Research Project 8

Splicing and epigenomic analyses in high-risk CLL for the development of novel therapeutic strategies

Objectives

 Over the last years more and more effective therapies have been developed for the treatment of  CLL patients. Nevertheless, therapy resistance and disease progression are still inevitable and most patients die from their disease or from therapy-related complications. There are a number of genomic features associated with adverse prognostic impact, such as del(17p), TP53mut, SF3B1mut, ATMmut, RPS15mut, NOTCH1mut, KRASmut and IGHV mutation status. Particularly SF3B1 and TP53 mutations are associated with poor clinical outcome. Furthermore, both - widespread changes in epigenetic patterning and alterative splicing events - are frequently found in CLL and strong evidence exists that they are associated with outcome of CLL patients. Some of these alterations have been further developed and have been shown to function as epigenetic prognostic biomarkers such as SATa regions and promoters of BCL2, MDR1, TCL1, hTERT and TWIST2 to name a few. Epigenetic signatures can even be used to subclassify IGHV-mutated patients with poor prognosis.

 Recently we have identified the epigenetic reader BRD4 (bromodomain protein 4) as a regulator of alternative splicing under heat shock. By binding to the heat shock factor 1 (HSF1), a central heat shock regulating factor, BRD4 is recruited to nuclear stress bodies and co-transriptional splicing is maintained. On the other side, it has been shown that knockdown of HSF1 or its chemical inhibition with triptolide in CLL leads to a partial depletion of downstream HSP90 clients including Bruton’s tyrosine kinase (BTK), c-RAF and cyclin-dependent kinase 4 (CDK4) and results in increased survival in Rag2-/-Il2Ryc-/- mice. Since we have shown that BRD4 localizes to nSB where HSF1 is localized and since the organization and recruitment of splicing factors to nSBs significantly contributes to the adjustment of the splicing program to stress conditions, splicing regulation over BRD4 might function synergistically with SF3B1. If this is the case this will open new therapeutic opportunities and will influence the selection of CLL patients based on genomic and epigenomic characteristics.

 Thus, the integration of epigenetic modifications and splicing events in the background of CLL-related genomic alterations are in the focus of our project. Investigations will be performed on CLL patients’ material as well as on specific mouse models which closely mirror the CLL pathogenesis. We will rely on state-of the art next generation technologies, already established in our laboratory, and combine it with focused functional investigations of SF3B1. This will guide us to an understanding of epigenetic directed splicing processes in refractory high-risk CLL. Achieved insights will also be explored for their usability as epigenetic biomarkers and investigated in selected mouse models as new therapeutic options for advanced CLL.