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2960 Non-Coding Mutations of BCL6 in Classical Hodgkin Lymphoma

Program: Oral and Poster Abstracts
Session: 621. Lymphomas: Translational – Molecular and Genetic: Poster II
Hematology Disease Topics & Pathways:
Research, Translational Research
Sunday, December 8, 2024, 6:00 PM-8:00 PM

Maria Cristina Pirosa, MD1, Matin Salehi1*, Alessio Bruscaggin, PhD1*, Lodovico Terzi Di Bergamo1*, Federico Jauk, MD1*, Gabriela Forestieri1*, Simone Bocchetta, PhD1*, Deborah Piffaretti, PhD2*, Riccardo Moia, MD3*, Vanessa Cristaldi, PhD4*, Martina di Trani5*, Georgia Alice Galimberti1*, Katia Pini1*, Valeria Spina6*, Claudia Giordano, MD7*, Adalgisa Condoluci, MD1, Salvatore Annunziata8*, Fabrizio Bergesio9*, Renzo Boldorini10*, Eugenio Borsatti11*, Pietro Bulian12*, Stephane Chauvie, PhD9*, Marco Cuzzocrea13*, Bernhard Gerber14*, Michal Kurlapski15*, Luigi Maria Larocca16*, Andrea Rinaldi17*, Marcello Rodari18*, Grzegorz Romanowicz19*, Gian Mauro Sacchetti20*, Anastasios Stathis, MD14*, Georg Stüssi, MD14*, Ilaria Zangrilli21*, Eleonora Calabretta5*, Francesco Corrado, MD5*, Antonello Pinto22*, Luca Mazzucchelli23*, Valter Gattei, MD24, Jan Maciej Zaucha15*, Armando Santoro, MD25*, Stefan Hohaus21*, Franco Cavalli, MD26*, Emanuele Zucca, MD14, Gianluca Gaidano, MD, PhD3, Carmelo Carlo-Stella, MD5, Alexandar Tzankov, MD, PhD27*, Luca Ceriani, MD13* and Davide Rossi, MD, PhD1

1Institute of Oncology Research, Laboratory of Experimental Hematology, Bellinzona, Switzerland
2Institute of Oncology Research, Laboratory of Experimental Hematology, Bellinzona, AL, Switzerland
3Division of Hematology, Department of Translational Medicine, Università del Piemonte Orientale, Novara, Italy
4Department of Biomedical Sciences, University of Humanitas, Milan, Italy
5Department of Biomedical Sciences, Humanitas University, Milan, Italy
6Laboratorio di Diagnostica Molecolare, Servizio di Genetica Medica EOLAB, Ente Ospedaliero Cantonale, Bellinzona, Switzerland
7Department of Clinical Medicine and Surgery, Federico II University Medical School, Naples, ITA
8UOC Medicina Nucleare, GSTeP Radiopharmacy TracerGLab, Fondazione Policlinico Universitario A. Gemelli IRCCS, Rome, Italy
9Medical Physics Division, AO Santa Croce e Carle, Cuneo, Italy
10Division of pathology, University of Eastern Piedmont, Novara, Italy
11Nuclear Medicine, Centro di Riferimento Oncologico, Aviano, Italy
12Clinical and Experimental Onco-Hematology Unit, Centro di Riferimento Oncologico, Aviano, Italy
13Imaging Institute of Southern Switzerland, Clinic of Nuclear Medicine and Molecular Imaging, Bellinzona, Switzerland
14Oncology Institute of Southern Switzerland (IOSI), Ente Ospedaliero Cantonale, Bellinzona, Switzerland
15Department of Hematology and Bone Marrow Transplantation, Medical University of Gdańsk, Gdańsk, Poland
16Fondazione Policlinico Universitario A. Gemelli. IRCCS, Rome, ITA
17Genomics Facility, Institute of Oncology Research, Bellinzona, Switzerland, Bellinzona, Switzerland
18Nuclear Medicine Unit, IRCCS Humanitas Research Hospital, Rozzano, Italy
19Department of Nuclear Medicine, Medical University of Gdańsk, Gdańsk, Poland
20Divsion of Nuclear Medicine, AOU Maggiore della Carità, Novara, ITA
21Sezione di Ematologia, Dipartimento di Scienze Radiologiche ed Ematologiche, Università Cattolica del Sacro Cuore, Rome, Italy
22Hematology-Oncology and Stem Cell Transplantation Unit, National Cancer Institut, Napoli, ITA
23Division of Pathology, Ente Ospedaliero Cantonale, Bellinzona, Switzerland
24Clinical and Experimental Onco-Hematology Unit, Centro di Riferimento Oncologico di Aviano (CRO) IRCCS, Aviano, Italy
25IRCCS Humanitas Research Hospital, Rozzano, Italy
26Fondazione per l’Istituto Oncologico di Ricerca, Bellinzona, Switzerland
27Institute for Pathology, University Hospital Basel, Basel, Switzerland

Introduction: Non-coding regions, constituting a substantial part of the human genome, are crucial for gene regulation. In cancer, somatic non-coding regulatory mutations can significantly alter gene expression by either disrupting or creating transcription factor-binding sites. This phenomenon is notably well-known in diffuse large B-cell lymphoma (DLBCL), where activation-induced cytidine deaminase (AID) misfires during the germinal center (GC) reaction, leading to numerous non-coding mutations that disrupt gene regulation and contribute to malignancy. Classical Hodgkin lymphoma (cHL) is the most prevalent lymphoma among young adults. Although cHL originates from GC B cells, research into AID-associated hypermutation affecting non-coding regulatory regions has not yet been carried out.

Methods: We investigated somatic mutations in 155 coding genes and thirty-three genomic regions (mostly non-coding) known to be foci of AID-associated hypermutation and contained within regulatory regions of genes involved in GC B cell biology and transformation. ctDNA was analyzed using LyV4.0 ctDNA CAPP-seq in 297 cHL patients from the IOSI-EMA003-NCT03280394 and FIL-RougeBIO-NCT05066555 studies, and in 235 DLBCL for comparison.

Results: Mutations of in at least one of region of AID-associated hypermutation was detected in 83% of cHL, a prevalence comparable to that observed in DLBCL (88%). The region of AID-associated hypermutation within the BCL6 locus was the most frequently mutated, found in 56% of cHL cases. Other hypermutated regions occurring in >10% of patients mapped in the SOCS1, CD83, RHOH, ST6GAL1, PAX5, BTG1, BCL7A, LRMP and HIST1H1E loci. We then scanned the sequenced non-coding spaces to detect recurrently mutated hotspots. This analysis involved evaluating overlapping 20 bp intervals through a sliding window of one bp while accounting for the nucleotide context. A hotspot located at chr3:187462671-187462703 of BCL6-intron 1 (designated as BCL6 Hotspot 7) was mutated in 30% of cases, a proportion that closely resembled that of DLBCL patients where it is a validated somatic expression quantitative trait locus (seQTL). BCL6 Hotspot 7 aligned with an area of accessible chromatin and heightened H3K27 acetylation in cHL, which was also nominated a super enhancer in 33% cHL cell lines. Moreover, ChIP-seq analysis of BCL6 Hotspot 7 revealed increase in H3K27ac in mutant cHL cell lines that suggested differential binding of transcription factors. BCL6 Hotspot 7 sequence predicted to bind the PRDM1 transcriptional repressor of BCL6, and mutations therein were found to impede BLIMP1 binding in cHL cells. Notably, BCL6 Hotspot 7 mutations were found to co-occur with BCL6 expression in cHL cell lines and HRS cells of primary biopsies, despite the co-expression of PRDM1. BCL6 protein expression ranging from weak to strong was detected in the nucleus of HRS cells of 68% of primary biopsies. The core set of genes that are directly bound and regulated by BCL6 exhibited similar expression levels in GC B cells and in BCL6 expressing cHL cell lines and in primary HRS cells. Furthermore, the core set of genes directly bound and regulated by BCL6 had a nearly identical chromatin accessibility patterns in GC B cells and in BCL6 expressing cHL cell lines. BCL6 protein degradation was observed with BI-3802 in four cHL cell lines expressing BCL6. After BCL6 degradation, the core set of genes directly bound and regulated by BCL6 was similarly derepressed in cHL cell lines as in the SU-DHL4 DLBCL cell line, indicating that BCL6 is involved in their repression in cHL as in DLBCL. Compared to the BI-5372 control molecule, treatment with BI-3802 significantly decreased proliferation in all cell lines where BCL6 degradation was observed, as well as in the BCL6-dependent SU-DHL4 DLBCL cell line, which was used as a control.

Conclusion: This study broadens the understanding of known oncogenic mechanisms in cHL development and identifies BCL6 as a vulnerability in a fraction of cHL and as a potential therapeutic target.

Disclosures: Pirosa: BeiGene: Honoraria, Other: travel grant; Janssen: Other: travel grant. Condoluci: AbbVie, BeiGene, BMS, Janssen Cilag AG: Honoraria; Gilead: Research Funding. Stathis: Abbvie; ADC Therapeutics; Amgen, Astra Zeneca; Bayer; BMS; Cellestia; Incyte, Loxo Oncology; Merck MSD; Novartis; Pfizer; Philogen; Prelude Therapeutics; Roche: Research Funding; Debiopharm, Janssen, AstraZeneca, Incyte, Eli Lilly, Novartis, Roche, Loxo Oncology: Consultancy; Incyte; AstraZeneca: Other: Travel grant. Stüssi: Bristol Myers Squibb: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Roche: Honoraria; Gilead: Honoraria; Incyte: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria, Research Funding. Santoro: Servier: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Consultancy; BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Arqule: Speakers Bureau; Beigene: Speakers Bureau; Novartis: Speakers Bureau; Sandoz: Speakers Bureau; Lilly: Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; MSD: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bayer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; EISAI: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Consultancy; Takeda: Speakers Bureau; Roche: Speakers Bureau; Abb-vie: Speakers Bureau; Amgen: Speakers Bureau; Celgene: Speakers Bureau; Astrazeneca: Speakers Bureau. Hohaus: Ipsen: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees; Beigene: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Lilly: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. Zucca: AstraZeneca, Beigene, Celgene/BMS, Incyte, Janssen, Roche: Research Funding; Abbvie: Honoraria; AbbVie, BeiGene, BMS, Curis, Eli/Lilly, Incyte, Ipsen, Merck, and Roche: Consultancy; AbbVie, AstraZeneca, BeiGene, and Gilead: Other: Travel grants. Gaidano: AbbVie: Honoraria; AstraZeneca: Honoraria; BeiGene: Honoraria; Hikma: Honoraria; Incyte: Honoraria; Janssen: Honoraria; Lilly: Honoraria. Carlo-Stella: Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen Oncology: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria; ADC Therapeutics: Consultancy, Honoraria, Research Funding; Takeda: Honoraria; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; SOBI: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene/BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; Merck Sharp & Dohme: Honoraria, Membership on an entity's Board of Directors or advisory committees; Karyopharm Therapeutics: Membership on an entity's Board of Directors or advisory committees; Scenic Biotech: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Genmab: Membership on an entity's Board of Directors or advisory committees; Humanitas University, Milano (Italy): Current Employment; Sanofi, ADC Therapeutics: Consultancy; ADC Therapeutics, Roche, Sanofi: Research Funding; AstraZeneca, Celgene/Bristol-Myers Squibb, Incyte, Janssen Oncology, Takeda, Novartis, ADC Therapeutics, Roche, Gilead, SOBI, Merck Sharp & Dohme: Honoraria; Sanofi, ADC Therapeutics, Celgene/Bristol-Myers Squibb, Karyopharm Therapeutics, Roche, Novartis, Scenic Biotech, Janssen Oncology, Merck Sharp & Dohme, SOBI, AbbVie, Genmab, AstraZeneca: Membership on an entity's Board of Directors or advisory committees. Rossi: AbbVie, Adaptive, AstraZeneca, BeiGene, Janssen: Research Funding; AbbVie, AstraZeneca, BeiGene, BMS, Janssen, Lilly: Consultancy, Honoraria.

*signifies non-member of ASH