Investigation of BRD proteins and BET-inhibitors in tumor intrinsic killing and regulation of the tumor microenvironment

Abstract

The epigenetic code is modulated through writers, readers and erasers, which add, interpret and remove post-translational modifications on DNA and histones, respectively. However, while this process is essential to regulate gene expression and adapt to environmental influences, it is also exploited by cancer cells to enhance expression of oncogenes or silence transcription of tumor suppressors. BET proteins are epigenetic readers, which bind to acetylated lysine residues on histones and modulate transcription of target genes including the oncogene MYC. BET-inhibitors displace BET proteins from the chromatin and thereby suppress transcription of target genes. However, the complete catalogue of BET targets is still unknown and the function of BET-inhibitors especially on immune cells is under high debate showing immune suppressive and activating capabilities. In this thesis, the effect of the small molecule BET-inhibitor RG6146 was elucidated by assessing potential resistance mechanisms of Multiple Myeloma cell lines in vitro. Different hypothesis including overexpression of MYC, modulation of proteins of the intrinsic apoptosis pathway and presence of the export transporter ABCB1 were assessed in sensitive and resistant cell lines. While the effect of BET-inhibitors on cancer cells has been the main focus of research studies thus far, only little is known on the function of BET-inhibitors on immune cells. Therefore, one part of this research also focused on discovering the effect of RG6146 on CD4+ and CD8+ T cell activation using various in vitro assays. While T cell proliferation was strongly suppressed at early time points, longer treatment with RG6146 even enhanced cell division. The main part of this thesis describes the effect of BET-inhibitors in the interface between immune and cancer cells. RG6146 and structurally distinct BET-inhibitors were shown to sensitize cancer cells to TNF induced cell death by modulating pro-survival proteins of the NF-kB pathway thereby enhancing cell death in combination with a T cell bispecific antibody in vitro and in vivo. This thesis in collaboration with other research works is the basis for future potential combination strategies of BET-inhibitors with small molecules or immunotherapy.