K-ras mutations in sinonasal cancers in relation to wood dust exposure

<p>Abstract</p> <p>Background</p> <p>Cancer in the sinonasal tract is rare, but persons who have been occupationally exposed to wood dust have a substantially increased risk. It has been estimated that approximately 3.6 million workers are exposed to inhalable wood dust in EU. In previous small studies of this cancer, <it>ras </it>mutations were suggested to be related to wood dust exposure, but these studies were too limited to detect statistically significant associations.</p> <p>Methods</p> <p>We examined 174 cases of sinonasal cancer diagnosed in Denmark in the period from 1991 to 2001. To ensure uniformity, all histological diagnoses were carefully reviewed pathologically before inclusion. Paraffin embedded tumour samples from 58 adenocarcinomas, 109 squamous cell carcinomas and 7 other carcinomas were analysed for K-<it>ras </it>codon 12, 13 and 61 point mutations by restriction fragment length polymorphisms and direct sequencing. Information on occupational exposure to wood dust and to potential confounders was obtained from telephone interviews and from registry data.</p> <p>Results</p> <p>Among the patients in this study, exposure to wood dust was associated with a 21-fold increased risk of having an adenocarcinoma than a squamous cell carcinoma compared to unexposed [OR = 21.0, CI = 8.0–55.0]. K-<it>ras </it>was mutated in 13% of the adenocarcinomas (seven patients) and in 1% of squamous cell carcinomas (one patient). Of these eight mutations, five mutations were located in the codon 12. The exact sequence change of remaining three could not be identified unambiguously. Among the five identified mutations, the G→A transition was the most common, and it was present in tumour tissue from two wood dust exposed adenocarcinoma patients and one patient with unknown exposure. Previously published studies of sinonasal cancer also identify the GGT → GAT transition as the most common and often related to wood dust exposure.</p> <p>Conclusion</p> <p>Patients exposed to wood dust seemed more likely to develop adenocarcinoma compared to squamous cell carcinomas. K-<it>ras </it>mutations were detected in 13% of adenocarcinomas. In this study and previously published studies of sinonasal cancer the found K-<it>ras </it>mutations, were almost exclusively G → A transitions. In conclusion, our study, based on a large representative collection of human SNC tumours, indicates that K-<it>ras </it>mutations are relatively infrequent, and most commonly occur in adenocarcinomas. Wood dust exposure alone was not found to be explanatory for the G→A mutations, but combination of exposure to tobacco, wood dust, and possibly other occupational agents may be a more likely explanation. Overall, the study suggests a limited role for K-<it>ras </it>mutations in development of sinonasal cancer.</p

Automated Quantification Of Proliferation With Automated Hot-Spot Selection In Phosphohistone H3/Mart1 Dual-Stained Melanomas : PS02.06 | ePoster Session II

INTRODUCtIOn / BACKGROUND: Staging of melanoma includes quantification of a pro- liferation index, i.e., presumed melanocytic mitoses of H&amp;E stains are counted manually in hot spots. Yet, its reproducibility and prognostic impact increases by im- munohistochemical dual staining for phosphohistone H3 (PHH3) and MART1, which also may enable fully automated quantification by image analysis. AIMS: To ensure manageable workloads and repeatable measurements in modern pathology, the study aimed to present an automated quantification of proliferation with automated hot-spot selection in PHH3/MART1- stained melanomas. METHODS: Formalin-fixed, paraffin-embedded tissue from 153 consecutive stage I/II melanoma patients was immuno- histochemically dual-stained for PHH3 and MART1, and whole slide images were captured. An algorithm that automatically detects the number of PHH3/MART1-pos- itive cells was developed using commercial software. In preprocessing, the intensity band of the HSI color model and color deconvolution including a standard deviation filter highlighted PHH3 positivity. Threshold- ing functions classified the image into brown PHH3, red MART1, and blue hematoxylin. Finally, postprocessing al- gorithms pinpointed PHH3/MART1-positive cells based on size, MART1 surrounding, color intensity, and nuclearirregularity. Based on the labels of image analysis, a hot spot was automatically selected by a processing step where circles that detect PHH3/MART1-positive cells produce a heat map according to their cluster. The number of PHH3/MART1-positive cells was counted both automatically and manually in the global tumor area and in a manually and automatically selected hot spot, i.e., a fixed 1-mm2 square. RESULTS: The mean difference between manual and automated global counts was 2.9 cells/mm2 (P = 0.0071) and 0.23 cells per hot spot (P = 0.96) for automated counts in manually and automatically selected hot spots. In 77% of cases, manual and automated hot spots overlapped. Fully manual hot-spot counts yielded the highest prog- nostic performance with an adjusted hazard ratio of 5.5 (95% CI, 1.3–24, P = 0.024) as opposed to 1.3 (95% CI, 0.61–2.9, P = 0.47) for automated counts with automated hot spots. In conclusion, the automated index and auto- mated hot-spot selection were highly correlated to their manual counterpart, but altogether their prognostic impact was noticeably reduced. Because correct rec- ognition of only one PHH3/MART1-positive cell seems important, extremely high sensitivity and specificity of the algorithm is required for prognostic purposes. The automated analysis may thus still aid and improve the pathologists’ detection of mitoses in melanoma and possibly be useful in other malignancies and future research studies

Sentinel node imaging

Breast cancer and melanoma metastasize predominantly via the lymphatic route. It has long been known that invasion into one or a few nodes draining the primary tumour, the sentinel nodes (SN), is the most important, early sign of dissemination. If no malignant cells are detected in the SN, dissemination is unlikely to be expected. For the last 10 years SN biopsy has become an important tool in staging cancers. Two kinds of tracers are used for SN detection: The blue dye, injected during operation, and radioactively labelled colloid, injected before operation. The lymphatic drainage can then be mapped by following the blue dye by visual inspection during the operation, and with gamma camera imaging before and probe detection during the operation. The variations in the tracers used, and the injection and imaging techniques are discussed. The pathologic examination has also undergone a rapid evolution with more detailed analysis including immunohistochemistry. The use of the SN technique has quickly spread worldwide for melanoma and breast cancer but is also being tested in several other cancers. Reports on the influence on morbidity and mortality reduction are becoming increasingly convincing. The near future of SN examination is finally briefly outlined

Molecular profiles as predictive marker for the effect of overall treatment time of radiotherapy in supraglottic larynx squamous cell carcinomas.

BACKGROUND AND PURPOSE: Reduction of the overall treatment time of radiotherapy increases the probability of local tumour control, but it does not benefit all patients. Identification of molecular marker profiles may aid in the selection of patients likely to benefit from accelerated radiotherapy. PATIENTS AND METHODS: Two hundred and nine patients with SCC of the supraglottic larynx received primary radiotherapy in the randomised DAHANCA trials to 66-68 Gy, 2 Gy/fx but with different overall treatment times of 9.5 week, 6.5 week and 5.5 week. Formalin-fixed paraffin embedded tumour slides were assessed by immunohistochemistry for expression of EGFr, E-cadherin, KI-67 and Bcl-2 and the TP53 mutation profile was determined using PCR-amplification, DHPLC and sequencing. The profiles were established using a hierarchical clustering algorithm with a Bayesian information criterion for cluster number optimisation. RESULTS: Full data-set were available for 158 patients and four almost equally sized clusters were identified. One of these clusters differed significantly with respect to local control compared to the other clusters: the cluster (n=36) characterised by wild type TP53, low expression of E-cadherin and Bcl-2, moderate KI-67 and EGFr, was not influenced by a reduction in the overall treatment time (P=0.6) whereas the other clusters showed an increase in local control when the overall treatment time of radiotherapy was reduced. This was also partially seen with disease specific survival as the endpoint. CONCLUSIONS: Molecular marker profiling may aid in the selection of patients that will benefit of a reduction in overall treatment time of radiotherapy in SCC of the supraglottic larynx