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

Abstracts
Methods. Three ALK + ALCL cell lines were transduced with C/EBPβ
shRNA by lentiviral gene transfer. The C/EBPβ knockdown was quantified
by RT-qPCR and Western Blot. MiRNA expression in ALK+ ALCL
cell lines with and without C/EBPβ knockdown, ALK − ALCL cells and
T-cells were analyzed by deep-sequencing, to determine differentially
regulated and expressed miRNAs in ALK+ ALCL. The influence of C/
EBPβ on the expression of miRNA candidates and the differential expression
in ALK+ ALCL was validated by RT-qPCR in cell lines and in
primary tumors.
Results. Next-generation sequencing analysis resulted in 80 significantly
regulated miRNAs after C/EBPβ knockdown in the three ALK+ ALCL
cell lines. Three of these miRNAs (miR-181a*, miR-146b-5p, miR-203)
were significantly regulated in all three cell lines. The C/EBPβ dependent
regulation of these miRNAs was confirmed in two cell lines by RTqPCR.
Comparison of the results of the ALK+ ALCL cell line SUDHL-1
and T-cells or SUDHL-1 and the ALK− ALCL cell line revealed 379 or
301 significantly regulated miRNAs respectively. ALK− ALCL cells and
T-cells showed a significant difference in the expression levels of 366
miRNAs. A hundredfold change in expression levels was observed for
a few interesting miRNAs of the different signatures. The differential
expression of some of the most remarkable miRNAs in ALK+ ALCL
was validated in primary human ALK+ and ALK− ALCLs by RT-qPCR.
Several of these miRNAs play important roles in diverse cancers with
tumor-suppressing or oncogenic functions.
Conclusions. Three miRNAs were found to be regulated by C/EBPβ in
two ALK+ ALCL cell lines. Numerous miRNAs are differentially expressed
in ALK+ or ALK− ALCL cells and T-cells. Several miRNAs which are
significant differentially expressed in ALK+ and ALK− ALCLs separate
ALCLs depending on their ALK status. We identified a miRNA profile
specific to ALK+ ALCLs.
DO-066
The role of C/EBPβ in the phenotype of ALK+ anaplastic large cell
lymphoma
J .-A . Schmidt 1 , I . Bonzheim1 , S . Schäfer1 , F . Fend1 , L . Quintanilla-Martinez1 1University Hospital Tübingen, Institute of Pathology and Neuropathology,
Tübingen
Aims. ALK+ anaplastic large cell lymphoma is characterized by the loss
of pan T-cell antigens and the unusual expression of myelomonocytic
surface markers. The forced overexpression of the transcription factor C/
EBPβ in B and T-cells has been shown to induce transdifferentiation into
macrophages. Since C/EBPβ has been shown to play a central role in the
pathogenesis of ALK+ ALCL, the aim of the study was to investigate its
influence on the unusual phenotype of ALK+ ALCL.
Methods. The expression of 242 surface antigens was investigated in
ALK+ ALCL cell lines before and after the specific knockdown of C/
EBPβ using the BD Lyoplate Human Cell Surface Marker Screening Panel
by flow cytometry. A highly specific C/EBPβ-shRNA was transduced
by lentiviral infection. The expression of significant surface markers was
validated by immunohistochemistry and Western blot in primary ALK+
and ALK− ALCL cases.
Results. The surface marker screening confirmed the loss of T-cell specific
markers, such as TCR chains and CD3, and overexpression of EMA
and activation markers CD25 and CD30 but did not support the unusual
expression of myeloid markers as CD13 or CD33, as previously described.
Activation surface markers, including CD25, CD30, CD97 and CD98,
showed downregulation after C/EBPβ knockdown indicating a role for
C/EBPβ in the activation of ALK + ALCL cells. CD147 (EMMPRIN) was
strongly expressed in ALK + ALCL cells, and was downregulated after C/
EBPβ knockdown. Interestingly, CD147 was differentially expressed in
ALK + ALCL when compared to ALK − ALCL primary cases.
Conclusions. Surface marker screening in ALK+ ALCL revealed an influence
of C/EBPβ on the activation phenotype of the neoplastic T-cells and
expression of CD147, an inductor of matrix metalloproteinases implica-
32 | Der Pathologe · Supplement 1 · 2012
ted in tumor progression and metastasis in solid tumors. The differential
expression of CD147 in ALK + ALCL suggests its involvement in the pathogenesis
of ALK + ALCL.
AG Orthopädische Pathologie
DO-079
Histomorphological and clinical characterisation of Epstein-Barr
virus-associated post-transplant smooth muscle tumours
L . Maegel1 , J . Rische1 , C . Tiede1 , J . Salem1 , F . Laenger1 , H . Kreipe1 , K . Hussein1 ,
D . Jonigk1 1Hannover Medical School, Institute of Pathology, Hannover
Aims. Histomorphological and clinical characterisation of Epstein-Barr
virus-associated post-transplant smooth muscle tumours.
Methods. Own cohort: Five PTSMT were examined by histology,
immunohistochemistry and molecular methods [mean age of patients
10 years; PTSMT developed after a mean interval of 44 months following
liver (n=2), heart (n=2) or bone marrow (n=1) transplantation]. Metadata
analysis: Data of 64 PTSMT cases (including our cohort) of the last
20 years were re-evaluated by applying survival analysis. Molecular analyses:
All specimen of our cohort underwent compartment-specific laser
microdissection and processed for further quantitative real-time PCR
analysis. Gene expression of transcripts for a set of 20 EBV-associated
endogenous human genes were analysed by qPCR: transcription factors
MYC, TP53, NFKB1; apoptosis factors such as BAX; JAK3/STAT signal
factors; cytokines such as VEGF; miR-155 and miR-146a. To evaluate the
origin of PTSMT – either from the recipient or from the donor – short
tandem repeat (STR)-PCR was performed (“molecular fingerprinting”).
Results. Histomorphology and immunoprofile (total cohort, n=64):
– Spindle shaped, leiomyogenous cells (actin+/desmin+/EBER+), local
invasion.
– Most PTSMT show no marked cellular atypia, no tumour necrosis,