96. Jahrestagung der Deutschen Gesellschaft für Pathologie e. V ...

Abstracts
SO-014
Genome-wide mRNA expression analysis reveals massive
transcriptional deregulation of cell proliferation and mitosis at
multiple steps as a key factor for tumor progression in gastrointestinal
stromal tumors (GISTs)
F . Haller 1 , D .J . Zhang 2 , I .-M . Schaefer 3 , S . Cameron 4 , B .M . Ghadimi 4 ,
A . Agaimy 5 , S . Wiemann 6 , Ö . Sahin 2
1 Albert Ludwigs University, Institute of Pathology, Freiburg, 2 German
Cancer Research Center, Heidelberg, 3 Georg August University, Institute
of Pathology, Göttingen, 4 Georg August University, Göttingen, 5 Friedrich
Alexander University, Institute of Pathology, Erlangen, 6 German Cancer
Research Center
Aims. Gastrointestinal stromal tumors (GISTs) carry mutations in the
receptor tyrosine kinases KIT and PDGFRA, leading to ligand-independent
autophosphorylation with constitutive activation of downstream
intracellular signalling cascades and accelerated cell proliferation. Next
to genotype, anatomical localisation and mitotic counts are two clinicopathological
parameters with significant impact on clinical behavior
in GISTs. The aim of the current study was to determine the effect of
genotype, anatomical localisation and mitotic counts on global mRNA
expression.
Methods. Genome-wide mRNA expression analyses were performed
using Sentrix HumanWG-6 arrays (Illumina, San Diego, CA) in a series
of 20 GISTs with either KIT or PDGFRA mutation from different anatomical
localisations, and with low and high mitotic counts.
Results. Using two-dimensional principal component analysis, tumors
were clearly separated according to genotype and anatomical localisation,
with clustering of tumors from the stomach, duodenum, jejunum/
ileum and rectum, respectively. Moreover, tumors with high mitotic
counts were separated from tumors with low mitotic counts. Group-wise
comparison of gene expression levels revealed significant upregulation
of 269 genes in GISTs with high mitotic counts, compared to 88 genes
that were downregulated. Further functional enrichment analysis using
this signature revealed a significant enrichment of genes allocated to 38
GO terms associated with cell proliferation and mitosis.
Conclusions. Deregulation of several key players of cell cycle regulation at
different steps of the cell cycle contributes to increased cell proliferation
and tumor progression in GISTs, and determination of their expression
may improve prognostication of clinical behavior.
SO-015
SRC signalling is crucial in the growth of synovial sarcoma cells
S . Michels1 , E . Sievers1 , M . Trautmann1 , D . Kindler1 , S . Huss1 , M . Renner2 ,
R . Penzel2 , O . Larsson3 , A . Kawai4 , S . Tanaka5 , P . Schirmacher2 , G . Mechtersheimer2
, E . Wardelmann1 , R . Büttner1 , W . Hartmann1 1 2 University Hospital Cologne, Institute of Pathology, Köln, University
Hospital Heidelberg, Institute of Pathology, Heidelberg, 3The Karolinska
Institute, Department of Oncology & Pathology, Stockholm, Sweden, 4Natio nal Cancer Center Hospital, Division of Orthopaedic Surgery, Tokyo, Japan,
5Hokkaido University Graduate School of Medicine, Laboratory of Molecular
& Cellular Pathology, Sapporo, Japan
Aims. Synovial sarcoma is a malignant soft tissue tumor, which affects
mainly adolescents and young adults. It is molecularly characterized
by a reciprocal balanced t(X;18) translocation, resulting in a chimeric
transcriptional modifier. Several receptor tyrosine kinases including
the IGF-IR and the EGFR have been shown to be expressed in synovial
sarcomas, leading to an activation of common intracellular kinase signalling
cascades. The SRC tyrosine kinase is an important interaction
partner for different growth factor receptors and effectors of intracellular
kinase signalling pathways and has been shown to be of particular
importance in a variety of tumors. This study was performed to examine
64 | Der Pathologe · Supplement 1 · 2012
the functional relevance SRC in synovial sarcomas and to evaluate if it
might represent a target for innovative therapeutic approaches.
Methods. Immunohistochemical analyses of differentially phosphorylated
SRC and the SRC regulators CSK and PTP1B were performed in a set
of 30 synovial sarcomas. Functional aspects of SRC signals in synovial
sarcomas and dependence of SRC activation on the SS18/SSX fusion proteins
were subsequently analyzed in vitro.
Results. Activated p-(Tyr416)-SRC was detected in the majority of synovial
sarcomas; deregulation of CSK and PTP1B could be excluded to be
the reason for the activation of the kinase. In a T-Rex293-based in vitro
model, expression of the SS18/SSX fusion proteins was associated with
increased p-(Tyr416)-SRC levels, at least partially due to an induction of
the Insulin-like growth factor signalling pathway. Four human synovial
sarcoma cell lines treated with the SRC inhibitor dasatinib displayed a
significant and dose-dependent inhibition of cellular growth in MTT assays,
which was accompanied by decreased phosphorylation of the SRC
targets FAK, STAT3, IGF-IR and AKT. In flow cytometric analyses, the
growth effects exerted by the inhibitor were shown to be due to a reduction
of cellular proliferation combined with an increase of apoptosis.
Concurrent exposure of synovial sarcoma cells to dasatinib and conventional
chemotherapeutic agents resulted in positive interactions. Finally,
synovial sarcoma cell migration and invasion was found to be dependent
on signals transmitted by SRC.
Conclusions. In summary, our data show that the SRC kinase might represent
a promising therapeutic target in synovial sarcomas.
SO-016
Targeting endometrial stromal sarcoma: histone deacetylase and
PI3K/Akt/mTOR signaling
P . Quan1 , E . Lederer1 , I . halbwedl1 , H . Denk1 , K . Zatloukal1 , J . Haybaeck1 1Med . Uni . Graz/Department of Pathology, Graz, Austria
Aims. Endometrial stromal sarcoma (ESS), derived from mesenchymal
cells, is a rare gynecological malignancy with an unclear molecular pathogenesis
and thus few therapeutic options. Previously, histone deacetylase
(HDAC) 2 expression was shown to be upregulated in human ESS
specimens. The HDAC inhibitor SAHA reduced in vitro growth of the
ESS cell line ESS-1 through the inhibition of mTOR signaling. The PI3K/
Akt/mTOR signaling, central to translational regulation and vital to
the growth and survival of cancer cells is an important target in cancer
therapy. This study aims at investigating (1) if HDAC and the PI3K/Akt/
mTOR signaling are involved in ESS pathogenesis and (2) how altered
HDAC levels regulate PI3K/Akt/mTOR signaling and its downstream
translation regulators in ESS.
Methods. The expression levels of HDAC1 and 2 in ESS were examined by
using a tissue microarray. The mRNA and protein levels of HDAC1 and 2
were determined by Q-PCR and Western blotting in ESS cell lines (ESS-1
and MESSA) and the appropriate control endometrial stromal cell line
HESC. The role of HDAC in regulating the PI3K signaling was checked
in cells treated with SAHA.
Results. 1.) HDAC1 and 2 are overexpressed in ESS tissues, compared to
normal endometrium. 2.) Higher levels of HDAC1 and 2, increased cell
growth, upregulated Akt and mTOR activation were detected in ESS cell
lines, relative to HESC. 3.) SAHA inhibited cell growth of three cell lines.
However, HESC is less sensitive to SAHA with a higher IC50 value than
other cells. 4.) SAHA reduced phosphorylated 4EBP1 and BCL-2 protein
levels in all cell lines. 5.) SAHA dose-dependently inhibited activation of
Akt and p70S6k in ESS-1, but not in MESSA and HESC cells.
Conclusions. HDACs and the PI3K signaling are involved in the ESS
pathogenesis. Despite of different drug sensitivity and response rates
among all cell lines, SAHA reduced cell growth via the PI3K/Akt/mTOR
signaling and its downstream effectors 4EBP1 and p70S6K, indicating
an integration of HDACs with the PI3K signaling and translation regulation.
This connection offers a promise for a combination therapy, i.e.
SAHA with various inhibitors for the PI3K signaling, which might be