Optimization of analytical and pre-analytical variables associated with an ex vivo cytokine secretion assay

Ray, Chad; Dumaual, Carmen M.; Willey, Mark B.; Fill, Jeffrey; O’Brien, Peter J.; Gourley, Ian; Devanarayan, Viswanath; Konrad, Robert J.

doi:10.1016/j.jpba.2005.10.013

Cited by 15 publications

(13 citation statements)

References 16 publications

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“…Processed plasma or serum should be frozen at −80°C as soon as possible in small aliquots to avoid repeated freeze-thaw cycles [107, 122]. Some reports proposed to keep samples refrigerated at 4–8°C (but not on ice) after clotting for the duration of processing as room temperature favours proinflammatory cytokine degradation such as IL-6 but conversely stabilises TNF-alpha [26, 34, 120, 123, 124]. Most cytokines are relatively stable with the well-known exception of TNF-alpha and IL-6 [42, 125, 126]; therefore, the interval between blood draw and separation should not exceed 3–24 hours, even when the tubes are stored at 4–8°C and only when EDTA tubes are used (TNF-alpha however cannot be reliably measured any longer), although many cytokines have not been sufficiently tested [26, 35, 37, 78].…”

Section: Cytokines As Biomarkersmentioning

confidence: 99%

Cytokines as Biomarkers in Rheumatoid Arthritis

Burska

Boissinot

Ponchel

2014

Mediators of Inflammation

182

139

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RA is a complex disease that develops as a series of events often referred to as disease continuum. RA would benefit from novel biomarker development for diagnosis where new biomarkers are still needed (even if progresses have been made with the inclusion of ACPA into the ACR/EULAR 2010 diagnostic criteria) and for prognostic notably in at risk of evolution patients with autoantibody-positive arthralgia. Risk biomarkers for rapid evolution or cardiovascular complications are also highly desirable. Monitoring biomarkers would be useful in predicting relapse. Finally, predictive biomarkers for therapy outcome would allow tailoring therapy to the individual. Increasing numbers of cytokines have been involved in RA pathology. Many have the potential as biomarkers in RA especially as their clinical utility is already established in other diseases and could be easily transferable to rheumatology. We will review the current knowledge's relation to cytokine used as biomarker in RA. However, given the complexity and heterogeneous nature of RA, it is unlikely that a single cytokine may provide sufficient discrimination; therefore multiple biomarker signatures may represent more realistic approach for the future of personalised medicine in RA.

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Section: Cytokines As Biomarkersmentioning

confidence: 99%

Cytokines as Biomarkers in Rheumatoid Arthritis

Burska

Boissinot

Ponchel

2014

Mediators of Inflammation

182

139

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“…(28). That is, to determine whether a biomarker method is fit-for-purpose, we should determine whether it is capable of distinguishing changes that are statistically significant based on the intra-and intersubject variation.…”

Section: In-study Validation and Sample Analysis Acceptance Criteriamentioning

confidence: 99%

Fit-for-Purpose Method Development and Validation for Successful Biomarker Measurement

et al. 2006

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Despite major advances in modern drug discovery and development, the number of new drug approvals has not kept pace with the increased cost of their development. Increasingly, innovative uses of biomarkers are employed in an attempt to speed new drugs to market. Still, widespread adoption of biomarkers is impeded by limited experience interpreting biomarker data and an unclear regulatory climate. Key differences preclude the direct application of existing validation paradigms for drug analysis to biomarker research. Following the AAPS 2003 Biomarker Workshop (J. W. Lee, R. S. Weiner, J. M. Sailstad, et al. Method validation and measurement of biomarkers in nonclinical and clinical samples in drug development. A conference report. Pharm Res 22:499-511, 2005), these and other critical issues were addressed. A practical, iterative, "fit-for-purpose" approach to biomarker method development and validation is proposed, keeping in mind the intended use of the data and the attendant regulatory requirements associated with that use. Sample analysis within this context of fit-for-purpose method development and validation are well suited for successful biomarker implementation, allowing increased use of biomarkers in drug development.

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“…Corticosteroids and phosphodiesterase 4 (PDE4) inhibitors are well-known inhibitors of TNF-α production in the human whole blood assay [11,15]. For this reason, we tested the activity of dexamethasone and roflumilast, a selective PDE4 inhibitor, in the rat whole blood assay.…”

Section: Dexamethasone and Roflumilast Inhibit Tnf-α Secretion In Ratmentioning

confidence: 99%

“…Blood is incubated with a stimulus which activates cytokine production from leukocytes, in the presence of various compound concentrations. The main advantage is the ease of evaluating a simple response in a complex system, where several types of cells and proteins coexist and interact with each other in a conserved physiological environment [11,12]. A wellcharacterized assay consists of lipopolysaccharide (LPS)-stimulated human blood leading to TNF-α secretion from monocytes [13].…”

Section: Introductionmentioning

confidence: 99%

Differential Pharmacological Behaviour of p38 Inhibitors in Regulating the LPS-Induced TNF-α Production in Human and Rat Whole Blood In Vitro

Pont-Giralt¹,

Godessart²,

Balagué³

2010

Inflammation

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p38 inhibitors are potent TNF-α suppressors in LPS-stimulated human whole blood and promote efficacy in the rat adjuvant arthritis model. However, the anti-TNF-α activity of p38 inhibitors in rat whole blood has not been explored, preventing the establishment of a potential relation between in vitro and in vivo activity data in the same species. We have pharmacologically characterized a rat whole blood assay based on LPS stimulation. While p38 inhibitors showed good activity in the human assay, they failed to inhibit TNF-α in the rat whole blood assay. At high LPS concentration some compounds even potentiated TNF-α production in the rat assay, which could be reverted in the presence of the ERK pathway inhibitor U0126. Our results suggest that p38 contributes directly to TNF-α production in human whole blood while playing a negative regulatory role in rat blood which can be overridden by p38 inhibition in the presence of high stimulus concentration.

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Optimization of analytical and pre-analytical variables associated with an ex vivo cytokine secretion assay

Cited by 15 publications

References 16 publications

Cytokines as Biomarkers in Rheumatoid Arthritis

Cytokines as Biomarkers in Rheumatoid Arthritis

Fit-for-Purpose Method Development and Validation for Successful Biomarker Measurement

Differential Pharmacological Behaviour of p38 Inhibitors in Regulating the LPS-Induced TNF-α Production in Human and Rat Whole Blood In Vitro

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