RESUMO
AIM: Testing for ALK rearrangements in advanced, non-squamous non-small-cell lung cancers that are wild-type for activating EGFR mutation has become standard care. Fluorescence in-situ hybridization is considered the gold standard for this evaluation. Pre-screening with immunohistochemistry has been suggested, to reduce testing costs and to make testing more widely available. By analysing the sensitivity and specificity of different ALK immunohistochemical assays, we aimed to identify the most reliable assay to detect ALK rearrangement. METHODS AND RESULTS: ALK screening performed by FISH analysis was compared with three different immunohistochemical assays, in which two ALK antibody clones (5A4 and D5F3) were used on two detection platforms (Dako AutostainerLink 48 and Ventana Benchmark GX). Data from 30 ALK FISH-positive cases show that the sensitivity of the immunohistochemical assays varies from 93.3% to 96.6%. Head-to-head comparison of the 5A4 and D5F3 ALK antibody clones demonstrates similar staining potency. In general, homogeneous, intermediate to strong staining of the ALK-positive samples was obtained. CONCLUSIONS: ALK immunohistochemistry can be considered as a pre-screen method if one accepts a sensitivity of 93.3-96.6%. Because ALK immunohistochemical staining needs to be performed close to the detection limit of the assay, vigilant quality control monitoring is required to guarantee trustworthy results.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Rearranjo Gênico , Imuno-Histoquímica/métodos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Quinase do Linfoma Anaplásico , Humanos , Hibridização in Situ Fluorescente/métodos , Receptores Proteína Tirosina Quinases/imunologia , Translocação GenéticaRESUMO
OBJECTIVE: The purpose of this study was to determine the significance of the intracellular NPxYxxL motif of LRP1 for the atheroprotective role of this multifunctional receptor. METHODS AND RESULTS: LRP1 knock-in mice carrying an inactivating mutation in the NPxYxxL motif were crossed with LDLR-deficient mice, a model for atherosclerosis. In this LDLR(-/-) background the mutated mice showed a more atherogenic lipoprotein profile, which was associated with a decreased clearance of postprandial lipids because of a compromised endocytosis rate and reduced lipase activity. On an atherogenic diet LRP1 mutant mice revealed a 50% increased development of atherosclerosis. This aggravation was accompanied by an increase in smooth muscle cell (SMC) and collagen content and apoptotic cells in the lesions. The mutation showed, however, a limited impact on basal PDGFR-beta expression and signaling and the antimigratory property of apoE on PDGF-BB-stimulated SMCs. Additionally, levels of LRP1 atherogenic ligands, like MMP2, t-PA, FVIII, and the inflammatory ligand TNF-alpha showed to be significantly elevated. CONCLUSIONS: These findings demonstrate that the NPxYxxL motif is essential for the atheroprotective role of LRP1. This motif is relevant for normal control of lipid metabolism and of atherogenic and inflammatory ligands, but has no pronounced effect on regulating PDGF-BB/PDGFR-beta signaling in SMCs.