Analytical Cellular Pathology - Volume 33, issue 2

Authors: Gu, Lei | Zhu, Xian-Hua | Visakorpi, Tapio | Alanen, Kalle | Mirtti, Tuomas | Edmonston, Tina Bocker | Nevalainen, Marja T.

Article Type: Research Article

Abstract: Background: Transcription factor Stat5a/b is highly critical for the viability of human prostate cancer cells in vitro and for prostate tumor growth in vivo. Stat5 is constitutively active in clinical prostate cancers but not in the normal human prostate epithelium. Moreover, Stat5a/b activation in prostate cancer is associated with high histological grade of prostate cancer. However, the molecular mechanisms underlying constitutive activation of Stat5a/b in prostate cancer are unclear. The receptor-associated tyrosine kinase Jak2 is a known key activator of Stat5a/b in prostate cancer cells in response to ligand stimulation. Recently, a single gain-of-function point mutation of JAK2 was described …in myeloproliferative diseases leading to constitutive Jak2 kinase activity, subsequent Stat5a/b activation and involvement of V617F Jak2 in the pathogenesis of myeloproliferative disorders. Materials and methods: We determined whether JAK2 undergoes the V617F activating mutation during clinical progression of human prostate cancer using a highly sensitive assay (amplification refractory mutation system) and a unique material of fresh specimens from organ-confined or castration-resistant prostate cancers. Results: The JAK2 V617F mutation was not found in any of the normal or malignant prostate samples analyzed in this study. Conclusions: Future work should focus on determining the molecular mechanisms other than V617F mutation of Jak2 resulting in continuous Stat5 activation in clinical prostate cancers. Show more

Keywords: JAK2 V617 mutation, prostate cancer

DOI: 10.3233/ACP-CLO-2010-0534

Citation: Analytical Cellular Pathology, vol. 33, no. 2, pp. 55-59, 2010

Authors: Jotzu, Constantin | Alt, Eckhard | Welte, Gabriel | Li, Jie | Hennessy, Bryan T. | Devarajan, Eswaran | Krishnappa, Srinivasalu | Pinilla, Severin | Droll, Lilly | Song, Yao-Hua

Article Type: Research Article

Abstract: Carcinoma-associated fibroblasts (CAF) are considered to contribute to tumor growth, invasion and metastasis. However, the cell type of origin remains unknown. Since human adipose tissue-derived stem cells (hASCs) are locally adjacent to breast cancer cells and might directly interact with tumor cells, we investigated whether CAFs may originate from hASCs. We demonstrated that a significant percentage of hASCs differentiated into a CAF-like myofibroblastic phenotype (e.g., expression of alpha smooth muscle actin and tenascin-C) when exposed to conditioned medium from the human breast cancer lines MDAMB231 and MCF7. The conditioned medium from MDAMB231 and MCF7 contains significant amounts of transforming growth …factor-beta 1 (TGFβ1) and the differentiation of hASCs towards CAFs is dependent on TGFβ1 signaling via Smad3 in hASCs. The induction of CAFs can be abolished using a neutralizing antibody to TGFβ1 as well as by pretreatment of the hASCs with SB431542, a TGFβ1 receptor kinase inhibitor. Additionally, we found that these hASC-derived CAF-like cells exhibit functional properties of CAFs, including the ability to promote tumor cell invasion in an in vitro invasion assay, as well as increased expression of stromal-cell-derived factor 1 (SDF-1) and CCL5. Taken together, these data suggest that hASCs are a source of CAFs which play an important role in the tumor invasion. Show more

Keywords: Breast cancer, mesenchymal stem cells, carcinoma-associated fibroblasts, transforming growth factor-beta 1, invasion

DOI: 10.3233/ACP-CLO-2010-0535

Citation: Analytical Cellular Pathology, vol. 33, no. 2, pp. 61-79, 2010

Authors: Shaw, Elisabeth J. | Haylock, Brian | Husband, David | du Plessis, Daniel | Sibson, D. Ross | Warnke, Peter C. | Walker, Carol

Article Type: Research Article

Abstract: Background: Oligodendroglial tumors with 1p/19q loss are more likely to be chemosensitive and have longer survival than those with intact 1p/19q, but not all respond to chemotherapy, warranting investigation of the biological basis of chemosensitivity. Methods: Gene expression profiling was performed using amplified antisense RNA from 28 oligodendroglial tumors treated with chemotherapy (26 serial stereotactic biopsy, 2 resection). Expression of differentially expressed genes was validated by real-time PCR. Results: Unsupervised hierarchical clustering showed clustering of multiple samples from the same case in 14/17 cases and identified subgroups associated with tumor grade and 1p/19q status. 176 genes were differentially …expressed, 164 being associated with 1p/19q loss (86% not on 1p or 19q). 94 genes differed between responders and non-responders to chemotherapy; 12 were not associated with 1p/19q loss. Significant differential expression was confirmed in 11/13 selected genes. Novel genes associated with response to therapy included SSBP2, GFRA1, FAP and RASD1. IQGAP1, INA, TGIF1, NR2F2 and MYCBP were differentially expressed in oligodendroglial tumors with 1p/19q loss. Conclusion: Gene expression profiling using serial stereotactic biopsies indicated greater homogeneity within tumors than between tumors. Genes associated with 1p/19q status or response were identified warranting further elucidation of their role in oligodendroglial tumors. Show more

Keywords: Oligodendroglioma, gene expression, chemosensitivity

DOI: 10.3233/ACP-CLO-2010-0533

Citation: Analytical Cellular Pathology, vol. 33, no. 2, pp. 81-94, 2010

Authors: Brosens, R.P.M. | Belt, E.J.T.H. | Haan, J.C. | Buffart, T.E. | Carvalho, B. | Grabsch, H. | Quirke, P. | Cuesta, M.A. | Engel, A.F. | Ylstra, B. | Meijer, G.A.

Article Type: Research Article

Abstract: Background: Around 30% of all stage II colon cancer patients will relapse and die of their disease. At present no objective parameters to identify high-risk stage II colon cancer patients, who will benefit from adjuvant chemotherapy, have been established. With traditional histopathological features definition of high-risk stage II colon cancer patients is inaccurate. Therefore more objective and robust markers for prediction of relapse are needed. DNA copy number aberrations have proven to be robust prognostic markers, but have not yet been investigated for this specific group of patients. The aim of the present study was to identify chromosomal aberrations that …can predict relapse of tumor in patients with stage II colon cancer. Materials and methods: DNA was isolated from 40 formaldehyde fixed paraffin embedded stage II colon cancer samples with extensive clinicopathological data. Samples were hybridized using Comparative Genomic Hybridization (CGH) arrays to determine DNA copy number changes and microsatellite stability was determined by PCR. To analyze differences between stage II colon cancer patients with and without relapse of tumor a Wilcoxon rank-sum test was implemented with multiple testing correction. Results: Stage II colon cancers of patients who had relapse of disease showed significantly more losses on chromosomes 4, 5, 15q, 17q and 18q. In the microsatellite stable (MSS) subgroup (n=28), only loss of chromosome 4q22.1–4q35.2 was significantly associated with disease relapse (p<0.05, FDR<0.15). No differences in clinicopathological characteristics between patients with and without relapse were observed. Conclusion: In the present series of MSS stage II colon cancer patients losses on 4q22.1–4q35.2 were associated with worse outcome and these genomic alterations may aid in selecting patients for adjuvant therapy. Show more

Keywords: DNA copy number changes, stage II, colon cancer, prognosis

DOI: 10.3233/ACP-CLO-2010-0531

Citation: Analytical Cellular Pathology, vol. 33, no. 2, pp. 95-104, 2010

Authors: Gaiser, Timo | Berroa-Garcia, Lissa | Kemmerling, Ralf | Dutta, Aparajita | Ried, Thomas | Heselmeyer-Haddad, Kerstin

Article Type: Research Article

Abstract: Background: The simultaneous detection of protein expression and gene copy number changes in patient samples, like paraffin-embedded tissue sections, is challenging since the procedures of immunohistochemistry (IHC) and Fluorescence in situ Hybridization (FISH) negatively influence each other which often results in suboptimal staining. Therefore, we developed a novel automated algorithm based on relocation which allows subsequent detection of protein content and gene copy number changes within the same cell. Methods: Paraffin-embedded tissue sections of colorectal cancers were stained for CD133 expression. IHC images were acquired and image coordinates recorded. Slides were subsequently hybridized with fluorescently labeled DNA probes. …FISH images were taken at the previously recorded positions allowing for direct comparison of protein expression and gene copy number signals within the same cells/tissue areas. Relocation, acquisition of the IHC and FISH images, and enumeration of FISH signals in the immunophenotyped tumour areas were done in an automated fashion. Results: Automated FISH analysis was performed on 13 different colon cancer samples that had been stained for CD133; each sample was scored for MYC, ZNF217 and Chromosome 6 in CD133 positive and negative glands. From the 13 cases four (31%) showed amplification for the MYC oncogene and seven of 13 (54%) cases were amplified for ZNF217. There was no significant difference between CD133 positive tumour and CD133 negative tumour cells. Conclusion: The technique and algorithm presented here enables an easy and reproducible combination of IHC and FISH based on a novel automated algorithm using relocation and automated spot counting. Show more

Keywords: Automated image analysis, fluorescence in situ hybridization, immunofluorescence, image relocation

DOI: 10.3233/ACP-CLO-2010-0532

Citation: Analytical Cellular Pathology, vol. 33, no. 2, pp. 105-112, 2010