Genomic and Transcriptomic Analyses of Malignant Pleural Mesothelioma (MPM) Samples Reveal Crucial Insights for Preclinical Testing

Cell lines are extensively used to study cancer biology. However, the use of highly passaged commercial cell lines has to be questioned, as they do not closely resemble the originating tumor. To understand the reliability of preclinical models for Malignant pleural mesothelioma (MPM) studies, we have performed whole transcriptome and whole exome analyses of fresh frozen MPM tumors and compared them to cell lines generated from these tumors, as well as commercial cell lines and a preclinical MPM mouse model. Patient-derived cell lines were generated from digested fresh tumors and whole exome sequencing was performed on DNA isolated from formalin-fixed, paraffin-embedded (FFPE) tumor samples, corresponding patient-derived cell lines, and normal tissue. RNA sequencing libraries were prepared from 10 fresh frozen tumor samples, the 10 corresponding patient-derived cell lines, and 7 commercial cell lines. Our results identified alterations in tumor suppressor genes such as FBXW7, CDKN2A, CDKN2B, and MTAP, all known to drive MPM tumorigenesis. Patient-derived cell lines correlate to a high degree with their originating tumor. Gene expressions involved in multiple pathways such as EMT, apoptosis, myogenesis, and angiogenesis are upregulated in tumor samples when compared to patient-derived cell lines; however, they are downregulated in commercial cell lines compared to patient-derived cell lines, indicating significant differences between the two model systems. Our results show that the genome and transcriptome of tumors correlate to a higher degree with patient-derived cell lines rather than commercial cell lines. These results are of major relevance for the scientific community in regard to using cell lines as an appropriate model, resembling the pathway of interest to avoid misleading results for clinical applications

Tumor Immune Microenvironment and Genetic Alterations in Mesothelioma

Malignant pleural mesothelioma (MPM) is a rare and fatal disease of the pleural lining. Up to 80% of the MPM cases are linked to asbestos exposure. Even though its use has been banned in the industrialized countries, the cases continue to increase. MPM is a lethal cancer, with very little survival improvements in the last years, mirroring very limited therapeutic advances. Platinum-based chemotherapy in combination with pemetrexed and surgery are the standard of care, but prognosis is still unacceptably poor with median overall survival of approximately 12 months. The genomic landscape of MPM has been widely characterized showing a low mutational burden and the impairment of tumor suppressor genes. Among them, BAP1 and BLM are present as a germline inactivation in a small subset of patients and increases predisposition to tumorigenesis. Other studies have demonstrated a high frequency of mutations in DNA repair genes. Many therapy approaches targeting these alterations have emerged and are under evaluation in the clinic. High-throughput technologies have allowed the detection of more complex molecular events, like chromotripsis and revealed different transcriptional programs for each histological subtype. Transcriptional analysis has also paved the way to the study of tumor-infiltrating cells, thus shedding lights on the crosstalk between tumor cells and the microenvironment. The tumor microenvironment of MPM is indeed crucial for the pathogenesis and outcome of this disease; it is characterized by an inflammatory response to asbestos exposure, involving a variety of chemokines and suppressive immune cells such as M2-like macrophages and regulatory T cells. Another important feature of MPM is the dysregulation of microRNA expression, being frequently linked to cancer development and drug resistance. This review will give a detailed overview of all the above mentioned features of MPM in order to improve the understanding of this disease and the development of new therapeutic strategies

Tumor Immune Microenvironment and Genetic Alterations in Mesothelioma

Malignant pleural mesothelioma (MPM) is a rare and fatal disease of the pleural lining. Up to 80% of the MPM cases are linked to asbestos exposure. Even though its use has been banned in the industrialized countries, the cases continue to increase. MPM is a lethal cancer, with very little survival improvements in the last years, mirroring very limited therapeutic advances. Platinum-based chemotherapy in combination with pemetrexed and surgery are the standard of care, but prognosis is still unacceptably poor with median overall survival of approximately 12 months. The genomic landscape of MPM has been widely characterized showing a low mutational burden and the impairment of tumor suppressor genes. Among them, BAP1 and BLM are present as a germline inactivation in a small subset of patients and increases predisposition to tumorigenesis. Other studies have demonstrated a high frequency of mutations in DNA repair genes. Many therapy approaches targeting these alterations have emerged and are under evaluation in the clinic. High-throughput technologies have allowed the detection of more complex molecular events, like chromotripsis and revealed different transcriptional programs for each histological subtype. Transcriptional analysis has also paved the way to the study of tumor-infiltrating cells, thus shedding lights on the crosstalk between tumor cells and the microenvironment. The tumor microenvironment of MPM is indeed crucial for the pathogenesis and outcome of this disease; it is characterized by an inflammatory response to asbestos exposure, involving a variety of chemokines and suppressive immune cells such as M2-like macrophages and regulatory T cells. Another important feature of MPM is the dysregulation of microRNA expression, being frequently linked to cancer development and drug resistance. This review will give a detailed overview of all the above mentioned features of MPM in order to improve the understanding of this disease and the development of new therapeutic strategies

Genomic and Transcriptomic Analyses of Malignant Pleural Mesothelioma (MPM) Samples Reveal Crucial Insights for Preclinical Testing

Cell lines are extensively used to study cancer biology. However, the use of highly passaged commercial cell lines has to be questioned, as they do not closely resemble the originating tumor. To understand the reliability of preclinical models for Malignant pleural mesothelioma (MPM) studies, we have performed whole transcriptome and whole exome analyses of fresh frozen MPM tumors and compared them to cell lines generated from these tumors, as well as commercial cell lines and a preclinical MPM mouse model. Patient-derived cell lines were generated from digested fresh tumors and whole exome sequencing was performed on DNA isolated from formalin-fixed, paraffin-embedded (FFPE) tumor samples, corresponding patient-derived cell lines, and normal tissue. RNA sequencing libraries were prepared from 10 fresh frozen tumor samples, the 10 corresponding patient-derived cell lines, and 7 commercial cell lines. Our results identified alterations in tumor suppressor genes such as FBXW7, CDKN2A, CDKN2B, and MTAP, all known to drive MPM tumorigenesis. Patient-derived cell lines correlate to a high degree with their originating tumor. Gene expressions involved in multiple pathways such as EMT, apoptosis, myogenesis, and angiogenesis are upregulated in tumor samples when compared to patient-derived cell lines; however, they are downregulated in commercial cell lines compared to patient-derived cell lines, indicating significant differences between the two model systems. Our results show that the genome and transcriptome of tumors correlate to a higher degree with patient-derived cell lines rather than commercial cell lines. These results are of major relevance for the scientific community in regard to using cell lines as an appropriate model, resembling the pathway of interest to avoid misleading results for clinical applications.ISSN:2072-669

MicroRNA profiling predicts positive nodal status in papillary thyroid carcinoma in the preoperative setting

BACKGROUND: The molecular characterization of thyroid nodules in cytological samples has so far been focused on discriminating between benign and malignant forms in a purely diagnostic setting. The evidence on the impact of molecular biomarkers to determine the risk of aggressiveness in cytologically “neoplastic” lesions is limited to genomic alterations (such as BRAF and TERT mutations). The aim of our study was to assess the preoperative role of microRNAs (miRNAs) in predicting the nodal status of patients with papillary thyroid cancer. METHODS: A pilot series of histological samples of papillary thyroid carcinoma with (6 cases) or without (6 cases) lymph node metastases, matched for other major clinical and pathological features, was analyzed for global miRNA expression in a screening phase. A set of miRNAs was then validated in a series of 63 consecutive cytological samples of papillary carcinomas: 48 pN-negative and 15 pN-positive at histology. RESULTS: Unsupervised cluster analysis segregated surgical pN-negative and pN-positive samples, except for 1 case. The 45 differentially expressed miRNAs in pN-positive versus pN-negative cases were predicted to regulate a wide range of cellular pathways, enriched for Wnt, gonadotropin-releasing hormone receptor, and cerulein/cholecystokinin receptor signaling. In agreement with their profiles in surgical samples, 4 miRNAs of the 10 selected for validation (miR-154-3p, miR-299-5p, miR-376a-3p, and miR-302E) had a significant differential expression in cytological samples of papillary carcinoma with lymph node metastases and predicted the positive nodal status with a relatively good performance. CONCLUSIONS: MiRNA profiling is a potential promising strategy to define papillary carcinoma aggressiveness in the preoperative setting

Detection of ALK fusion variants by RNA-based NGS and clinical outcome correlation in NSCLC patients treated with ALK-TKI sequences

Introduction: Anaplastic lymphoma kinase (ALK) fusions identify a limited subset of non -small cell lung cancer (NSCLC) patients, whose therapeutic approach have been radically changed in recent years. However, diagnostic procedures and clinical-radiological responses to specific targeted therapies remain heterogeneous and intrinsically resistant or poor responder patients exist. Methods: A total of 290 patients with advanced NSCLC defined as ALK+ by immunohisto-chemistry (IHC) and/or fluorescent in situ hybridisation (FISH) test and treated with single or sequential multiple ALK inhibitors (ALKi) from 2011 to 2017 have been retrospectively retrieved from a multicentre Italian cancer network database. In 55 patients with enough left-over tumour tissue, specimens were analysed with both targeted and customised next generation sequencing panels. Identified fusion variants have been correlated with clinical outcomes. Results: Of the 55 patients, 24 received crizotinib as first-line therapy, 1 received ceritinib, while 30 received chemotherapy. Most of the patients (64%) received ALKi in sequence. An ALK fusion variant was identified in 73% of the cases, being V3 variant (E6A20) the most frequent, followed by V1 (E13A20) and more rare ones (e.g. E6A19). In three specimens, four new EML4-ALK fusion breakpoints have been reported. Neither fusion variants nor brain metastases were significantly associated with overall survival (OS), while it was predictably longer in patients receiving a sequence of ALKi. The presence of V1 variant was associated with progression-free survival (PFS) improvement when crizotinib was used (p = 0.0073), while it did not affect cumulative PFS to multiple ALKi. Conclusion: Outcomes to sequential ALKi administration were not influenced by fusion variants. Nevertheless, in V1+ patients a prolonged clinical benefit was observed. Fusion variant identification by NGS technology may add relevant information about rare chromosomal events that could be potentially correlated to worse outcomes. (C) 2022 Elsevier Ltd. All rights reserved