Evaluation of a New Approach of the Diagnosis of Constitutional Functional Disorders of Platelets

Toulouse University Hospital

Status: Completed

Conditions: Platelet Dysfunction

Treatments: Other: Blood punction

Study type: Interventional

Funder types: Other

Identifiers

NCT01957345

PHRCI 2012 (Other Grant/Funding Number)

12 068 08

Details and patient eligibility

About

The primary purpose of the study is to evaluate a standardized method of screening for platelet signalling defects in patients with constitutional disorders of platelet function of unknown origin. We hypothesize that such defects are under-diagnosed in patients, due to heavy workup and requirement of relatively large blood sample by conventional biochemical methods. We propose to analyse kinase signalling downstream platelet membrane receptors using multiplex flow cytometry quantification and fluorescent platelet barcoding.

Full description

Little is known of the molecular basis of disorders of signalling pathways potentially responsible of constitutional defects of platelet functions (adhesion, aggregation or secretion) (1-5). Indeed, in routine practice, this investigation is limited by the complexity of the analyse using biochemical methods (western blotting), and the requirement of large amount of platelets.

We have designed a new approach for the quantification of platelet cytoplasmic phosphoproteins by flow cytometry. Fresh platelets in platelet rich plasma are analysed at baseline or after stimulation by major agonists (ADP, TRAP, thromboxane analogue, or collagen-related peptide), with and without relevant inhibitors of each pathway. Multiplex barcoding is used to identify each condition, allowing a high throughput analysis (6, 7). Platelet signalling profiling of Akt, Slp76, P38 MAPK and LIMK can be obtained from a blood sample of less than 10 ml, and within 6h The main objective of the study is to standardize the method between clinical laboratories with a standard expertise in flow cytometry. The study will be performed in 4 academic hospitals members of the French reference network of rare platelet diseases. Three groups of patients referred for mild or severe bleeding disorders will be included: 1) a control group of patients (30 per centre) with a bleeding disorder definitely other than of platelet origin (e.g. "low" von Willebrand); 2) a group of 10 patients per centre with definite constitutional platelet disorder (e.g. Glanzmann thrombasthenia) and 3) a group of 10 patients per centre with a defect of platelet function of unknown origin, potentially defective in signalling pathway.

The control group will serve to standardize the method between centres and to establish the reference values. A quality control will be set up by using frozen platelet preparations. The patients with definite platelet disorder will be useful for detecting potential signalling defects still not described in these pathologies. Platelet signalling defects which could be evidenced in these groups will be further identified by conventional biochemical and molecular methods after confirmation on a new sample.

If this new approach can be proposed to clinical laboratories working on rare platelet diseases, we expect an advance in our knowledge in the field. In addition the method has a potential in pharmaceutical innovation, for identifying (8) or monitoring new antiplatelet agents (9, 10), or identifying platelet defects induced by new "target therapies" designed for other diseases such as cancer or immune pathologies (10).

Enrollment: 322 patients

Sex: All

Ages: 18+ years old

Volunteers: No Healthy Volunteers

Inclusion criteria

• patient consulting for hemorrhagic symptomatology with a bleeding disorder definitely other than of platelet origin (e.g. "low" von Willebrand) / or constitutional platelet disorder (e.g. Glanzmann thrombasthenia) / or with a defect of platelet function of unknown origin, potentially defective in signalling pathway.
• Informed consent form
• patient with social security insurance or equivalent

Exclusion criteria

• treatment interfering with platelets function within 7 days prior to enrollment
• thrombocytopenia <100G/L
• pregnant or lactating females
• subjects under juridical protection guardianship or tutelage measure
• subjects involved in another clinical trial

Trial design

Primary purpose: Diagnostic

Allocation: Non-Randomized

Interventional model: Parallel Assignment

Masking: None (Open label)

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322 participants in 3 patient groups

bleeding disorder

Other group

Description:

Blood punction at patients with bleeding disorder definitely other than of platelet origin (e.g. "low" von Willebrand)

Treatment:

Other: Blood punction

constitutional platelet disorder

Other group

Description:

Blood punction at patients with constitutional platelet disorder (e.g. Glanzmann thrombasthenia)

Treatment:

Other: Blood punction

defect of platelet function

Other group

Description:

Blood punction at patients with defect of platelet function of unknown origin, potentially defective in signalling pathway.

Treatment:

Other: Blood punction