PMID- 22213594
OWN - NLM
STAT- MEDLINE
DCOM- 20120831
LR  - 20211203
IS  - 1552-4957 (Electronic)
IS  - 1552-4949 (Linking)
VI  - 82
IP  - 3
DP  - 2012 May
TI  - Assay validation of phosphorylated S6 ribosomal protein for a pharmacodynamic 
      monitoring of mTOR-inhibitors in peripheral human blood.
PG  - 151-7
LID - 10.1002/cyto.b.21005 [doi]
AB  - BACKGROUND: Therapeutic drug monitoring (TDM) of immunosuppressive drugs after 
      organ transplantation is based on measuring blood levels alone, which often 
      results in under- or over-immunosuppression. Previous studies have shown the 
      potential of measuring pharmacodynamic drug effects for TDM, but assessment of 
      biomarkers for individual drugs is still not clinical routine. Therefore, we 
      validated a specific assay to measure the pharmacodynamic effects of mammalian 
      target of rapamycin (mTOR)-inhibitors on phosphorylated S6 ribosomal protein 
      (p-S6RP), a downstream target of mTOR. METHODS: Clinical relevant concentrations 
      of sirolimus (SRL, 0.9-91.4 mug/L), cyclosporine A (CsA, 75.1-1202 mug/L), 
      mycophenolate acid (MPA, 0.08-3.2 mg/L), or dexamethasone (DEX, 0.5-200 ng/mL) 
      were added to whole-blood from healthy volunteers. Activated whole-blood was 
      analyzed by phospho-flow cytometry to measure p-S6RP in T cells. RESULTS: 
      Phospho-flow analysis revealed that SRL suppressed p-S6RP in human T cells in a 
      dose-dependent manner with a half-maximal inhibitory concentration (IC(50)) at 
      19.8 nM and a maximal inhibitory effect (I(max) %) at 91.9%. Neither CsA, MPA, 
      nor DEX inhibited mTOR-related S6RP-phosphorylation. Coefficient of variations 
      from 0.03 to 0.05, 0.12 to 0.25, and 0.14 to 0.38 for intra-, interassay, and 
      interindividual variability respectively, showed robustness of our assay. 
      Furthermore, samples can be stored at RT or 4 degrees C up to 2 h after withdrawal. 
      CONCLUSION: We validated a robust whole-blood assay that allows the specific 
      measurement of SRL- and everolimus-induced inhibition of T cells' function 
      through detection of p-S6RP. Future studies in organ transplanted recipients will 
      show if this assay has the potential to enhance a TDM for mTOR-inhibitor drugs in 
      combination therapies.
CI  - Copyright (c) 2011 International Clinical Cytometry Society.
FAU - Dieterlen, Maja-Theresa
AU  - Dieterlen MT
AD  - Department of Cardiac Surgery, University of Leipzig, Heart Center Leipzig, 
      Leipzig, Germany.
FAU - Bittner, Hartmuth B
AU  - Bittner HB
FAU - Klein, Sara
AU  - Klein S
FAU - von Salisch, Sandy
AU  - von Salisch S
FAU - Mittag, Anja
AU  - Mittag A
FAU - Tarnok, Attila
AU  - Tarnok A
FAU - Dhein, Stefan
AU  - Dhein S
FAU - Mohr, Friedrich W
AU  - Mohr FW
FAU - Barten, Markus J
AU  - Barten MJ
LA  - eng
PT  - Journal Article
PT  - Validation Study
DEP - 20111223
PL  - United States
TA  - Cytometry B Clin Cytom
JT  - Cytometry. Part B, Clinical cytometry
JID - 101235690
RN  - 0 (Immunosuppressive Agents)
RN  - 0 (Ribosomal Protein S6)
RN  - 7S5I7G3JQL (Dexamethasone)
RN  - 83HN0GTJ6D (Cyclosporine)
RN  - EC 2.7.1.1 (MTOR protein, human)
RN  - EC 2.7.11.1 (TOR Serine-Threonine Kinases)
RN  - HU9DX48N0T (Mycophenolic Acid)
RN  - W36ZG6FT64 (Sirolimus)
SB  - IM
MH  - Cyclosporine/blood/pharmacology
MH  - Dexamethasone/blood/pharmacology
MH  - Drug Monitoring/*methods
MH  - Flow Cytometry/methods
MH  - Humans
MH  - Immunosuppressive Agents/blood/*pharmacology
MH  - Mycophenolic Acid/blood/pharmacology
MH  - Phosphorylation/drug effects
MH  - Ribosomal Protein S6/*blood
MH  - Sirolimus/blood/pharmacology
MH  - T-Lymphocytes/*drug effects/*metabolism
MH  - TOR Serine-Threonine Kinases/*antagonists & inhibitors
EDAT- 2012/01/04 06:00
MHDA- 2012/09/01 06:00
CRDT- 2012/01/04 06:00
PHST- 2011/07/27 00:00 [received]
PHST- 2011/11/24 00:00 [revised]
PHST- 2011/11/30 00:00 [accepted]
PHST- 2012/01/04 06:00 [entrez]
PHST- 2012/01/04 06:00 [pubmed]
PHST- 2012/09/01 06:00 [medline]
AID - 10.1002/cyto.b.21005 [doi]
PST - ppublish
SO  - Cytometry B Clin Cytom. 2012 May;82(3):151-7. doi: 10.1002/cyto.b.21005. Epub 
      2011 Dec 23.