The deubiquitinase CYLD removes K63-linked ubiquitin chains from molecules involved,
e.g., in NF-kB, Wnt/ß-catenin and BCL-3 signaling pathways. CYLD is dysregulated in
different cancers, including hepatocellular carcinoma (HCC). Loss of CYLD is characterized
by enhanced NF-κB-signaling and increased cell survival. In previous studies, we demonstrated
decreased levels of CYLD expression in human HCC. In recent analysis of CYLD expression
in tissue microarrays, survival analysis of HCC patients after resection revealed
a more favorable outcome of patients with positive nuclear CYLD expression. We hypothesize
that reconstitution of CYLD is a promising approach for the treatment of liver diseases
and the sensitization of HCC cells towards therapy-induced cell death. To assess the
regulation of CYLD by different treatment strategies, HCC cell lines will be treated
with chemotherapeutic agents and targeted agents. After treatment, CYLD expression
levels including subcellular localization will be measured and cell death induction
will be assessed by different analysis. Those targeted agents, which result in a decrease
or no alteration of CYLD levels will be used for treatment of HCC cells, previously
transfected with CYLD expression plasmids. Cell death induction will be assessed to
test, whether CYLD upregulation sensitizes HCC cells to the respective targeted therapy.
Another subproject intends to investigate the influence of CYLD on the migration and
invasive potential of HCC cell lines in vitro. After CYLD downregulation, cells will
be grown in 3D polystyrene scaffolds, which provide a suitable surrounding for long
time cell growth. Scaffolds will be further processed for cryosectioning followed
by immunostaining to investigate the expression levels of CYLD, Bcl-2 proteins and
markers for EMT, migration and proliferation. The invasion depth will be assessed
as well.
Corresponding author: Elssner, Christin
E-Mail:
christin.elssner@nct-heidelberg.de
