New Methods for ALK Status Diagnosis in Non–Small-Cell Lung Cancer: An Improved ALK Immunohistochemical Assay and a New, Brightfield, Dual ALK IHC–In Situ Hybridization Assay

Introduction:The demonstration of anaplastic lymphoma kinase (ALK) positivity in non–small-cell lung cancer (NSCLC) has been hindered by the technical complexity and interpretative challenges of fluorescence in situ hybridization methods for detection of ALK gene rearrangement and by the inadequate sensitivity of existing immunohistochemistry (IHC) methods for ALK protein detection. In this study, we sought to increase the sensitivity of ALK IHC detection and to develop a brightfield assay for concurrent detection of ALK protein expression and ALK gene rearrangement.Methods:We developed a horseradish peroxidase–based IHC detection system using the novel, nonendogenous hapten 3-hydroxy-2-quinoxaline (HQ) and tyramide. We also developed a dual gene protein ALK assay combining a brightfield break-apart in situ hybridization ALK assay with another sensitive IHC method using the novel, nonendogenous hapten 5-nitro-3-pyrazole. We examined the sensitivity and accuracy of these methods using surgically resected NSCLC cases examined with ALK fluorescence in situ hybridization.Results:The new HQ-tyramide IHC detection system offered readily interpretable staining with substantially greater sensitivity than conventional ALK IHC, and produced heterogeneous and homogeneous patterns of ALK protein staining among ALK-positive NSCLC surgical cases. The new 5-nitro-3-pyrazole–based IHC detection system was similar in ALK detection sensitivity to the HQ-tyramide IHC system and was compatible with the brightfield in situ hybridization assay.Conclusion:The new HQ-tyramide IHC reagent system allows more sensitive assessment of ALK protein status in NSCLC cases. The new ALK gene-protein assay allows the concurrent visualization of ALK gene and ALK protein status in single cells, allowing more accurate ALK status determination even in heterogeneous specimens

Optical properties–microstructure–texture relationships of dried apple slices: Spatially resolved diffuse reflectance spectroscopy as a novel technique for analysis and process control

The potential of spatially resolved diffuse reflectance spectroscopy in the 500–1000 nm range by means of a fiber-optics probe was investigated for acquiring scattering and absorption properties of air dried apple rings subjected to different pre-treatment conditions: without osmo-dehydration (TQ) and with osmo-dehydration for 1 (OSMO1) and 3 h (OSMO2). The fresh apple rings were produced from ‘Golden Delicious’ apples at harvest (H) and 5 month storage at 2 conditions: controlled atmosphere (CA) and normal atmosphere (NA). Microstructure properties of the dried apple rings were also obtained from X-ray micro-CT measurements. The TQ samples were found to have significantly higher scattering properties, thicker tissue, smaller pore sizes, were less crispy, and required higher snapping work or rupture energy than the OSMO1 and OSMO2 samples. On the other hand, no significant differences were observed between the scattering properties, microstructure, and textural quality of the OSMO1 and OSMO2 apple rings. From these results, it was concluded that there is a clear process–microstructure–quality relation in osmo-air-dried apples which can be measured non-destructively with spatially resolved diffuse reflectance spectroscopy. Therefore, this study confirmed the potential of spatially resolved diffuse reflectance spectroscopy for non-destructive quality assessment of air-dried apple slices, which provides perspectives for drying process optimization

Comprehensive genomic profiles of small cell lung cancer

We have sequenced the genomes of 110 small cell lung cancers (SCLC), one of the deadliest human cancers. In nearly all the tumours analysed we found bi-allelic inactivation of TP53 and RB1, sometimes by complex genomic rearrangements. Two tumours with wild-type RB1 had evidence of chromothripsis leading to overexpression of cyclin D1 (encoded by the CCND1 gene), revealing an alternative mechanism of Rb1 deregulation. Thus, loss of the tumour suppressors TP53 and RB1 is obligatory in SCLC. We discovered somatic genomic rearrangements of TP73 that create an oncogenic version of this gene, TP73Dex2/3. In rare cases, SCLC tumours exhibited kinase gene mutations, providing a possible therapeutic opportunity for individual patients. Finally, we observed inactivating mutations in NOTCH family genes in 25% of human SCLC. Accordingly, activation of Notch signalling in a pre-clinical SCLC mouse model strikingly reduced the number of tumours and extended the survival of the mutant mice. Furthermore, neuroendocrine gene expression was abrogated by Notch activity in SCLC cells. This first comprehensive study of somatic genome alterations in SCLC uncovers several key biological processes and identifies candidate therapeutic targets in this highly lethal form of cancer

Prediction of optimal boiling time for potatoes by hyperspectral imaging and heat transfer model

Cooking of potatoes causes changes in the microstructure and composition of starch. These changes affect the interaction of light with the starch granules at different regions inside the potato. In this research, the potential of hyperspectral imaging in combination with chemometric tools for contactless detection of the cooking front in potatoes has been investigated. Boiled potatoes cooked for 0 to 30 minutes were cut into halves and their cut surfaces were scanned by a hyperspectral camera in the wavelength range 400–1000 nm. Partial least squares discriminant analysis (PLSDA) was employed to discriminate between the pixel spectra for the cooked regions and those for the remaining raw regions. From each of the resulting images with detected cooking fronts, the relative area, which is the ratio of the remaining raw part area over the total potato area, was calculated. Then a heat transfer model was applied to plot the relative area against the cooking time and the optimal cooking time was defined as the time when the relative area became zero. It was found that the optimal cooking time could be precisely predicted by measuring one potato sample without cooking and another cooked for 12 to 21 minutes. The results illustrate the potential of hyperspectral imaging as a process monitoring tool for the potato processing industry.status: publishe

Bright-Field Dual-Color Chromogenic In Situ Hybridization for Diagnosing Echinoderm Microtubule-Associated Protein-Like 4-Anaplastic Lymphoma Kinase-Positive Lung Adenocarcinomas

IntroductionA subset of lung cancers harbors an EML4-ALK (echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase) gene fusion, and detecting this subset may hold therapeutic implications. Many prior studies used fluorescence in situ hybridization (FISH) analysis for this detection, but FISH may have disadvantages including signal decay and dark-field examination that may obscure tissue architecture. In this study, we explored the potential of the ALK-break-apart chromogenic in situ hybridization (CISH) method to detect ALK-rearranged lung cancer.MethodsWe examined 15 lung adenocarcinomas with reverse-transcriptase polymerase chain reaction-proven EML4-ALK fusion transcripts and 30 ALK-negative cases. One hundred tumor cells were evaluated by CISH and FISH for each case, and a detailed signal profile was recorded and compared.ResultsCISH preserved tissue architecture and cytomorphology considerably and facilitated the signal evaluation using a routine light microscope. Positive rearrangement signals (splits or isolated 3′ signals) were identified in 13 to 78% (mean ± SD, 41% ± 19%) of tumor cells in the ALK-positive cohort and in 0 to 15% (mean ± SD, 6% ± 4%) of cells in the ALK-negative cohort. The two groups were best separated by a cutoff value of 20%, with a sensitivity of 93% and a specificity of 100%. The only false-negative tumor having only 13% CISH-positive cells displayed predominantly (76%) isolated 5′ signals unaccompanied by 3′ signals. FISH showed largely similar signal profiles, and the results were completely concordant with CISH.ConclusionsWe have successfully introduced CISH for diagnosing EML4-ALK-positive lung adenocarcinoma. This method allows simultaneous visualization of genetics and tumor cytomorphology and facilitates the molecular evaluation and could be applicable in clinical practice to detect lung cancer that may be responsive to ALK inhibitors