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Title: Characterization of rare cell phenotypes in peripheral blood and their utilisation as a surrogate tissue in pharmacodynamic biomarker assays
Authors: Uitrakul, Suriyon
Issue Date: 2019
Publisher: Newcastle University
Abstract: Rare peripheral blood cells are defined as white blood cells (WBC) at a population frequency of less than 1 in 1,000. Because blood is a relatively non-invasive and accessible sample, it has the potential to be exploited as a surrogate tissue for biomarker development. This PhD project therefore focused on the development and validation of pharmacodynamic biomarkers in rare WBCs using flow cytometry techniques. These biomarkers were developed and utilised for use in three clinical studies, the LY3143921 phase I trial, the SIOP Ependymoma II trial and the PROSPECT-NE observational study. An assay focusing on phosphorylated MCM2 (pMCM2) and ki-67 staining of peripheral blood cells was developed for an early phase clinical trial of LY3143921, a novel CDC7 inhibitor. These proteins were tested in vitro, ex vivo and in vivo. The results indicated a significant association between the number of ki-67 positive cells as a percentage of pMCM2 positive WBCs and drug exposure. Furthermore, an appropriate method was successfully developed to preserve pMCM2 and ki-67 signal in samples being transported from national clinical centres, to facilitate its incorporation in the clinical trial. An assay for the detection of acetylated histone H4 (acH4) was developed for patients receiving valproic acid, a HDAC inhibitor, as a part of the SIOP Ependymoma II trial. The applicability of the assay was tested in vitro, ex vivo and in patient samples. AcH4 signal was associated with drug exposure in vitro and ex vivo but was not associated with dose or concentration of valproic acid WBCs in patient samples. This finding was potentially related to the relatively low plasma drug concentrations observed at the dose levels studied. Additional patients with higher drug exposures need to be recruited to future studies to further investigate this issue. The protein of interest for the PROSPECT-NE observational study was ki-67, which was detected using the same assay as developed for the LY3143921 trial. This study focussed on the prognostic utility of ki-67, rather than its potential predictive or mechanistic properties. The results suggested significant differences in overall survival between patients with high and low ki-67 positive WBCs. There was no association observed between ki-67 positive WBCs and plasma ki-67 in this study. In conclusion, pMCM2 positivity in ki-67 positive cells was identified as a potentially useful pharmacodynamic biomarker, with applicability to an ongoing clinical trial. For acH4 positive cells, their utility as a pharmacodynamic biomarker remains questionable and requires further investigation. Lastly, a high proportion of ki-67 positive WBCs was found to be associated with decreased overall survival in people with cancer recruited to a Phase I clinical trial. These findings highlight the potential utility of peripheral rare WBCs as pharmacodynamic and prognostic biomarkers for clinical studies in an oncology setting.
Description: PhD Thesis
Appears in Collections:Northern Institute for Cancer Research

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