Expression of the carbohydrate tumour marker Sialyl Lewis A, Sialyl Lewis X, Lewis Y and Thomsen-Friedenreich antigen in normal squamous epithelium of the uterine cervix, cervical dysplasia and cervical cancer.

The carbohydrate molecules Sialyl Lewis X (SLeX), Sialyl Lewis A (SLeA), Lewis Y (LeY) and Thomsen-Friedenreich antigen (TF) are known to mediate the adhesion between tumor cells and endothelium. They are used as serum markers in diagnosis and treatment in a broad spectrum of human carcinomas, but their expression profile and role in the development of cervical cancer remains unclear. The aim of this study was to investigate the expression of SLeX, SLeA, LeY and TF in normal cervical squamous epithelium, cervical dysplasia and cervical cancer. Slides of paraffin-embedded tissue were fixed and incubated with monoclonal antibodies against SLeX, SLeA, LeY and TF. Immunohistochemical staining was evaluated by using a semi-quantitative score (IRS Score). We found a significant difference of SLeA expression in invasive cervical cancer compared to normal epithelium (p=0.006) and all grades of dysplasia (p=0.002). The expression of SLeX in normal epithelium was less intense than in carcinoma in situ (p=0.036). Staining for LeY showed the weakest results of the investigated markers. Significant differences were found when normal epithelium was compared to CIN I (p=0.011), to CIN II (p=0.013) and to invasive cervical cancer (p=0.005). For TF, significant differences were found in normal epithelium compared to CIN I (p=0.011), CIN II (p=0.013) and compared to invasive cervical cancer (p=0.005). This is the first study on the expression of SLeA, SLeX, LeY and TF in normal cervical endothelium, cervical dysplasia, carcinoma in situ and invasive cervical cancer. Further studies and higher numbers are desirable.

Topoisomerase IIalpha expression rather than gene amplification predicts responsiveness of adjuvant anthracycline-based chemotherapy in women with primary breast cancer.

INTRODUCTION: Adjuvant anthracycline-based chemotherapy (AbCTX) is a standard treatment for patients with primary breast cancer. Its main target is topoisomerase IIalpha (TopIIa), a nuclear protein which is important for DNA replication and mitosis. We propose that the overexpression of the TopIIa protein or amplification of the TopIIa gene may be useful in predicting increased responsiveness towards AbCTX. METHODS: Tumor tissues of 118 patients who received adjuvant AbCTX were examined by immunohistochemistry (IHC) and fluorescence in situ hybridisation (FISH) for TopIIa and HER2. For IHC, the primary antibodies 485 (Dako) and NCL-TOPOIIA (Novocastra) were used. FISH analysis was performed with the SPEC HER2/CEP 17 Dual Color Probe (Zytovision) and LSI TOP 2A Spectrum Orange/CEP 17 Spectrum Green probe (Abbott). TopIIa IHC was evaluated by the immunoreactive score (IRS). FISH amplification was stated at an HER2-TopIIa/CEP 17 ratio > or = 2, deletion of TopIIa at a ratio <0.8. RESULTS: The median age of the patient population was 50 years (range 23-77), 76 (64%) had tumors >2 cm in size, 98 (85%) were nodal positive, and 72 (66%) estrogen-receptor positive. Chemotherapy regimes consisted of epirubicin-cyclophosphamide (EC 40 pts), EC-CMF (18 pts), FAC/FEC (33 pts), anthracycline-taxane combinations (23 pts) and others (4 pts). After IHC, it was found that 19% of the tumors were positive for HER2 (3+) and the median IRS for TopIIa staining was 2 (49% positive); 28 (24%) tumors showed HER2 amplification, therefrom 20/22 (91%) within the HER2 3+ group. TopIIa gene was amplified in 17 cases (16%) and deletion was seen in 6 (5%) tumors. Of all cases with HER2 gene amplification, 14 (50%) cases of TopIIa co-amplification and one case of deletion were seen. Looking at histological parameters, TopIIa IHC correlated with nodal status (P = 0.018) and high grading (G3) (P = 0.047). After a median follow-up of 42 months (range 1-242), a significant prognostic factor for local recurrence was HER2 positivity (IHC P = 0.013 and FISH P = 0.023). Thirty-two patients developed metastasis (27%), which was correlated with HER2 FISH positivity (P = 0.024) and, as a trend, Top IIa IHC negativity (P = 0.094); 25 (21%) patients died from the disease. Negative prognostic parameters were the lack of estrogen-receptor expression (P = 0.008), lymphangiosis (P = 0.02), and TopIIa IHC negativity (P = 0.03). CONCLUSION: In this cohort of patients, HER2 positivity indicated higher rates of local and distant recurrence. In contrast, TopIIa IHC positivity predicted lower risk of metastases and death, thus being a positive-predictive factor for the responsiveness to AbCTX. TopIIa gene amplification did not add predictive information. Therefore, we conclude that TopIIa protein expression might rather be the target of anthracyclines independent from gene copy number.

CpG-island methylation of the ER promoter in colorectal cancer: analysis of micrometastases in lymph nodes from UICC stage I and II patients.

Patients with UICC stage II colorectal cancer (CRC) have a risk of approximately 20% to develop disease recurrence after tumour resection. The presence and significance of micrometastases for locoregional recurrence in these patients lacking histopathological lymph node involvement on routine stained HE sections is undefined. Oestrogen receptor (ER) promoter methylation has earlier been identified in CRC. Therefore, we evaluated the methylation status of the ER promoter in lymph nodes from 49 patients with CRC UICC stage I and II as a molecular marker of micrometastases and predictor of local recurrence. DNA from 574 paraffin-embedded lymph nodes was isolated and treated with bisulphite. For the detection of methylated ER promoter sequences, quantitative real-time methylation-specific PCR was used. Of the 49 patients tested, 15 (31%) had ER methylation-positive lymph nodes. Thirteen of those (86%) remained disease free and two (14%) developed local recurrence. In the resected lymph nodes of 34 of the 49 patients (69%), no ER promoter methylation could be detected and none of these patients experienced a local relapse. The methylation status of the ER promoter in lymph nodes of UICC stage I and II CRC patients may be a useful marker for the identification of patients at a high risk for local recurrence.