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1600 Somatic and Germline Cohesin Genes Alterations in Pediatric Acute Lymphoblastic Leukemia

Program: Oral and Poster Abstracts
Session: 618. Acute Lymphoblastic Leukemias: Biomarkers, Molecular Markers and Minimal Residual Disease in Diagnosis and Prognosis: Poster I
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Lymphoid Leukemias, ALL, Translational Research, genomics, bioinformatics, hematopoiesis, blood banking, Diseases, cell expansion, Lymphoid Malignancies, computational biology, Biological Processes, Technology and Procedures, gene editing, pathogenesis, omics technologies
Saturday, December 9, 2023, 5:30 PM-7:30 PM

Stefano Rebellato1*, Claudia Saitta1*, Laura Rachele Bettini, MD1*, Daniela Silvestri1*, Orietta Spinelli2*, Agata Pastorczak, MD, PhD3*, Danielle Brandes4*, Ute Fischer, PhD4*, Arndt Borkhardt5, Sabine Strehl, PhD6*, Luca Lo Nigro, MD7, Barbara Buldini, MD, PhD8, Julia Hauer9*, Franziska Auer, PhD10*, Lennart Kester11*, Roland P. Kuiper, PhD11,12, Luciana Vinti, MD, PhD13, Franco Locatelli, MD, PhD13,14, Andrea Biondi, MD1,15, Grazia Fazio, PhD1* and Giovanni Cazzaniga, PhD1,16

1Tettamanti Center, Fondazione IRCCS San Gerardo dei Tintori, Monza, Italy
2Hematology and Bone Marrow Transplant Unit, ASST Papa Giovanni XXIII, Bergamo, Italy
3Department of Pediatrics, Oncology and Hematology, Medical University of Lodz, Lodz, Poland
4Department of Pediatric Oncology, Hematology and Clinical Immunology, Heinrich Heine University, Medical Faculty, Duesseldorf, Germany
5Department of Pediatric Oncology, Hematology and Clinical Immunology, Medical faculty, Heinrich Heine University Düsseldorf, Duesseldorf, Germany
6St. Anna Children’s Cancer Research Institute, Vienna, Austria
7Cytogenetic-Cytofluorimetric-Molecular Biology Laboratory Center of Pediatric Hematology Oncology, Azienda Policlinico "G. Rodolico",, Catania, Italy
8Department of Pediatric Hematology and Oncology, University-Hospital of Padova, Padova, Italy
9Technical University of Munich, School of Medicine, Department of Pediatrics, Munich, Germany
10Technical University of Munich, School of Medicine, Department of Pediatrics, Neufahrn Bei Freising, Bavaria, Germany
11Princess Maxima Center for Pediatric Oncology, Utrecht, Netherlands
12Department of Genetics, Utrecht University Medical Center, Utrecht, Netherlands
13IRCCS Ospedale Pediatrico Bambino Gesù, Roma, Italy
14Catholic University of the Sacred Heart, Roma, Italy
15Pediatrics, Fondazione IRCCS San Gerardo dei Tintori, Monza, Italy
16Medical Genetics, Università di Milano Bicocca, Monza, Italy

Leukemia is a complex disease and the molecular mechanisms of malignant transformation are not fully understood yet. Cohesins are essential proteins, forming a complex which plays critical roles in various cellular processes, not only by canonical DNA binding and chromosome segregation but also in non-canonical regulation of gene expression in both proliferating and post-mitotic cells.

Mutations or gene expression alterations of genes encoding the components of the cohesins’ complex may affect chromosomal stability and potentially contribute to the pathogenesis of leukemia. Somatic cohesins mutations have been already reported to occur in myeloproliferative disorders and pediatric Acute Lymphoblastic Leukemia (ALL), while on the other hand, germline mutations are causative of Cohesinopathies such as Cornelia de Lange Syndrome (CdLS). Our previous first report of a patient affected by both CdLS and ALL suggested a potential involvement of cohesin germline mutations in the pathogenesis of ALL. Moreover, we demonstrated that germline mutations in STAG1 gene could play a crucial role in predisposition to pediatric hematological malignancies, including ALL. These findings suggest that both somatic and germline alterations of cohesin genes may be associated with the development of ALL.

This study aims to understand the molecular mechanisms affected by Cohesin genes alterations, that could potentially pave the way for the development of targeted therapies and improved treatment strategies for leukemia patients.

By employing a DNA-targeted next-generation sequencing (NGS) screening of a cohort of 120 consecutive pediatric ALL cases (enrolled in the AIEOP-BFM protocol at various Italian centers), we discovered a total of 11 germline cohesin mutations, further categorized as follows: 6 out of 11 were missense, 4 out of 11 were located in the 3' or 5' untranslated regions (UTR) and 1 out of 11 was a splice-acceptor variant. To investigate the functional implications of selected germline mutations in STAG1 gene, we took advantage of the CRISPR/Cas9 technique to generate in vitro models of two Lymphoblastoid Cell Lines (LCLs) harboring specific variants already found in two patients of our cohort. Additionally, we established control LCLs from healthy donors' samples using the same CRISPR/Cas9 approach. Subsequently, we performed RNA sequencing (RNAseq) analysis on these cell lines, and our preliminary findings revealed a total of 619 differentially expressed genes (DEGs) compared to the control group. Many of them are involved in critical intracellular pathways, like ribosomal transcription, suggesting potential disruptions of essential cellular processes caused by germline STAG1 mutations.

Moreover, we have successfully developed an advanced computational analysis pipeline with the primary objective of detecting fusion transcripts from RNAseq data. This pipeline was applied to a dataset of 529 consecutive diagnose and 95 relapse samples of pediatric B/T-ALL patients and in five B-ALL cases a fusion transcript involving the STAG1 (n=1), STAG2 (n=3), and NIPBL (n=1) genes was identified. The significance of these findings was further reinforced by additional cases from both pediatric and adult cohorts through an international collaborative effort and five more fusion transcripts of STAG2 (n=2) and NIPBL (n=3) were uncovered. Investigation to clarify the underlying functional consequences of these fusion genes in pediatric ALL are ongoing.

Collectively, these observations endeavor to elucidate the role of germline and somatic cohesin events in pediatric ALL. In particular, the investigation seeks to unveil the non-canonical functions of these proteins, thus paving the way for the identification and characterization of a novel and distinct subcategory of leukemia within the pediatric population.

Disclosures: Lo Nigro: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceutical: Consultancy, Membership on an entity's Board of Directors or advisory committees; Clinigen: Consultancy, Membership on an entity's Board of Directors or advisory committees. Vinti: Amgen / Clinigen: Consultancy; Amgen / Nequil / Takeda: Other: Travel/Accommodations/Expenses; Merck Sharp & Dohme: Research Funding; Amgen / Takeda: Speakers Bureau. Biondi: Colmmune: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Membership on an entity's Board of Directors or advisory committees; Novartis: Speakers Bureau; Agmen: Speakers Bureau; Galapagos: Membership on an entity's Board of Directors or advisory committees.

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