Live cell analyses of synaptonemal complex dynamics and chromosome movements in cultured mouse testis tubules and embryonic ovaries

During mammalian meiotic prophase, homologous chromosomes connect through the formation of the synaptonemal complex (SC). SYCP3 is a component of the lateral elements of the SC. We have generated transgenic mice expressing N- or C-terminal fluorescent-tagged SYCP3 (mCherry-SYCP3 (CSYCP) and SYCP3-mCherry (SYCPC)) to study SC dynamics and chromosome movements in vivo. Neither transgene rescued meiotic aberrations in Sycp3 knockouts, but CSYCP could form short axial element-like structures in the absence of endogenous SYCP3. On the wild-type background, both fusion proteins localized to the axes of the SC together with endogenous SYCP3, albeit with delayed initiation (from pachytene) in spermatocytes. Around 40% of CSYCP and SYCPC that accumulated on the SC was rapidly exchanging with other tagged proteins, as analyzed by fluorescent recovery after photobleaching (FRAP) assay. We used the CSYCP transgenic mice for further live cell analyses and observed synchronized bouquet configurations in living cysts of two or three zygotene oocyte nuclei expressing CSYCP, which presented cycles of telomere clustering and dissolution. Rapid chromosome movements were observed in both zygotene oocytes and pachytene spermatocytes, but rotational movements of the nucleus were more clear in oocytes. In diplotene spermatocytes, desynapsis was found to proceed in a discontinuous manner, whereby even brief chromosome re-association events were observed. Thus, this live imaging approach can be used to follow changes in the dynamic behavior of the nucleus and chromatin, in normal mice and different infertile mouse models

Assessment of a New ROS1 Immunohistochemistry Clone (SP384) for the Identification of ROS1 Rearrangements in Patients with Non–Small Cell Lung Carcinoma: the ROSING Study

Introduction: The ROS1 gene rearrangement has become an important biomarker in NSCLC. The College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology testing guidelines support the use of ROS1 immunohistochemistry (IHC) as a screening test, followed by confirmation with fluorescence in situ hybridization (FISH) or a molecular test in all positive results. We have evaluated a novel anti-ROS1 IHC antibody (SP384) in a large multicenter series to obtain real-world data. Methods: A total of 43 ROS1 FISH-positive and 193 ROS1 FISH-negative NSCLC samples were studied. All specimens were screened by using two antibodies (clone D4D6 from Cell Signaling Technology and clone SP384 from Ventana Medical Systems), and the different interpretation criteria were compared with break-apart FISH (Vysis). FISH-positive samples were also analyzed with next-generation sequencing (Oncomine Dx Target Test Panel, Thermo Fisher Scientific). Results: An H-score of 150 or higher or the presence of at least 70% of tumor cells with an intensity of staining of 2+ or higher by the SP384 clone was the optimal cutoff value (both with 93% sensitivity and 100% specificity). The D4D6 clone showed similar results, with an H-score of at least 100 (91% sensitivity and 100% specificity). ROS1 expression in normal lung was more frequent with use of the SP384 clone (p < 0.0001). The ezrin gene (EZR)-ROS1 variant was associated with membranous staining and an isolated green signal FISH pattern (p = 0.001 and p = 0.017, respectively). Conclusions: The new SP384 ROS1 IHC clone showed excellent sensitivity without compromising specificity, so it is another excellent analytical option for the proposed testing algorithm

Assessment of a New ROS1 Immunohistochemistry Clone (SP384) for the Identification of ROS1 Rearrangements in Patients with Non-Small Cell Lung Carcinoma : the ROSING Study

Altres ajuts: Funding for this study was provided bythe iLUNG Program (B2017/BMD-3884) from the Comunidad de Madrid.Introduction: The ROS1 gene rearrangement has become an important biomarker in NSCLC. The College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology testing guidelines support the use of ROS1 immunohistochemistry (IHC) as a screening test, followed by confirmation with fluorescence in situ hybridization (FISH) or a molecular test in all positive results. We have evaluated a novel anti-ROS1 IHC antibody (SP384) in a large multicenter series to obtain real-world data. Methods: A total of 43 ROS1 FISH-positive and 193 ROS1 FISH-negative NSCLC samples were studied. All specimens were screened by using two antibodies (clone D4D6 from Cell Signaling Technology and clone SP384 from Ventana Medical Systems), and the different interpretation criteria were compared with break-apart FISH (Vysis). FISH-positive samples were also analyzed with next-generation sequencing (Oncomine Dx Target Test Panel, Thermo Fisher Scientific). Results: An H-score of 150 or higher or the presence of at least 70% of tumor cells with an intensity of staining of 2+ or higher by the SP384 clone was the optimal cutoff value (both with 93% sensitivity and 100% specificity). The D4D6 clone showed similar results, with an H-score of at least 100 (91% sensitivity and 100% specificity). ROS1 expression in normal lung was more frequent with use of the SP384 clone (p < 0.0001). The ezrin gene (EZR)-ROS1 variant was associated with membranous staining and an isolated green signal FISH pattern (p = 0.001 and p = 0.017, respectively). Conclusions: The new SP384 ROS1 IHC clone showed excellent sensitivity without compromising specificity, so it is another excellent analytical option for the proposed testing algorithm

Assessment of a New ROS1 Immunohistochemistry Clone (SP384) for the Identification of ROS1 Rearrangements in Patients with Non–Small Cell Lung Carcinoma: the ROSING Study

Introduction: The ROS1 gene rearrangement has become an important biomarker in NSCLC. The College of American Pathologists/International Association for the Study of Lung Cancer/Association for Molecular Pathology testing guidelines support the use of ROS1 immunohistochemistry (IHC) as a screening test, followed by confirmation with fluorescence in situ hybridization (FISH) or a molecular test in all positive results. We have evaluated a novel anti-ROS1 IHC antibody (SP384) in a large multicenter series to obtain real-world data. Methods: A total of 43 ROS1 FISH-positive and 193 ROS1 FISH-negative NSCLC samples were studied. All specimens were screened by using two antibodies (clone D4D6 from Cell Signaling Technology and clone SP384 from Ventana Medical Systems), and the different interpretation criteria were compared with break-apart FISH (Vysis). FISH-positive samples were also analyzed with next-generation sequencing (Oncomine Dx Target Test Panel, Thermo Fisher Scientific). Results: An H-score of 150 or higher or the presence of at least 70% of tumor cells with an intensity of staining of 2+ or higher by the SP384 clone was the optimal cutoff value (both with 93% sensitivity and 100% specificity). The D4D6 clone showed similar results, with an H-score of at least 100 (91% sensitivity and 100% specificity). ROS1 expression in normal lung was more frequent with use of the SP384 clone (p < 0.0001). The ezrin gene (EZR)-ROS1 variant was associated with membranous staining and an isolated green signal FISH pattern (p = 0.001 and p = 0.017, respectively). Conclusions: The new SP384 ROS1 IHC clone showed excellent sensitivity without compromising specificity, so it is another excellent analytical option for the proposed testing algorithm

A year in the life of the EU-CardioRNA COST action: CA17129 catalysing transcriptomics research in cardiovascular disease

The EU-CardioRNA Cooperation in Science and Technology (COST) Action is a European-wide consortium established in 2018 with 31 European country members and four associate member countries to build bridges between translational researchers from academia and industry who conduct research on non-coding RNAs, cardiovascular diseases and similar research areas. EU-CardioRNA comprises four core working groups (WG1-4). In the first year since its launch, EU-CardioRNA met biannually to exchange and discuss recent findings in related fields of scientific research, with scientific sessions broadly divided up according to WG. These meetings are also an opportunity to establish interdisciplinary discussion groups, brainstorm ideas and make plans to apply for joint research grants and conduct other scientific activities, including knowledge transfer. Following its launch in Brussels in 2018, three WG meetings have taken place. The first of these in Lisbon, Portugal, the second in Istanbul, Turkey, and the most recent in Maastricht, The Netherlands. Each meeting includes a scientific session from each WG. This meeting report briefly describes the highlights and key take-home messages from each WG session in this first successful year of the EU-CardioRNA COST Action