Development of a Sensitive Phospho-p70 S6 Kinase ELISA to Quantify mTOR Proliferation Signal Inhibition

Hartmann, Bertram; He, Xuemei; Keller, Frieder; Fischereder, Michael; Guba, Markus and Schmid, Holger (April 2013): Development of a Sensitive Phospho-p70 S6 Kinase ELISA to Quantify mTOR Proliferation Signal Inhibition. In: Therapeutic Drug Monitoring, Vol. 35, No. 2: pp. 233-239

Abstract

Background: Drug blood levels can only serve as a surrogate because of the lack of information on the drug's direct pharmacological effects in the individual patient. Measurement of the mammalian target of rapamycin (mTOR) activity dependent on the phosphorylation status of p70 S6 kinase (p70 S6K) offers a practical way for monitoring pharmacodynamic drug activity, with the potential to better assess the state of immunosuppression in individual patients. Material and Methods: Here, we established a novel in vitro model system by treating Jurkat cells and peripheral blood mononuclear cells with different concentrations of sirolimus after stimulation with phorbol 12-myristate 13-acetate. Results: A dose-dependent reduction of the p70 S6K phosphorylation status was demonstrated by Western blot and a newly established enzyme-linked immunosorbent assay (ELISA). Relative phospho-p70 S6K values from ELISA and relative densities from Western blot analysis in peripheral blood mononuclear cells revealed a strong correlation (Spearman correlation coefficient r(s) = 0.7, P = 0.01). Finally, parallel assays confirmed a sirolimus dose-dependent reduction of cytokine production and cell proliferation in the in vitro model. Conclusions: Pharmacodynamic monitoring of mTOR inhibition with a p70 S6K ELISA could guide mTOR inhibitor immunosuppression therapy toward a more individualized therapy. The usage of this technique now has to be evaluated in a clinical series of patients.

• Item Type: Journal article
• Form of publication: Publisher's Version
• Keywords: pharmacodynamic monitoring; immunosuppression; mTOR inhibitor; p70 S6 kinase; sirolimus; therapeutic drug monitoring
• Faculties: Medicine
• Subjects: 600 Technology > 610 Medicine and health
• ISSN: 0163-4356
• Place of Publication: 530 WALNUT ST, PHILADELPHIA, PA 19106-3621 USA
• Alliance/National Licence: This publication is with permission of the rights owner freely accessible due to an Alliance licence and a national licence (funded by the DFG, German Research Foundation) respectively.
• Language: English