Single-cell pharmacodynamic monitoring of S6 ribosomal protein phosphorylation in AML blasts during a clinical trial combining the mTOR inhibitor sirolimus and intensive chemotherapy.
Publication/Presentation Date
3-15-2012
Abstract
PURPOSE: Integration of signal transduction inhibitors into chemotherapy regimens generally has generally not led to anticipated increases in response and survival. However, it remains unclear whether this is because of inadequate or inconsistent inhibition of target or other complex biology. The mTOR signaling pathway is frequently activated in acute myelogenous leukemia (AML) and we previously showed the safety of combining the mTOR inhibitor, sirolimus, with mitoxantrone, etoposide, and cytarabine (MEC) chemotherapy. However, we did not reliably determine the extent of mTOR inhibition on that study. Here, we sought to develop an assay that allowed us to serially quantify the activation state of mTOR kinase during therapy.
EXPERIMENTAL DESIGN: To provide evidence of mTOR kinase activation and inhibition, we applied a validated whole blood fixation/permeabilization technique for flow cytometry to serially monitor S6 ribosomal protein (S6) phosphorylation in immunophenotypically identified AML blasts.
RESULTS: With this approach, we show activation of mTOR signaling in 8 of 10 subjects' samples (80%) and conclusively show inhibition of mTOR in the majority of subjects' tumor cell during therapy. Of note, S6 phosphorylation in AML blasts is heterogeneous and, in some cases, intrinsically resistant to rapamycin at clinically achieved concentrations.
CONCLUSIONS: The methodology described is rapid and reproducible. We show the feasibility of real-time, direct pharmacodynamic monitoring by flow cytometry during clinical trials combining intensive chemotherapy and signal transduction inhibitors. This approach greatly clarifies pharmacokinetic/pharmacodynamic relationships and has broad application to preclinical and clinical testing of drugs whose direct or downstream effects disrupt PI3K/AKT/mTOR signaling.
Volume
18
Issue
6
First Page
1716
Last Page
1725
ISSN
1557-3265
Published In/Presented At
Perl, A. E., Kasner, M. T., Shank, D., Luger, S. M., & Carroll, M. (2012). Single-cell pharmacodynamic monitoring of S6 ribosomal protein phosphorylation in AML blasts during a clinical trial combining the mTOR inhibitor sirolimus and intensive chemotherapy. Clinical cancer research : an official journal of the American Association for Cancer Research, 18(6), 1716–1725. https://doi.org/10.1158/1078-0432.CCR-11-2346
Disciplines
Medicine and Health Sciences
PubMedID
22167413
Department(s)
Hematology-Medical Oncology Division
Document Type
Article