ECP 2023 Abstracts

S131 Virchows Archiv (2023) 483 (Suppl 1):S1–S391 13 origin, both sensitivity and specificity is higher for GAD2 than for PR. The combination of PR and GAD2 further increases sensitivity and specificity. PS-23-015 RNF135 promoter methylation as a potential diagnostic bio- marker distinguishing hepatocellular carcinoma from other adenocarcinomas B. Ranković*, T. Draškovič, N. Zidar, N. Hauptman *Institute of Pathology, Slovenia Background & objectives: Differentiation of hepatocellular carcinoma (HCC) from liver metastases can be challenging. We investigated the methylation status of the promoter region of RING finger protein 135 (RNF135) and its significance as DNA methylation biomarker in dif- ferent adenocarcinomas. Methods: DNA was isolated from formalin-fixed paraffin-embedded tissue samples from 14 HCCs, 12 cholangiocarcinomas, 15 pancreatic adenocarcinomas, 6 stomach adenocarcinomas, 7 colorectal cancers and their paired normal tissue samples. The methylation status of RNF135 promoter region was determined by methylation-sensitive high-resolution melting analysis. The methylation status was used to determine sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV). Results: Our results show that RNF135 promoter region was hyper- metylated in HCC and hypomethylated in other investigated adenocar- cinomas and their paired adjacent normal samples. Methylation status of RNF135 promoter region successfully discriminates between HCC and its adjacent normal liver tissue with a sensitivity of 77%, specific- ity of 93%, PPV of 0.91 and NPV 0.81. Furthermore, this methylation biomarker can differentiate between HCC and investigated adenocarci- nomas with a sensitivity of 77%, specificity of 100%, PPV 1, and NPV 0.93. When testing all tumour and normal samples, methylation status of RNF135 promoter showed an overall sensitivity of 77%, specificity of 99%, PPV of 0.91, and NPV of 0.97 for HCC. Conclusion: DNA methylation in promoter region of gene RNF135 showed high sensitivity, specificity, PV and NPV for discriminating HCC from other adenocarcinomas and normal samples. PS-23-016 TIMP1 as venous invasion marker in pancreatic ductal adenocarcinoma Y. Sung*, M. Kim, S. Hong *Republic of Korea Background & objectives: Venous invasion is known to be the cause of poor prognosis in pancreatic ductal adenocarcinoma (PDAC), but the exact mechanism is still not well elucidated. In this study, gene expression array was performed and biomarkers of venous invasion were investigated. Methods: To select venous invasion specific genes, Nanostring nCoun- ter analysis was performed on the following groups: 1) vein with cancer invasion; 2) cancer without vein invasion; and 3) normal vein. The selected genes were validated by immunohistochemical staining in protein level. The role of potential biomarker in venous invasion was investigated by invasion assay and western blot analysis. Results: Four genes (CXCR4, TIMP1, OLFML2B, CYP1B1) were spe- cifically high expressed in vein with cancer invasion group. Among them, high protein expression of CXCR4 and TIMP1 were validated by immu- nohistochemical staining and in particular, high TIMP1 expression was confirmed by 3D image in venous invasion areas. TIMP1-expression group was related to lymphovascular invasion (p < 0.001) and low 5 year-survival rate (p = 0.032). Furthermore, TIMP1 inhibition by siRNA reduced cancer cell invasion ability in the presence of cancer-associated fibroblast (CAF). In co-culture condition, TIMP1 was increased along with PI3Kp110 and phospho-AKT in pancreatic cancer cells. Therefore, TIMP1 in pancreatic cancer cells may induce venous invasion through PI3K/AKT pathway. Conclusion: This study discovered TIMP1 as a biomarker of venous invasion of pancreatic cancer and revealed that TIMP1/PI3K/AKT pathway affect venous invasion of pancreatic cancer. PS-24 | Poster Session Molecular Pathology PS-24-001 A real-world experience from a single centre (LPCE, Nice, France) highlights the urgent need to abandon immunohistochemistry for ROS1 rearrangement screening of advanced non-squamous non- small cell lung cancer C. Bontoux*, V. Hofman, S. Goffinet, E. Long-Mira, S. Lassalle, M. Ilié, P. Hofman *Laboratory of Clinical and Experimental Pathology, Hospital-Integrated Biobank (BB- 0033-00025), Pasteur Hospital, IRCAN Team 4, Inserm U1081, CNRS 7284, FHUOncoAge, Université Côte d’Azur, Nice, France Background & objectives: Detection of ROS1 rearrangements in metastatic NSCLC is mostly based on a testing algorithm associ- ating ROS1 immunohistochemistry (IHC) screening followed by mandatory ROS1 FISH and/or next generation sequencing (NGS) analyses in the case of IHC positivity to confirm the results. Methods: We here evaluated RNA NGS used as reflex testing for ROS1 rearrangement assessment in NS-NSCLC with the aim of replacing ROS1 IHC as a screening method. ROS1 IHC and RNA and DNA NGS were prospectively performed from January 2021 to Janu- ary 2023 for 810 NS-NSCLC. Positive results were additionally ana- lysed by ROS1 FISH. The turnaround times (TATs) were compared. Results: ROS1 IHC was positive in 26/810 (3.2%) cases that showed variable staining intensity while NGS detected ROS1 rear- rangements in 12/810 (1.5%) cases. ROS1 FISH was positive in 14/26 (54%) of ROS1 IHC positive cases and in all positive ROS1 NGS cases. Obtaining both ROS1 IHC and ROS1 FISH reports took an average of 6 days, while obtaining ROS1 IHC, DNA, and RNA NGS reports took an average of 3 days. Conclusion: Taken together, these results showed that systematic screening for the ROS1 status using IHC must be replaced by NGS reflex testing in cases of metastatic NS-NSCLC. PS-24-002 Pulmonary Adenocarcinoma: ALK, ROS1 and RET imbalance sequencing concomitant with other target mutations L. Carvalho*, M.R. Silva, A. Alarcão, A.F. Ladeirinha, T. Ferreira, C. Eliseu, M. Viseu, V. Almeida, G. Fontinha, V. Sousa *Institute of Anatomical and Molecular Pathology, Faculty of Medicine of the University of Coimbra, Coimbra, Portugal, CIMAGO – Research Center for Environment, Genetics and Oncobiology, Faculty of Medi- cine, University of Coimbra, Portugal; Pathology Service, University Hospital Anatomical Pathology Coimbra, Portugal Background & objectives: Gene fusions have significant prognostic and predictive value being screened as part of molecular pathology test- ing for patient management. Different approaches have been developed to detect fusion, in next generation sequencing, 3’/5’ imbalance value can evaluate novel fusions for diagnosis. Methods: Oncomine™ Precision Assay Panel workflow applied to fusion detection using expression “imbalance” in Oncomine Reporter™ Software, generates 3’/5’ imbalance value, reporting the difference in expression between 5’ assay and 3’ assay of each driver gene ALK, ROS1 and RET. Fluorescence in situ hybridization (FISH) and Immunohistochemistry (IHC) were applied to 3’/5’ imbalance cases to confirm these targets in pulmonary adenocarcinomas.