Analytical Cellular Pathology - Volume 32, issue 1-2

Authors: Weber, Achim | Chung, Hye-Jung | Springer, Erik | Heitzmann, Dirk | Warth, Richard

Article Type: Research Article

Abstract: Background: The general transcription factor II H (TFIIH), comprised of a core complex and an associated CAK-complex, functions in transcription, DNA repair and cell cycle control. Mutations of the two largest subunits, p89 (XPB) and p80 (XPD), cause the hereditary cancer-prone syndrome xeroderma pigmentosum. Methods: The TFIIH subunit p89 was monitored during interphase and cell division by immunofluorescence staining, GFP-fusion constructs including deletions, live cell imaging and immuno-precipitations. Results: Here we demonstrate that during cell division, from prophase until telophase, the TFIIH core subunit p89, but not other subunits of TFIIH, associates with the centrosomes and the adjacent …parts of the mitotic spindle. With overall constant levels throughout mitosis, p89 re-localizes to the newly formed nuclei by the end of mitosis. Furthermore, p89 interacts with the centrosomal protein γ-tubulin. Truncations of p89 result in an abnormal subcellular distribution during interphase and abolished centrosomal association during mitosis. Conclusions: Our observations suggest a so far unappreciated role for p89 in cell cycle regulation, and may be the structural basis for a long known, but hitherto unexplained interaction between p89 and tubulin. Show more

Keywords: TFIIH, mitosis, centrosome

DOI: 10.3233/CLO-2009-0509

Citation: Analytical Cellular Pathology, vol. 32, no. 1-2, pp. 121-130, 2010

Authors: Yang, Nan | Nijhuis, Esther R. | Volders, Haukeline H. | Eijsink, Jasper J.H. | Lendvai, Ágnes | Zhang, Bo | Hollema, Harry | Schuuring, Ed | Wisman, G. Bea A. | van der Zee, Ate G.J.

Article Type: Research Article

Abstract: Objectives: To determine methylation status of nine genes, previously described to be frequently methylated in cervical cancer, in squamous intraepithelial lesions (SIL). Methods: QMSP was performed in normal cervix, low-grade (L)SIL, high-grade (H)SIL, adenocarcinomas and squamous cell cervical cancers, and in corresponding cervical scrapings. Results: Only CCNA1 was never methylated in normal cervices and rarely in LSILs. All other genes showed methylation in normal cervices, with CALCA, SPARC and RAR-β2 at high levels. Methylation frequency of 6 genes (DAPK, APC, TFPI2, SPARC, CCNA1 and CADM1) increased with severity of the underlying cervical lesion. DAPK showed the highest …increase in methylation frequency between LSIL and HSIL (10% vs. 40%, p<0.05), while CCNA1 and TFPI2 were most prominently methylated in cervical cancers compared to HSILs (25% vs. 52%, p<0.05, 30% vs. 58%, p<0.05). CADM1 methylation in cervical cancers was related to depth of invasion (p<0.05) and lymph vascular space involvement (p<0.01), suggesting a role in invasive potential of cervical cancers. Methylation ratios in scrapings reflected methylation status of the underlying lesions (p<0.05). Conclusion: Methylation of previously reported cervical cancer specific genes frequently occurs in normal epithelium. However, frequency of methylation increases during cervical carcinogenesis, with CCNA1 and DAPK as the best markers to distinguish normal/LSIL from HSIL/cancer lesions. Show more

Keywords: Methylation, cervical (intraepithelial) neoplasia, DAPK, CCNA1, CADM1

DOI: 10.3233/CLO-2009-0510

Citation: Analytical Cellular Pathology, vol. 32, no. 1-2, pp. 131-143, 2010

Authors: Meijer, Gerrit

Article Type: Other

DOI: 10.3233/CLO-2010-0513

Citation: Analytical Cellular Pathology, vol. 32, no. 1-2, pp. 147-148, 2010

Authors: Moelans, Cathy B. | Kibbelaar, Robby E. | van den Heuvel, Marius C. | Castigliego, Domenico | de Weger, Roel A. | van Diest, Paul J.

Article Type: Research Article

Abstract: Background: Testing for HER2 amplification and/or overexpression is currently routine practice to guide Herceptin therapy in invasive breast cancer. At present, HER2 status is most commonly assessed by immunohistochemistry (IHC). Standardization of HER2 IHC assays is of utmost clinical and economical importance. At present, HER2 IHC is most commonly performed with the HercepTest which contains a polyclonal antibody and applies a manual staining procedure. Analytical variability in HER2 IHC testing could be diminished by a fully automatic staining system with a monoclonal antibody. Materials and methods: 219 invasive breast cancers were fully automatically stained with the monoclonal antibody-based Oracle …HER2 Bond IHC kit and manually with the HercepTest. All cases were tested for amplification with chromogenic in situ hybridization (CISH). Results: HercepTest yielded an overall sharper membrane staining, with less cytoplasmic and stromal background than Oracle in 17% of cases. Overall concordance between both IHC techniques was 89% (195/219) with a kappa value of 0.776 (95% CI 0.698–0.854), indicating a substantial agreement. Most (22/24) discrepancies between HercepTest and Oracle showed a weaker staining for Oracle. Thirteen of the 24 discrepant cases were high-level HER2 amplified by CISH, and in 12 of these HercepTest IHC better reflected gene amplification status. All the 13 HER2 amplified discrepant cases were at least 2+ by HercepTest, while 10/13 of these were at least 2+ for Oracle. Considering CISH as gold standard, sensitivity of HercepTest and Oracle was 91% and 83%, and specificity was 94% and 98%, respectively. Positive and negative predictive values for HercepTest and Oracle were 90% and 95% for HercepTest and 96% and 91% for Oracle, respectively. Conclusion: Fully-automated HER2 staining with the monoclonal antibody in the Oracle kit shows a high level of agreement with manual staining by the polyclonal antibody in the HercepTest. Although Oracle shows in general some more cytoplasmic staining and may be slightly less sensitive in picking up HER2 amplified cases, it shows a higher specificity and may be considered as an alternative method to evaluate the HER2 expression in breast cancer with potentially less analytical variability. Show more

Keywords: HER2, Oracle, HercepTest, CISH

DOI: 10.3233/CLO-2010-0514

Citation: Analytical Cellular Pathology, vol. 32, no. 1-2, pp. 149-155, 2010

Authors: Meijer, Gerrit A.

Article Type: Other

DOI: 10.3233/CLO-2010-0512

Citation: Analytical Cellular Pathology, vol. 32, no. 1-2, pp. 156-156, 2010