Effects of common haplotypes of the ileal sodium dependent bile acid transporter gene on the development of sporadic and familial colorectal cancer: A case control study

<p>Abstract</p> <p>Background</p> <p>The genetics of sporadic and non-syndromic familial colorectal cancer (CRC) is not well defined. However, genetic factors that promote the development of precursor lesions, i.e. adenomas, might also predispose to CRC. Recently, an association of colorectal adenoma with two variants (c.507C>T;p.L169L and c.511G>T;p.A171S) of the ileal sodium dependent bile acid transporter gene (<it>SLC10A2</it>) has been reported. Here, we reconstructed haplotypes of the <it>SLC10A2 </it>gene locus and tested for association with non-syndromic familial and sporadic CRC compared to 'hyper-normal' controls who displayed no colorectal polyps on screening colonoscopy.</p> <p>Methods</p> <p>We included 150 patients with sporadic CRC, 93 patients with familial CRC but exclusion of familial adenomatous polyposis and Lynch's syndrome, and 204 'hyper-normal' controls. Haplotype-tagging <it>SLC10A2 </it>gene variants were identified in the Hapmap database and genotyped using PCR-based 5' exonuclease assays with fluorescent dye-labelled probes. Haplotypes were reconstructed using the PHASE algorithm. Association testing was performed with both SNPs and reconstructed haplotypes.</p> <p>Results</p> <p>Minor allele frequencies of all <it>SLC10A2 </it>polymorphisms are within previously reported ranges, and no deviations from Hardy-Weinberg equilibrium are observed. However, we found no association with any of the <it>SLC10A2 </it>haplotypes with sporadic or familial CRC in our samples (all P values > 0.05).</p> <p>Conclusion</p> <p>Common variants of the <it>SLC10A2 </it>gene are not associated with sporadic or familial CRC. Hence, albeit this gene might be associated with early stages of colorectal neoplasia, it appears not to represent a major risk factor for progression to CRC.</p

PD-L1 expression in HNPCC-associated colorectal cancer

Background: PD-L1 immunohistochemistry is predictive for molecular inhibitors of PD-1/PD-L1 immune checkpoint. Therefore, this study evaluated the PD-L1 expression in patients with Hereditary Non-Polyposis Colorectal Cancer (HNPCC). Methods: Immunohistochemical expression of PD-L1 in carcinoma cells, stromal macrophages and lymphocytes of 40 HNPCC-patients with colorectal cancer was scored semi-quantitatively. Results: Focal (2 cases) to extensive (2 cases) PD-L1-immunopositivity of carcinoma cells was detected in 4 out of 40 cases (10.0%). Stromal macrophages were immunopositive in 28 out of 40 cases (70.0%). Lymphocytes showed PD-L1 expression in 3 out of 40 cases (7.5%). Simultaneous immunopositivity of stromal macrophages and tumor cells was detected in two MLH1/PMS2-deficient and two MSH2/MSH6-deficient cases. Conclusion: A subset of HNPCC-associated colorectal cancers in this study clearly showed PD-L1 expression of tumor epithelia and immune cells, therefore, the detection of PD-L1 status is useful

Tumoral expression of nuclear cofactor FHL2 is associated with lymphatic metastasis in sporadic but not in HNPCC-associated colorectal cancer

Background: Four and a half LIM domain protein-2 (FHL2) is part of the focal adhesion structures modulating cell motility. FHL2 may translocate into the nucleus serving as a transcriptional cofactor binding several transcription factors. Overexpression of FHL2 has been linked to cancer progression in various neoplasias. The aim of the present study was to determine, whether FHL2's function as nuclear cofactor plays a prognostic role in invading tumor cells of sporadic and HNPCC-associated colorectal cancer (CRC). Design: Immunohistochemical staining intensity of nuclear FHL2 was quantified by Remmele score analysing 47 sporadic and 42 HNPCC-associated colorectal cancers. Analysis was restricted to carcinoma cells of the tumoral invasion front. Results: Confocal microscopy detected nuclear expression of FHL2 in colon cancer cells and absence of nuclear FHL2 signal in normal colon enterocytes. In colon cancer, nuclear FHL2 expression was predominantly observed in low-differentiated, often mucinous tumor areas. 42.55% of sporadic and 54.76% of HNPCC-associated CRC showed enhanced (Remmele score 6-12) nuclear FHL2 expression in the carcinoma cells of the tumoral advancing edge. Enhanced nuclear FHL2 expression was significantly linked to lymphatic metastasis in sporadic CRC (p = 0.0197) and almost reached significance in HNPCC-associated CRC (p = 0.0545). In contrast, nuclear FHL2 expression was neither associated with hematogenic metastasis in sporadic (p = 0.7087) nor in HNPCC-associated colorectal cancer (p = 0.3007). Conclusions: We recently demonstrated that enhanced nuclear FHL2 expression in tumor stroma of sporadic colon cancer is associated with lymphatic metastasis. The results of the present study indicate a synergistic effect of nuclear cofactor FHL2 in tumor cells as well as in peritumoral stroma cells promoting lymphatic metastasis in sporadic CRC. As HNPCC-associated tumors did not show a significant association between tumoral nuclear FHL2 expression and lymphatic metastasis we speculate, that the intensive lymphocytic immune response in HNPCC precludes a direct contact of tumor cells and stromal cells resulting in reduced lymphatic spread. (C) 2014 Elsevier GmbH. All rights reserved

Enhanced Tumoral MLH1-Expression in MLH1-/PMS2-Deficient Colon Cancer Is Indicative of Sporadic Colon Cancer and Not HNPCC

Hereditary Non-Polyposis Colorectal Cancer (HNPCC) is caused by germline mutations of mismatch-repair (MMR) genes MLH1, MSH2, MSH6 and PMS2. MLH1-/PMS2-deficient colorectal carcinomas might be HNPCC-associated but also caused by MLH1-promoter methylation in sporadic colon carcinoma. This study analyzed semiquantitatively whether the MLH1 staining pattern might be indicative of sporadic or HNPCC-associated colorectal cancer. Using a semiquantitative score ranging from 0 (negative) to 12 (maximum immunopositivity) we analyzed MLH1 expression patterns in 130 MLH1-/PMS2-deficient colorectal cancers. The collective consisted of 70 HNPCC-associated colorectal cancers and 60 sporadic colon cancers. In tumor cells of 70 HNPCC-associated colorectal cancers, 64 cases (91.43%) showed no MLH1 staining, 5 cases weak (7.14%) and 1 case (1.43%) stronger staining intensity. In contrast, in tumor cells of 60 sporadic colorectal cancers 45 cases (75.0%) showed no MLH1 staining, 10 cases weak (16.67%) and 5 cases (8.33%) stronger staining intensity to a varying extent. In immunopositive cases, MLH1 showed a characteristic dot-like nuclear staining pattern in the tumor cells. We compared cases with absent to weak MLH1-staining (immunoscores 0 to 2) to cases with elevated immunoscores (3 to 12) detecting a statistically significant difference between HNPCC-associated and sporadic colon cancers (pvalue = 0.0031, Fisher's exact test). Taken together, enhanced tumoral MLH1 expression in MLH1-/PMS2-deficient colorectal carcinomas seems to be indicative of sporadic origin. In contrast, HNPCC-associated colorectal cancer showed absent or very weak MLH1 immunopositivity. Therefore, this semiquantitative and easy to exert MLH1 immunoscore might help to identify sporadic MLH1-/PMS2-deficient colorectal cancer cases prior to time-consuming methylation analysis

TGF-beta 1-dependent induction and nuclear translocation of FHL2 promotes keratin expression in pilomatricoma

Pilomatricoma is a tumour derived from hair matrix cells, which shows progressive keratin expression. Tumorigenesis is frequently associated with activating mutations in beta-catenin gene inducing nuclear expression of beta-catenin protein. The present study analysed the role of transforming growth factor-beta 1 (TGF-beta 1) and four-and-a-half LIM domain protein 2 (FHL2) in pilomatricoma in synopsis with their expression patterns in human anagen hair. Human anagen hair showed TGF-beta 1 and nuclear FHL2 expression in the outer root sheath layer separated from nuclear beta-catenin staining, which was observed in cells of matrix and inner root sheath layers. Correspondingly, 41 out of 50 pilomatricomas showed co-labelling of TGF-beta 1 and nuclear FHL2 in tumour cells, which mostly lacked nuclear beta-catenin expression. Tumoural proliferation (ki67) was associated with nuclear beta-catenin staining but not with expression of nuclear FHL2. In early pilomatricomas, TGF-beta 1 expression was observed in few peripheral tumour cells showing absent or faint nuclear FHL2 co-staining. TGF-beta 1 expression extended in growing tumours going along with strong nuclear FHL2 co-labelling as well as progressive keratin 14 and keratin 1 expression. In vitro, cultured human keratinocytes showed weak to marked autocrine TGF-beta 1 expression; in case of enhanced TGF-beta 1 expression associated with keratin 10 staining. TGF-beta 1-treatment of cultured human keratinocytes induced nuclear and cytoplasmatic FHL2 staining as well as keratin 14 staining. Accordingly, siRNA-mediated FHL2 knockdown of TGF-beta 1-stimulated keratinocytes reduced keratin 14 staining. In conclusion, tumoural TGF-beta 1 secretion seems to induce nuclear translocation of co-factor FHL2 mediating progressive keratin expression in pilomatricoma

Prognostic Significance of DNA Cytometry in Carcinoma of the Uterine Cervix FIGO Stage IB and II

Objective: To assess the prognostic value of DNA‐image cytometry in cervical carcinoma of the uterus and its relation to other established prognostic factors. Study design: The study included 116 cases of cervical carcinoma FIGO stages IB and II which were treated with radical abdominal hysterectomy. The median follow‐up was 55 months (range 1–162 months). DNA image cytometry was performed on cytologic specimens prepared by enzymatic cell separation from formalin‐fixed, paraffin‐embedded tissues. DNA stemline ploidy, DNA stemline aneuploidy, 5c exceeding rate, 9c exceeding rate, 2c deviation index, and DNA malignancy grade were computed. DNA‐variables as well as various clinical and histological variables were related to survival rates. Results: In multivariate statistical analysis DNA stemline ploidy using 2.2c as a cut‐off value and FIGO stage showed to be statistically significant available presurgery predictors of survival, whereas the postsurgical parameters lymphonodal status, tumor size and parametrial involvement were significantly correlated with survival. The synopsis of all parameters in a multivariate Cox model indicated that – with declining relevance – the number of positive pelvic lymph nodes, DNA stemline ploidy using a cut‐off level at a modal value of 2.2c, largest pelvic lymph node, 5c exceeding rate, and ratio of carcinoma area to cervix area, were of predictive value for survival. Conclusions: Our results suggest that prognostic information deducted from classical staging parameters is successfully complemented by DNA image cytometry which can be applied pretherapeutically