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

Methods. We performed MCPyV-FISH of formalin fixed and paraffin
embedded (FFPE) MCCs (n=62) on tissue microarrays (TMAs), determined
the hybridization patterns and correlated these with qPCR data
on the basis of a determined cut-off. MCPyV-FISH was established using
the MKL-1 cell line which harbors integrated copies of MCPyV DNA.
For MCPyV-qPCR a LT primer pair (Becker et al., 2009) was used on
whole tissue sections.
Results. MCPyV-FISH on FFPE MKL-1 cells revealed punctate signals
compatible with viral integration. The MCPyV-FISH positive MCC cores
(76%) mainly revealed two different signal patterns: a punctate pattern
(85%) which correlated with a moderate relative viral abundance
and in some areas the punctate pattern was combined with a diffuse pattern
(15%) indicating the episomal presence of the virus which is linked
to viral replication. A mixed hybridization pattern was associated with
very high qPCR values. Comparing MCPyV-FISH and qPCR data the results
highly correlated (83%) with the MCPyV positive evaluated group,
whereas the negative group showed a concordance of 93%. The mean of
the qPCR values of all MCPyV positive cores differed significantly from
the negative cores (p=0.0076). In some tumor areas of one and the same
patients the FISH signals were heterogeneous in intensity, pattern and
nuclear localization.
Conclusions. A strong correlation between MCPyV FISH and the relative
MCPyV abundance by qPCR was detected. Thus, while presence
of MCPyV can be verified by qPCR, the quality of the presence can be
visualized by MCPyV specific FISH analysis. In this regard, MCPyV
qPCR and MCPyV FISH are important complementary tools to gain
maximum biological information of the presence of MCPyV in MCC
and thus to further elucidate MCPyV related carcinogenesis.
FR-007
A novel BRAF mutation in a patient with metastatic melanoma
R . Schneider-Stock 1 , L . Heinzerling2 , E . Kämpgen2 , M . Erdmann2 , P . Keikavoussi2
, A . Agaimy1 , A . Hartmann1 , G . Schuler2 1University of Erlangen-Nuremberg, Institute of Pathology, Erlangen,
2University of Erlangen-Nuremberg, Department of Dermatology, Erlangen
Aims. BRAF mutations in melanoma have been identified as a new target
for therapy. The V600E mutation is found in 50–68% of malignant
melanomas and can be specifically treated with e.g., the BRAF inhibitor
PLX4032 now registered as Vemurafenib. We describe a patient with a
metastasized nodular melanoma on the right lumbar region which was
excised two years before but had already spread to the inguinal lymph
nodes. Subsequently, the patient developed multiple metastases of the
brain, metastases of the mediastinal, cervical, retroperitioneal, retrocrural,
mesenterial, paraaortal and interaortocaval lymph nodes, in the
liver, the stomach and the intestines. Previous therapy included lymphadenectomy,
radiation therapy, hyperthermia, radiochemotherapy
with temozolomide, sorafenib, fotemustine and paclitaxel/carboplatin.
Despite these attempts he showed progressive disease and inclusion into
a BRAF inhibitor study was considered. Thus the tumor was sent for mutation
analysis.
Methods. Formalin-fixed and paraffin-embedded tumor sample was
used for the extraction of genomic DNA. Mutational analysis was carried
out by Pyrosequencing and for confirmation by ABI capillary sequencing
of exon 15 of BRAF.
Results. In this melanoma patient a new complex mutation was found
in BRAF with base substitution of a valine residue at position 600 for
glutamic acid (GTG GAA) and deletion of codon 601 (p.V600EK601del;
c.1799_1801del3). With this mutation the patient did not qualify for the
BRAF inhibitor study. The clinical course of the patient was rapidly progressive
with various acute complications (renal insufficiency, ileus) and
the patient deceased only 2 months after analysis of the mutation.
Conclusions. The V600E mutation constitutively activates RAF/MEK
signaling, a major driver of carcinogenesis in various malignancies.
Other rather rare mutations like V600K, V600R or V600D might also
cause this constitutive activation and are discussed to be correlated with
a yet more aggressive behaviour. It is unclear whether the new mutation
described in this case can be considered for targeted BRAF inhibitor therapy.
FR-008
The aid of immunohistochemistry in differential diagnosis between
benign and malignant phenotype of difficult melanocytic
lesions
T . Papadopoulos1 1Klinikum Nürnberg, Institute of Pathology, Nürnberg
Aims. A panel of biological markers differentially expressed in common
nevi, dysplastic nevi, Spitz nevi and malignant melanomas are introduced
providing a potential tool to differentiate benign from malignant
phenotype in difficult melanocytic lesions. The panel includes Cyclin
D1, p16, p21 and p53.
Methods. Cyclin D1 is markedly expressed in radial growth phase malignant
melanomas but is negative in common nevi and in the deepest
part of vertical growth phase melanomas as well. The cell cycle inhibitor
p16 is positive in benign nevi and radial growth phase melanomas
but becomes often negative as malignant melanoma progresses to the
vertical growth phase. The cell cycle inhibitor p21 is positive in radial
growth phase melanomas and in thin vertical growth phase melanomas
but becomes negative in thick malignant melanomas. p21 is negative in
common melanocytic nevi. Both cell cycle inhibitors p16 and p21 are
upregulated in Spitz nevi and Spitzoid nevi. Finally, p53 is negative or
positive at a very low level in melanocytic nevi. Its expression increases
with tumor progression from dysplastic nevus to radial growth phase
malignant melanoma and vertical growth phase malignant melanomas.
Conclusions. This presentation demonstrates characteristic expression
patterns of the above mentioned panel that may enable pathologists to
differentiate benign from malignant melanocytic lesions even in cases
when morphology by itself may not allow a clear classification of the melanocytic
lesion.
Notiz an die Gutachter des Abstracts: Der Abstract wurde nach Rücksprache
und auf Wunsch von Prof . Meister (München) eingereicht . Vorgesehen ist ein
etwa 15- bis 20-minütiger Vortrag, quasi als Review für die Immunhistochemie
unklarer melanozytärer Läsionen .
AG Dermatopathologie und AG Zytopathologie II –
Endokrine Themen
FR-009
Epidermal growth factor receptor mutation analysis in pleural
effusions of advanced non-small cell lung cancer patients:
When only cells are available
A . Zimpfer1 , B . Schneider1 , A . Polak1 , A . Bier2 , J . Kölbel1 , J .C . Virchow2 ,
A . Erbersdobler1 1 2 University of Rostock, Institute of Pathology, Rostock, University of Rostock,
Rostock
Aims. Epidermal growth factor receptor (EGFR) mutations are associated
with an improved clinical outcome due to response to tyrosine kinase
inhibitors in patients with non-small cell lung cancer (NSCLC). The
overwhelming majority of bronchial cancer is diagnosed on often very
limited tumor material which is also requested for secondary mutational
analysis. This study assessed the feasibility of using PCR and gene-sequencing
to screen for (EGFR) mutations in pleural effusions of advanced
NSCLC patients.
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