Development of RAD51 as Biomarker for Sensitivity to the Histone Deacetylase (HDAC) Inhibitor PCI-24781 in Lymphoma.

Histone deacetylase inhibitors are currently under investigation in the clinic for a variety of malignancies, and one was recently approved for treating cutaneous T-cell lymphoma. We demonstrate here that a novel, orally bioavailable HDAC inhibitor currently in clinical trials, PCI-24781, demonstrates very good activity against lymphoid tumor lines in vitro as well as against primary tumor cells from lymphoma patients cultured ex vivo. Further, we demonstrate the potential of predicting sensitivity to PCI-24781 in lymphoma by measuring expression levels of RAD51, a protein essential for DNA repair by homologous recombination (HR) following exposure to DNA damaging agents. We have recently shown that RAD51 is downregulated by PCI-24781 treatment in solid tumor lines, and this leads to the inhibition of homologous recombination, absence of RAD51 foci and ultimately to apoptosis. However, little is known about the role of RAD51 in lymphoma. In this work, we show that RAD51 is expressed in human lymphoma by immunohistochemical staining of tissue microarrays (TMA) containing over 90 primary tumor sections (50 follicular lymphomas and 40 DLBCL). We found that a majority of these (65%) express significant levels of RAD51. Further, we measured RAD51 protein and mRNA levels in 8 non-Hodgkins lymphoma cell lines, including those derived from B- and T-cell lymphomas, before and after a 24 hour exposure to PCI-24781 at physiologically relevant concentrations. We found that there was a strong correlation between the percentage of apoptotic cells following PCI-24781 treatment as assayed using Annexin V/propidium iodide and the decrease in the level of RAD51 expression, with each of the five cell lines with >70% apoptosis following drug treatment also showing >4 fold decrease in RAD51 mRNA. These cell lines also had the highest pretreatment levels of RAD51. Thus, both the initial level of RAD51 in tumors as well as the fold decrease after treatment are predictive for PCI-24781 activity, and will be used as biomarkers in clinical studies to identify patients most likely to respond to PCI-24781.