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Rare BRAF mutations distinguish anorectal from cutaneous melanoma
Posters: Colon / Pathology - Research and Practice 200 (2004) 320-325 tic for the hereditary non polyposis colorectal cancer (HNPCC)syndrome and is also found in approximately 10% of all sporadic colorectal cancers. MSI analysis is expensive, timeconsuming, and preselection of patients according to the so called "Bethesda guidefines" prior to MSI analysis is routinely performed focusing on the identification of patients with HNPCC syndrome. Now there is growing evidence that MSI appears to be of prognostic significance no matter if the DNA mismatch repair (MMR) system is silenced by genetic or epigenetic mechanisms. This will urge the need for highly efficient screening strategies. The main purpose of the present study was to establish a set of key markers that can simplify MSI genotyping for high throughput applications in order to overcome preselection of patients prior to MSI analysis. Methods: The MSI status of 74 colorectal carcinomas from patients classified as "at risk" for HNPCC syndrome upon clinical criteria, was determined using the ICG-HNPCC microsatellite marker panel. Tissue samples were additionally classified with a set of four 3'UTR mononucleotide repeats. Results: Using the Bethesda marker panel for evaluation of MSI, 61% (45/74) colorectal cancer specimens were classified as MSIH. The clinical Bethesda criteria however, were only fullfilled by 40 of the 45 MSI-H tumor patients. For two of the 5 Bethesda negative patients a hereditary component could be identified. The diagnostic sensitivity for mononucleotide repeats was the highest for the new MSI-marker (45/45) followed by BAT26 (44/45), BAT25 (44/45), PPP3CA (41/45) CTNNB1 (37/44) and GTF2E1 (37/45). The dinucleotid repeats displayed a diagnostic sensitivity less than 80%. Mean base pair deletions were the largest for BAT26 (8.6 bp, SD _+3.51), new MSI marker (7.6 bp SD _+2.87), and BAT25 (6.0 SD + 2.6). The remaining mononucleotide repeats showed markedly decreased mean basepair deletions of < 3 bp. Conclusion: We identified a monomorphic Mononucleotide Microsatellite Marker as a useful marker for the evaluation of MSI status in colorectal cancer tissue samples. A simplified testing procedure for high throughput MSI diagnosis of further cancer types is currently developed on the basis of this particular microsatelfite.
247 Rare BRAF Mutations Distinguish Anorectal from Cutaneous Melanoma
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248 Analysis of TolMike receptors 2, 4 and MD2 in inflamed and non-inflamed gut tissues in inflammatory bowel disease F. AUTSCHBACH, T. GIESE 1, N. GASSLER, I. BERGER, B. SIDO 2, G. HEUSCHEN 2, S.C. MEUEW, H.F. OTTO Pathologisches Institut, Universit~it Heidelberg *Institut fur Immunologie, Universitat Heidelberg 2Chimrgische Universitatsklinik, Universitat Heidelberg
Aims: A perturbed host response to the enteric microbial flora plays a role in the pathogenesis of chronic inflammatory bowel diseases (IBD) like Crohn's disease (CD) and ulcerative colitis (UC). In this study, we investigated the intestinal expression of Toll-like receptors (TLRs), important signaling receptors which can be activated by bacterial wall products such as fipopolysaccharide. Methods: Expression of mRNA-transcripts for TLR2, TLR 4 and the TLR-4-related coreceptor MD2 was analyzed by real-time RTPCR on a large number of transmural gut samples from actively inflamed and non-inflamed/inactive areas in IBD (n = 83) and controls (n = 27). Cells expressing TLR2 and TLR4 protein were characterized by immunohistochemistry (IH). Additional paraffin sections were immunostained for Ki-67 antigen (Mib-1) to investigate the proliferative activity of lamina propria leucocytes (LPLs). Results: Transcript levels for TLR2, TLR4 and MD2 were significantly elevated in samples with active CD and UC as compared with normal controls. IH revealed LPLs to be the predominant TLR-expressing cell type. No increased expression of TLRmRNAs was observed in noninfiamed/inactive areas in IBD. Elevated TLR-transcripts in IBD were accompanied by a significant increase in the number of Ki-67-positive proliferating LPLs. In the case of TLR4, transcript levels were significantly higher in inflamed CD as compared with UC or non-IBD-related inflammatory controls. Conclusions: Induction of TLRs and receptor signalling via bacteri-al wall products contribute to the exaggerated immune response of intestinal LPLs in actively inflamed, but not in inactive areas in IBD. TLR4 mRNA is predominantly induced in CD as compared with UC.
B.M. HELMKE, J. MOLLENHAUER l , C. HEROLD-MENDE 2'3, A. BENNER 4, M. DEICHMANN 5, H.F. OTTO Institute of Pathology, University of Heidelberg, Germany Department of Molecular Genome Analysis, Deutsches Krebsforschungszentrum, Heidelberg, Germany 2Molecular Biology Laboratory, Neurosurgery Hospital, University of Heidelberg, Germany 3Department of Head and Neck Surgery, Molecular Cell Biology Group, University of Heidelberg, Germany 4Central Unit Biostatistics, Deutsches Krebsforschungszentrum, Heidelberg, Germany 5Department of Dermatology, University of Heidelberg, Germany
Aims: Anorectal melanoma (AM) is a rare but highly malignant tumor, displaying histological and immunohistochemical features similar to cutaneous melanoma (CM). As BRAF mutations were identified in the majority of CM and nevi, we investigated AM for mutations of this oncogene. Methods: DNA from formalin-fixed and paraffin-embedded AM was PCR amplified and sequenced. Results: We detected BRAF exon 15 mutations in only one tumor (1/19 cases), a novel 1800A to T mutation resulted in a K700N transition of the BRAF protein. The oncogenic V599E mutation preponderating in CM was not found in any of the investigated AM. Conclusion: In regard to BRAF kinase domain mutations, AM is clearly different from CM and has molecular aspects of nonmelanocytic tumors.
249
Adenocarcinoid of the Appendix Vermiformis A Case Report I. HIRSCH, H.-U. KASPER, J. BRABENDER l, A.H. HOLSCHEW, H.P. DIENES Institut fOr Pathologic, Univcrsit~it K01n 1Klinik und Poliklinik for Viszeral- und Gef~igchirurgie, Universitat Ktln, Joseph-Stezlmann-Str. 9, 50924 K61n
Aims: Adenocarcinoids are rare tumors with histological features of both adenocarcinoma and carcinoid tumor. The most common site of occurrence is the appendix (2% of primary appendicular tumors). The mean age is 58 years. Methods/Results: A 63-year-old woman suffered from abdominal pain in the lower right quadrant, fever and nausea. A CT-scan demonstrated a thickened wall of the appendix. Under the diagnosis of an acute appendicitis an appendectomy was performed. The appendix revealed a severe gangrenous appendicitis. In addition to the signs of inflammation there was a diffuse infiltration of the appendicular wall by groups of tumor cells containing PASpositive mucin. Staining for CEA was strongly positive, some tumor cells were positive for Synaptophysin, Chromogranin and IgA. Staining for NSE, Serotonin and S-100 was negative. The tumor showed a perineural and vascular invasion. The pathological
247 Rare BRAF Mutations Distinguish Anorectal from Cutaneous Melanoma
321
248 Analysis of TolMike receptors 2, 4 and MD2 in inflamed and non-inflamed gut tissues in inflammatory bowel disease F. AUTSCHBACH, T. GIESE 1, N. GASSLER, I. BERGER, B. SIDO 2, G. HEUSCHEN 2, S.C. MEUEW, H.F. OTTO Pathologisches Institut, Universit~it Heidelberg *Institut fur Immunologie, Universitat Heidelberg 2Chimrgische Universitatsklinik, Universitat Heidelberg
Aims: A perturbed host response to the enteric microbial flora plays a role in the pathogenesis of chronic inflammatory bowel diseases (IBD) like Crohn's disease (CD) and ulcerative colitis (UC). In this study, we investigated the intestinal expression of Toll-like receptors (TLRs), important signaling receptors which can be activated by bacterial wall products such as fipopolysaccharide. Methods: Expression of mRNA-transcripts for TLR2, TLR 4 and the TLR-4-related coreceptor MD2 was analyzed by real-time RTPCR on a large number of transmural gut samples from actively inflamed and non-inflamed/inactive areas in IBD (n = 83) and controls (n = 27). Cells expressing TLR2 and TLR4 protein were characterized by immunohistochemistry (IH). Additional paraffin sections were immunostained for Ki-67 antigen (Mib-1) to investigate the proliferative activity of lamina propria leucocytes (LPLs). Results: Transcript levels for TLR2, TLR4 and MD2 were significantly elevated in samples with active CD and UC as compared with normal controls. IH revealed LPLs to be the predominant TLR-expressing cell type. No increased expression of TLRmRNAs was observed in noninfiamed/inactive areas in IBD. Elevated TLR-transcripts in IBD were accompanied by a significant increase in the number of Ki-67-positive proliferating LPLs. In the case of TLR4, transcript levels were significantly higher in inflamed CD as compared with UC or non-IBD-related inflammatory controls. Conclusions: Induction of TLRs and receptor signalling via bacteri-al wall products contribute to the exaggerated immune response of intestinal LPLs in actively inflamed, but not in inactive areas in IBD. TLR4 mRNA is predominantly induced in CD as compared with UC.
B.M. HELMKE, J. MOLLENHAUER l , C. HEROLD-MENDE 2'3, A. BENNER 4, M. DEICHMANN 5, H.F. OTTO Institute of Pathology, University of Heidelberg, Germany Department of Molecular Genome Analysis, Deutsches Krebsforschungszentrum, Heidelberg, Germany 2Molecular Biology Laboratory, Neurosurgery Hospital, University of Heidelberg, Germany 3Department of Head and Neck Surgery, Molecular Cell Biology Group, University of Heidelberg, Germany 4Central Unit Biostatistics, Deutsches Krebsforschungszentrum, Heidelberg, Germany 5Department of Dermatology, University of Heidelberg, Germany
Aims: Anorectal melanoma (AM) is a rare but highly malignant tumor, displaying histological and immunohistochemical features similar to cutaneous melanoma (CM). As BRAF mutations were identified in the majority of CM and nevi, we investigated AM for mutations of this oncogene. Methods: DNA from formalin-fixed and paraffin-embedded AM was PCR amplified and sequenced. Results: We detected BRAF exon 15 mutations in only one tumor (1/19 cases), a novel 1800A to T mutation resulted in a K700N transition of the BRAF protein. The oncogenic V599E mutation preponderating in CM was not found in any of the investigated AM. Conclusion: In regard to BRAF kinase domain mutations, AM is clearly different from CM and has molecular aspects of nonmelanocytic tumors.
249
Adenocarcinoid of the Appendix Vermiformis A Case Report I. HIRSCH, H.-U. KASPER, J. BRABENDER l, A.H. HOLSCHEW, H.P. DIENES Institut fOr Pathologic, Univcrsit~it K01n 1Klinik und Poliklinik for Viszeral- und Gef~igchirurgie, Universitat Ktln, Joseph-Stezlmann-Str. 9, 50924 K61n
Aims: Adenocarcinoids are rare tumors with histological features of both adenocarcinoma and carcinoid tumor. The most common site of occurrence is the appendix (2% of primary appendicular tumors). The mean age is 58 years. Methods/Results: A 63-year-old woman suffered from abdominal pain in the lower right quadrant, fever and nausea. A CT-scan demonstrated a thickened wall of the appendix. Under the diagnosis of an acute appendicitis an appendectomy was performed. The appendix revealed a severe gangrenous appendicitis. In addition to the signs of inflammation there was a diffuse infiltration of the appendicular wall by groups of tumor cells containing PASpositive mucin. Staining for CEA was strongly positive, some tumor cells were positive for Synaptophysin, Chromogranin and IgA. Staining for NSE, Serotonin and S-100 was negative. The tumor showed a perineural and vascular invasion. The pathological