J.C. van der Linden

Peripheral primitive neuroectodermal tumour and extra-osseous Ewing's sarcoma: a histological, immunohistochemical and DNA flow cytometric study

Although peripheral primitive neuroectodermal tumour (pPNET) and extra-osseous Ewings sarcoma (EES) are thought to be closely related neoplasms, their clinical behaviour differs considerably. To determine the clinical relevance of the Schmidt classification scheme for differentiating pPNET and EES, 20 tumour specimens of poorly differentiated round cell tumours were evaluated. In addition, the diagnostic value of several neural markers and the prognostic value of quantitative morphological variables (DNA ploidy, S-phase fraction, and the mitotic activity) were assessed. Homer-Wright rosettes were present in 9 tumours. Neuron specific enolase (NSE) was expressed in 11 tumours, 8 of which expressed a second neural marker (CD57, S100, or neurofilament). According to the Schmidt classification, 11 pPNET and 5 EES were distinguished. HBA-71 was exclusively expressed in pPNET and EES. The remaining tumours were classified as sarcoma not otherwise specified (n=2), rhabdomyosarcoma (n=1), and desmoplastic tumour with divergent differentiation (n=1). EES611 patients fared significantly better than the pPNET patients (100% versus 42% 5-year survival). Neither DNA ploidy nor S-phase fraction assessed in 12 evaluative histograms (9 pPNET and 3 EES), nor mitotic activity yielded information of additional prognostic value. On the basis of this study and the Schmidt classification scheme, it can be concluded that if the diagnosis of EES and pPNET is based on light microscopy (Homer-Wright rosettes) and/or immunohistochemistry (at least two neural markers, i.e. NSE, S-100, CD57, and neurofilament), the classification provides important clinical information. Furthermore, positivity for HBA-71 is helpful in differentiating pPNET and EES from all other small round cell tumours.

Prognostic importance of DNA flow cytometric variables in rhabdomyosarcomas.

AIM--To determine whether DNA ploidy patterns and S phase fraction offer prognostic information in patients with rhabdomyosarcoma (RMS). METHODS--DNA flow cytometry was performed on formalin fixed, paraffin wax embedded samples from primary tumours, and metastatic deposits or recurrences in 70 patients. DNA histogram analysis was done using a semi-automated cell cycle analysis program. RESULTS--Of the 70 primary tumours, 23 were DNA diploid, 32 DNA aneuploid, eight DNA multiploid, and seven DNA tetraploid. The prognosis for DNA aneuploid patterns was favourable, intermediate within the group of DNA tetraploid tumours and poor among patients with DNA diploid and DNA multiploid tumours (p = 0.009). In multivariate analysis (Cox regression model) DNA ploidy was an important independent prognostic factor, along with TNM stage, localisation, and histopathological classification. Ten out of 32 patients with a high S phase fraction (> 15%) with primary RMS achieved long term survival in contrast to 20 out of 29 patients with a low S phase fraction (< or = 15%) (p = 0.008). In 24 cases the DNA ploidy of cases of relapse was analysed. Of the 15 cases, in which stem line changes had occurred, 13 died of disease. No stem line changes were noted in nine cases and in this group four patients died of disease (p = 0.02). CONCLUSIONS--Assessment of DNA ploidy and S phase fraction in primary RMS and evaluation of stem line changes in cases of relapse are important variables in predicting prognosis.

Micro-metastases in stages I and II colon cancer are a predictor of the development of distant metastases and worse disease-free survival

Approximately 30% of the patients with Dukes A/B colon carcinoma will develop loco-regional recurrence or distant metastases. The aim of this study was to evaluate if patients with micro-metastases are at higher risk for developing distant metastases and therefore a worse disease-free survival and overall survival. In the period January 2000-January 2002, 137 patients underwent curative surgery for colon cancer. When patients had a Dukes A/B colon carcinoma, additional staining and sectioning on the harvested lymph nodes were performed retrospectively. Lymph nodes were examined using 4 multilevel sections at 250-microm intervals and stained with Pan-Cytokeratin. There were 11 patients with a Dukes A and 61 patients with a Dukes B colon carcinoma. Twenty-two patients developed metastases in time (group I) whereas 50 patients did not (group II). After additional staining and sectioning 41% of the patients of group I and 16% of the patients of group II showed micro-metastases (p<0.05). The 5-year overall survival rate in the group with micro-metastases was 62% against 79% in the group without micro-metastases. The disease-free survival (DFS) was 51% and 72% (p<0.05), respectively. Patients with micro-metastases develop significant more distant metastases in time and have a significant worse DFS.

The number of high-risk factors is related to outcome in stage II colonic cancer patients.

BACKGROUND A subgroup of stage II colonic cancer patients are considered to be at high-risk for recurrent/metastatic disease based on 1) tumour obstruction/perforation 2) <10 lymph nodes 3) T4 lesions and 4) lymphangio-invasion. Their prognosis is regarded as comparable to stage III (T1-4N+M0) colonic cancer and it is therefore strongly advised to treat them with adjuvant chemotherapy. The purpose of this study was i) to determine the magnitude of prognostic significance of the conventional high-risk factors and ii) to determine whether the number of high-risk factors influences outcome. METHODS We retrospectively analyzed 212 stage II colonic cancer patients undergoing surgery between January 2002 and December 2008. No adjuvant chemotherapy was given. Survival analyses were performed. RESULTS 154/212 (73%) patients were considered to be high-risk patients based on conventional high-risk factors. 58 patients did not meet any high-risk factor, 125 patients met 1 high-risk factor and 29 patients met ≥2 high-risk factors. Median follow up was 40 months. Multivariate analysis identified four independent risk factors for recurrent/metastatic disease: age, obstruction, perforation and lymphangio-invasion. The three-year-DFS-rates for the low-risk group, the high-risk group with 1 high-risk factor and the high-risk group with ≥2 high-risk criteria are 90.4%, 87.6% and 75.9% respectively. Patients meeting ≥2 conventional high-risk criteria had a significantly worse three-year disease free survival (p < 0.002). CONCLUSIONS Four independent high-risk factors were identified. The number of high-risk factors does influence outcome. More attention should be given to the definition and treatment of high-risk stage II colonic cancer patients.

Use of cytokeratin 8 immunohistochemistry for assessing cell death after radiofrequency ablation of breast cancers

Abstract The effects of minimally invasive therapies such as radiofrequency ablation (RFA) and laser induced thermal therapy on breast carcinoma lesions usually is assessed by NADH diaphorase enzyme histochemistry for cell viability. NADH staining requires frozen material, however, with its associated poor morphology. We aimed to validate cytokeratin 8 (CK 8) immunohistochemistry as an alternative that works on paraffin sections. RFA was performed ex vivo on 20 breast resections after surgery and in vivo in eight patients who underwent general anesthesia followed by immediate resection. After treatment, specimens were lamellated and the tumors were divided into two equal parts. One part was fixed in neutral buffered formaldehyde for routine histopathological evaluation using hematoxylin and eosin (H & E) staining and CK 8 immunostaining. The other section was snap frozen and stored at −80° C for staining with NADH diaphorase. Both NADH diaphorase and CK 8 immunostaining demonstrated a clear and comparable demarcation between viable and nonviable tissues. The morphology of the CK 8 immunostained slides was much better, and fatty tissues could be judged readily by contrast to the NADH stained frozen sections, which had poor morphology and whose fatty parts were difficult to interpret. CK 8 immunohistochemistry seems to be well suited for assessing cell viability in breast tissue and for assessing the effects of RFA for breast cancer treatment. Because it can be applied to paraffin fixed material, it provides much better morphology than NADH staining and also can be applied to fatty tissues that usually are difficult to work up for frozen sections. Therefore, CK 8 immunohistochemistry may be preferred over NADH diaphorase staining for daily pathology practice for assessing the viability of breast carcinoma cells after RFA treatment.

Epidermal growth factor receptor (EGFR) and prostaglandin-endoperoxide synthase 2 (PTGS2) are prognostic biomarkers for patients with resected colorectal cancer liver metastases.

Background:Resection of colorectal cancer liver metastasis (CRCLM) with curative intent has long-term benefit in ∼40% of cases. Prognostic biomarkers are needed to improve clinical management and reduce futile surgeries. Expression of epidermal growth factor receptor (EGFR) and prostaglandin-endoperoxide synthase 2 (PTGS2; also known as cyclooxygenase-2) has been associated with carcinogenesis and survival. We investigated the prognostic value of EGFR and PTGS2 expression in patients with resected CRCLM.Methods:Formalin-fixed paraffin-embedded CRCLM tissue and corresponding primary tumour specimens from a multi-institutional cohort of patients who underwent liver resection between 1990 and 2010 were incorporated into tissue microarrays (TMAs). TMAs were stained for EGFR and PTGS2 by immunohistochemistry. The hazard rate ratio (HRR) for the association between expression in CRCLM and overall survival was calculated using a 500-fold cross-validation procedure.Results:EGFR and PTGS2 expression could be evaluated in 323 and 351 patients, respectively. EGFR expression in CRCLM was associated with poor prognosis (HRR 1.54; P<0.01) with a cross-validated HRR of 1.47 (P=0.03). PTGS2 expression was also associated with poor prognosis (HRR 1.60; P<0.01) with a cross-validated HRR of 1.63 (P<0.01). Expression of EGFR and PTGS2 remained prognostic after multivariate analysis with standard clinicopathological variables (cross-validated HRR 1.51; P=0.02 and cross-validated HRR 1.59; P=0.01, respectively). Stratification for the commonly applied systemic therapy regimens demonstrated prognostic value for EGFR and PTGS2 only in the subgroup of patients who were not treated with systemic therapy (HRR 1.78; P<0.01 and HRR 1.64; P=0.04, respectively), with worst prognosis when both EGFR and PTGS2 were highly expressed (HRR 3.08; P<0.01). Expression of PTGS2 in CRCLM was correlated to expression in patient-matched primary tumours (P=0.02, 69.2% concordance).Conclusions:EGFR and PTGS2 expressions are prognostic molecular biomarkers with added value to standard clinicopathological variables for patients with resectable CRCLM.

High-risk human papillomavirus detection in self-sampling compared to physician-taken smear in a responder population of the Dutch cervical screening: Results of the VERA study

In 2017 the cervical cancer screening program in The Netherlands will be revised. Cervical smears will primarily be tested for the presence of high-risk human papillomavirus (hrHPV) instead of cytology, and vaginal self-sampling will be offered to non-responders. This includes a potential risk that part of the women who would otherwise opt for a cervical smear will wait for self-sampling. However, self-sampling for hrHPV in a responder population has never been studied yet. The aim of this study was to investigate the applicability and accuracy of self-sampling in detecting hrHPV in a screening responder population. A total of 2049 women, aged 30-60years, participating in the screening program in The Netherlands were included from April 2013 to May 2015. After they had their cervical smear taken, women self-collected a cervicovaginal sample with a brush-based device, the Evalyn Brush. Both the cervical smear and self-sample specimen were tested with the COBAS 4800 HPV platform. The hrHPV prevalence was 8.0% (95% CI 6.9-9.2) among the physician-taken samples, and 10.0% (95% CI 8.7-11.3) among the self-samples. There was 96.8% (95% CI 96.0-97.5) concordance of hrHPV prevalence between self-samples and physician-taken samples. Women in our study evaluated self-sampling as convenient (97.1%), user-friendly (98.5%), and 62.8% preferred self-sampling over a physician-taken sampling for the next screening round. In conclusion, self-sampling showed high concordance with physician-taken sampling for hrHPV detection in a responder screening population and highly acceptable to women. Implementation of HPV-self-sampling for the responder population as a primary screening tool may be considered.