focus Technologies, one of the first privately owned microbiology laboratory in Washington D.C. and Cypress CA. In 2006 Focus Bio-Inova joined the Eurofins group. Since 15 years, the company always focused on the development of new biomarkers allowing to follow the effects of drugs in clinical trials.

About Eurofins Medinet

Eurofins Medinet has been serving the pharmaceutical industry with a full central laboratory service package for over 20 years.  We are proud of our unsurpassed quality standards in laboratory testing, data management, logistics services, and project management. We also provide comprehensive support in genomic testing, bioanalytical services, and anti-infective services to support clinical trials.  For more information, please visit www.eurofinsmedinet.com.