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809 Chromatin Accessibility Profiling to Increase Diagnostic Accuracy and Refine Cell-of-Origin Classification of Mature T-Cell Lymphomas

Program: Oral and Poster Abstracts
Type: Oral
Session: 621. Lymphomas: Translational—Molecular and Genetic: Sequencing and Subtyping
Hematology Disease Topics & Pathways:
Translational Research, Clinically Relevant
Monday, December 13, 2021: 5:30 PM

Edith Julia1,2*, Sylvain Mareschal3*, Amel Chebel4*, Camille Golfier, MD5*, Tony Andreas Müller6*, Camille Lours7*, Sofiane Hadj-Hamou8*, Nathalie Bissay8*, Manon Tschanz8*, Michele Ceribelli, PhD9*, Dimitri Chartoire8*, Aurélie Vernay4*, Sylvain Carras10*, David Sibon, MD, PhD11*, Ambroise Marçais, MD, PhD12*, Camille Laurent, MD, PhD13*, Laurent Martin, MD, PhD14*, Vahid Asnafi, MD, PhD15*, Lea Payen16*, Damien Sanlaville17*, Claire Bardel-Danjean18*, Mikael Roussel, MD, PhD19*, Vanessa Szablewski, MD20*, Fanny Drieux, MD, PhD21*, Philippe Ruminy22*, Marco Herling, MD23,24*, Catherine Chassagne-Clément, MD25*, Gilles Salles, MD, PhD26,27, Pierre Sujobert, MD, PhD28*, Alexandra Traverse-Glehen, MD, PhD29*, Lucile Baseggio, MD30*, Laurence De Leval, MD31, Louis M. Staudt, MD, PhD32, Philippe Gaulard, MD, PhD33*, Laurent Genestier, PhD34* and Emmanuel Bachy, MD, PhD35*

1Hematology deparment - Centre hospitalier Lyon Sud, Pierre-Bénite, France
2Lymphoma Immunobiology Research Unit- Medical Faculty Lyon Sud, Pierre Bénite, France
3Cancer Research Center of Lyon, INSERM U1052 UMR CNRS 5286, Lyon, FRA
4Lymphoma Immunobiology Research Unit - Medical Faculty Lyon Sud, Pierre-Bénite, France
5Department of Hematology, Lyon-Sud Hospital, Pierre-Benite, France
6Department of Internal Medicine - Köln, Köln, Germany
7HCL, Pierre Bénite, FRA
8Lymphoma Immunobiology Research Unit- Medical Faculty Lyon Sud, Pierre-Bénite, France
9National Cancer Institute, NIH, Bethesda, MD
10Centre Hospitalier Universitaire Grenoble, La Tronche, FRA
11Hematology Department, Necker University Hospital, Paris, France
12Hematology Department, Necker University Hospital, PARIS, France
13Institut universitaire du cancer Toulouse-Oncopole CHU Toulouse, Toulouse, France
14Pathology Department, University Hospital F. Mitterrand and Inserm UMR 1231, Dijon, France
15Biological Hematology, Necker University Hospital, Paris, France
16Centre Hospitalier Lyon Sud, Pierre-Bénite, France
17HCL, UCBL, Bron Cedex, FRA
18Hospices civils de Lyon, Lyon, France
19CHU Rennes, Rennes, France
20Pathological Anatomy and Cytology, CHU Montpellier, Montpellier, FRA
21INSERM U1245, Centre Henri Becquerel, UNIROUEN, University of Normandie, Rouen, France
22Centre Henri Becquerel, Rouen, FRA
23Department I of Internal Medicine, Center for Integrated Oncology (CIO), Aachen-Bonn-Cologne-Duesseldorf, University of Cologne, Cologne, Germany
24Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany
25Department of Pathology, Leon Berard Center, Lyon, France
26Centre Hospitalier Lyon-Sud, Hospices Civils de Lyon, Université Claude Bernard, Pierre Bénite, France
27Memorial Kettering Cancer Center, New York, NY
28Laboratory of Hematology, Centre Hospitalier Lyon Sud, Pierre Bénite, France
29Department of Pathology, Hopital Lyon Sud, Pierre-Benite, France
30Department of Hematology, Centre Hospitalier Lyon-Sud, Hospices Civils de Lyon, Université Claude Bernard, Pierre Benite, France
31Hôpital Universitaire de Lausanne, Lausanne, Switzerland
32Lymphoid Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD
33Département de Pathologie, Hôpitaux Universitaires Henri Mondor, Créteil, France
34Lymphoma Immunobiology Research Unit - Lyon Sud Medical Faculty, Oullins, FRA
35Department of Hematology, Lyon Sud Hospital, Lyon, France

Background:
Mature T-cell lymphomas and leukemias (MTCL) are heterogeneous diseases with dismal prognosis. Differentiating between the numerous entities requires specialized pathology expertise and studies show up to 20% change in diagnosis after expert review of cases (Laurent, JCO, 2017). Assay for transposase accessible chromatin sequencing (ATAC-seq) is a simple technique to profile open chromatin regions (OCR) proven to be highly discriminant for cell-of-origin identification regardless of cell activation status (Shih, Cell, 2016). We applied ATAC-seq to MTCL in order to explore the epigenetic landscape of these diverse entities, compared them to normal T-cell subtypes and built a predictive model to help diagnosis.

Method:
Ten-thousand FACS-sorted single cells from primary MTCL samples and 50µm section of frozen tumoral tissue from the TENOMIC French T-cell Lymphoma Consortium were processed according to the previously published FAST-ATAC and OMNI-ATAC protocols respectively (Corces, Nat Genetics, 2016 & Nat. Methods, 2017). Concurrently we applied FAST ATAC to different normal T- and NK-cell subsets sorted from healthy donor PBMC or lymph node suspensions. Sequencing data were processed by an adapted version of ENCODE ATAC-seq pipeline. Matrix of insertion events in peaks by sample was obtained, normalized and most variant peaks were selected for UMAP projection.

Results:
In total, 678 normal and tumoral samples were sequenced to provide a comprehensive landscape of chromatin accessibility in MTCL. Epigenetic profiling by ATAC-seq of FACS-sorted tumoral samples resulted in a complete segregation of the known MTCL entities (AITL, TFH-PTCL, ALK+ and ALK- ALCL, HSTL, CTCL, ATLL, LGL and T-PLL). Most PTCL-NOS (13/17) clustered with a pre-defined MTCL subtype (mainly AITL/TFH-phenotype PTCL, CTCL and lymphomas exhibiting cytotoxic features). All but one discordant diagnosis between pathology and ATAC-seq (1/11) led to revised diagnosis after pathology review.
Unsupervised clustering of normal NK- and T-cell subtypes (N=49) and sorted tumoral lymphoma cells (N=104) confirmed that AITL derive from TFH cells. HSTL and LGL closely segregated with NK- and gamma-delta T cells, in line with their known innate-like phenotype.
Surprisingly, the cell-of-origin of T-PLL seems to be naïve T cells despite the known expression of central memory markers on leukemic cells.
Beyond epigenetic classification, background reads from ATAC-seq profiles were used to detect copy number variation (CNV), such as isochromosome 7q in HSTL. In addition, HTLV1 and EBV viral sequence detection in ATAC-seq reads strengthened identification of ATLL and NKTCL cases.
Finally, using unsupervised deconvolution approaches, we were able to discriminate different MTCL subtypes from 223 processed bulk frozen samples. All known MTCL subtypes were differentiated (AITL/PTCL-TFH, HSTL, NKTCL, ATLL, ALK- and ALK+ ALCL, MEITL, EATL). A subgroup of PTCL-NOS harboring GATA3 OCRs and a distinctly high CNV number was isolated that might correspond to previously described PTCL-GATA3 subtypes (Iqbal, Blood, 2019).
A random forest model was trained to predict diagnosis based on chromatin-accessibility clusters defined in the discovery cohort of patients. The model showed accurate prediction performance by cross-validation. External validation on 172 samples collected from 5 tertiary care centers will be presented at the meeting.

Conclusion:
ATAC-seq is a fast and cost-effective technique to help and refine MTCL pathological classification and allows for putative cell-of-origin identification in lymphoma. Training of a machine learning model to predict MTCL entity diagnosis based on ATAC-seq analysis of fresh or frozen samples shows promising results.

Disclosures: Sibon: Janssen: Consultancy; Abbvie: Consultancy; iQone: Consultancy; Takeda: Consultancy; Roche: Consultancy. Drieux: Genexpath: Patents & Royalties: The author is a potential inventor on a patent application for the LymphoSign, which has been licensed for by Genexpath Patents & Royalties.. Ruminy: Genexpath: Patents & Royalties: The author is a potential inventor on a patent application for the LymphoSign, which has been licensed for by Genexpath Patents & Royalties. . Salles: Bristol-Myers Squibb/Celgene: Consultancy, Honoraria; Bayer: Honoraria; BeiGene: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Kite/Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Morphosys: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Miltenyi: Consultancy; Loxo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Genmab: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Roche/Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Ipsen: Consultancy; Epizyme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Debiopharm: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Rapt: Consultancy, Membership on an entity's Board of Directors or advisory committees; Regeneron: Consultancy, Honoraria; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Velosbio: Consultancy, Membership on an entity's Board of Directors or advisory committees; Allogene: Consultancy. Gaulard: Alderaan: Research Funding; Sanofi: Research Funding; Innate Pharma: Research Funding; Gilead: Consultancy; Takeda: Consultancy, Honoraria. Bachy: Novartis: Honoraria; Daiishi: Research Funding; Roche: Consultancy; Takeda: Consultancy; Incyte: Consultancy; Kite, a Gilead Company: Honoraria.

*signifies non-member of ASH