Coupling imaging mass cytometry with Alcian blue histochemical staining for a single-slide approach.

Imaging mass cytometry (IMC) is a metal mass spectrometry-based method allowing highly multiplex immunophenotyping of cells within tissue samples. However, some limitations of IMC are its 1-µm resolution and its time and costs of analysis limiting respectively the detailed histopathological analysis of IMC-produced images and its application to small selected tissue regions of interest (ROI) of one to few square millimeters. Coupling on a single-tissue section, IMC and histopathological analyses could permit a better selection of the ROI for IMC analysis as well as co-analysis of immunophenotyping and histopathological data until the single-cell level. The development of this method is the aim of the present study in which we point to the feasibility of applying the IMC process to tissue sections previously Alcian blue-stained and digitalized before IMC tissue destructive analyses. This method could help to improve the process of IMC in terms of ROI selection, time of analysis, and the confrontation between histopathological and immunophenotypic data of cells.

HER2 Mutated and Nonmutated Non-small Cell Lung Carcinomas Can Harbor Heterogeneous HER2 Gene Amplification and HER2 Protein Expression.

Molecular analyses have become mandatory for treatment choices in patients with advanced non-small cell lung cancers (NSCLC). Among them, HER2 gene mutation, HER2 gene amplification, and HER2 protein expression consist in potential targets of various treatments. Tumor heterogeneity and overlapping of molecular alterations may cause dilemmas in treatment choices but to date there are few that reported about HER2 with discrepant data. We led a retrospective study evaluating HER2 protein expression and HER2 gene/chromosome 17 copy number variations across different tumor areas and samples from patients with advanced NSCLC harboring HER2 gene mutations and other oncogenic mutations. Among patients with HER2-mutated (10 patients) and nonmutated lung adenocarcinomas (10 patients), we observed frequent heterogeneous HER2 protein expression with no correlation with HER2 gene copy number variations. HER2 gene amplification was observed in 6 patients (3 HER2-mutated and 3 HER2-nonmutated), but with intrasample heterogeneity in 2 cases and intersample heterogeneity in another case. Our small case series emphasizes the potential overlapping and spatial heterogeneity of HER2 alterations in NSCLC, which must be taken into account as a limitation in building predictive strategies accompanying the development of anti-HER2 therapeutic strategies in patients with advanced NSCLC.

Fluorescent In Situ Hybridization Must be Preferred to pan-TRK Immunohistochemistry to Diagnose NTRK3-rearranged Gastrointestinal Stromal Tumors (GIST).

Tyrosine kinase inhibitors have revolutionized the treatment of patients with gastrointestinal stromal tumors (GISTs). Nevertheless, some GISTs do not contain any targetable KIT or PDGFRA mutations classically encountered in this field. Novel approved therapies targeting TRK chimeric proteins products of NTRK genes fusions consist in a promising approach to treat some patients with GISTs lacking any identified driver oncogenic mutation in KIT, PDGFRA or BRAF genes. Thus, an adequate testing strategy permitting to diagnose the rare NTRK-rearranged GISTs is required. In this work, we studied about the performances of pan-TRK immunohistochemistry (IHC) and NTRK1/2/3 fluorescent in situ hybridization in a series of 39 GISTs samples. Among 22 patients with GISTs lacking KIT or PDGFRA mutations, BRAFV600E IHC permitted to diagnose 2/22 (9%) BRAFV600E-mutated GISTs and, among the 20 KIT, PDGFRA, and BRAF wild type tumors, 1/20 (5%), NTRK3-rearranged tumor was diagnosed using NTRK3 fluorescent in situ hybridization. Pan-TRK IHC using EPR17341 and A7H6R clones was negative in this NTRK3-rearranged sample. Pan-TRK IHC was frequently positive in NTRK not rearranged tumors without (24 samples analyzed) or with (15 samples analyzed) KIT or PDGFRA mutations with major discrepancies between the 2 IHC clones (intraclass correlation coefficient of 0.3042). Given the new therapeutic opportunity offered by anti-TRK targeted therapies to treat patients with advanced cancers including GISTs, it is worth to extend molecular analysis to NTRK fusions testing in KIT, PDGFRA, and BRAF wild type GISTs. Pan-TRK IHC appears not relevant in this field but performing a simple NTRK3 fluorescent in situ hybridization test consists in a valuable approach to identify the rare NTRK3-rearranged GISTs treatable using anti-TRK therapies.