-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

456 An Llmpp Study: Genomic Characterization of Novel PTCL- Biological Subtypes Reveal Distinctive Therapeutic Vulnerabilities

Program: Oral and Poster Abstracts
Type: Oral
Session: 621. Lymphomas: Translational – Molecular and Genetic: Single-cell and Spatial Analyses in Aggressive and T Cell Lymphomas
Hematology Disease Topics & Pathways:
Research, Translational Research, Genomics, Immune mechanism, Treatment Considerations, Biological therapies, Immunology, Biological Processes, Monoclonal Antibody Therapy, Molecular biology, Pathogenesis
Sunday, December 8, 2024: 10:45 AM

Dylan T. Jochum, BS1,2, Alyssa Bouska, PhD1*, Sunandini Sharma, PhD1*, Suchita Vishwakarma, M.Pharm3*, Catalina Amador, MD4*, Waseem Gul Lone, PhD1*, Ab Rauf Shah, Ph.D1*, Abdul Rouf Mir, PhD1*, Mahfuza Afroz Soma, MS1*, Syed Mohsin Mehmood, MBBS1*, Zaina Nasser5*, Andrew L. Feldman, MD6, Timothy C. Greiner, MS, MD1*, Julie M. Vose, MD, MBA7, James R. Cook, MD, PhD8, Sarah L. Ondrejka, DO8, Elaine S. Jaffe, MD9*, Andreas Rosenwald10*, German Ott, MD11*, Phillipp W. Raess, MD, PhD12*, Kerry J. Savage, MD, MSc13, Graham W. Slack, MD14*, David W. Scott, MBChB, PhD15, Joo Y Song, MD16*, Elias Campo, MD, PhD17,18*, Louis M. Staudt, MD, PhD19, Lisa M. Rimsza, MD20, Dennis D Weisenburger, MD1*, Giorgio Inghirami, MD21*, Amar Natarajan, PhD2,3*, Javeed Iqbal, PhD, MSc1* and Wing Chung Chan, MD16

1Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE
2Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE
3Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE
4Division of Hematopathology, University of Miami, Miami, FL
5College of Arts and Sciences, University of Nebraska, Lincoln, NE
6Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN
7Department of Hematology and Oncology, University of Nebraska Medical Center, Omaha, NE
8Department of Pathology and Laboratory Medicine, Cleveland Clinic, Cleveland, OH
9Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, Bethesda, MD
10Institute of Pathology, University Hospital of Wuerzburg, and Comprehensive Cancer Center Mainfranken, Würzburg, Germany
11Department of Clinical Pathology, Robert-Bosch-Krankenhaus, and Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany
12Department of Pathology and Laboratory Medicine, Oregon Health & Science University, Portland, OR
13BC Cancer, Vancouver, BC, Canada
14Centre for Lymphoid Cancer, BC Cancer, Vancouver, BC, Canada
15Centre for Lymphoid Cancer, British Columbia Cancer, Vancouver, BC, Canada
16Department of Pathology, City of Hope National Medical Center, Duarte, CA
17Hematopathology Unit, Hospital Clinic, Barcelona, Spain
18Institute of Biomedical Research August Pi i Sunyer (IDIBAPS), University of Barcelona, Barcelona, Spain
19Metabolism Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD
20Department of Laboratory Medicine and Pathology, Mayo Clinic, Scottsdale, AZ
21Department of Pathology and Laboratory Medicine, Weill Cornell Medicine, New York, NY

Introduction

Peripheral T-cell lymphoma (PTCL) encompasses a heterogeneous group of clinically aggressive entities. ~30% of cases cannot be further classified and are designated as PTCL – not otherwise specified (NOS). We delineated two novel molecular subgroups among PTCL-NOS with distinct clinical and biological features: PTCL-GATA3 which is characterized by high expression of GATA3, a master regulator of T-helper-2-cell (TH2) differentiation, and its target genes (i.e., CCR4, IL4, IL13) and PTCL-TBX21 which is characterized by high expression of TBX21, a master regulator of TH1 differentiation, and its target genes (i.e., CXCR3, IFNG), with the GATA3 subtype showing significantly worse clinical outcomes (overall survival). Nonsense mutations in CCR4 are found predominately in PTCL-GATA3, often within the cytoplasmic domain and gain-of-function in nature.

Methods

Mutation and copy number data were generated from whole exome sequencing (TFH phenotype excluded following LLMPP pathology review; n = 153). Single nucleotide variants were determined using Mutect2 and Varsvan2 using in-house filtering. Copy number (CN) profiles were generated using CNVkit. Four CCR4 cytoplasmic domain mutants and controls (cytoplasmic domain knockout [CDKO], wild-type CCR4 overexpression (CCR4-WT), and empty vector [EV]) were transduced into healthy-donor CD4+ T cells (n = 2). Polarization studies were generated using TH1- or TH2-associated cytokine exposure. Surface plasmon resonance was used to determine the binding affinity of CCR4 proteins to CCL17 and CCL22. In silico docking studies were done through Schrödinger's BioLuminate. Antibody-dependent cell-mediated cytotoxicity (ADCC) assay was performed through co-incubation of NK92 (CD16 and CD56) and CCR4 edited primary CD4+ T-cells (stained with calcein AM) for 2 hours at a 5:1 NK:T cell ratio by 7-AAD flow cytometry.

Results

PTCL-GATA3 had a higher frequency of CN aberrations (e.g. TP53, PTEN CN loss) and a more aberrant genome (p < 0.001) than PTCL-TBX21, including mutations with genes associated with DNA repair and damage (TP53, ATM) and T-cell differentiation (CCR4). CCR4 mutations were virtually mutually exclusive with TP53 mutations, and these cases showed higher CCR4 mRNA and protein expression than CCR4-WT cases (p < 0.05).

Primary CD4+ T-cells with ectopic expression of the CCR4 variants, especially CCR4-Q330X, did not require CD3/CD28 for proliferation, bypassing CD3/CD28 ligation for T-cell activation. In addition, proliferation rates were similar between TH2-conditional media and normal media, but proliferation and TBX21 expression in TH1-conditional media were inhibited, implying a novel role of CCR4 in T-cell receptor (TCR) activation and TH-differentiation. CCR4 variant proteins showed higher binding affinity for their natural ligands, CCL22 and CCL17, with mutants exhibiting tighter binding to both ligands (1.5- to 3-fold decrease in KD) by surface plasmon resonance and in silico modeling studies. The in silico models demonstrated mutant CCR4 bound mogamulizumab tighter than CCR4-WT, consistent with a higher ADCC-induced apoptosis rate, highest with CCR4-Q330X mutation. CCR4-Y331X exhibited a worse docking score compared to CCR4-WT and a lower ADCC-induced apoptosis rate.

Conclusions

Our findings reveal distinct genetic characterizations including aberrant TP53 and PI3K signaling and CCR4 mutations in PTCL-GATA3. These CCR4 mutations are shown to have a key role in PTCL-GATA3 pathogenesis by dysregulating TH-differentiation and aberrantly activating the TCR. CCR4 mutation promotes the TH2 phenotype by blocking TH1 differentiation. CCR4 mutant proteins demonstrated an increased binding affinity towards its known ligands, CCL17 and CCL22, and mogamulizumab, consistent with previous studies exhibiting mogamulizumab efficacy against CCR4 mutations. However, the most frequent CCR4 mutant, Y331X, showed worse binding compared to other CCR4 mutants or CCR4-WT, a mutation that showed Mogamulizumab-resistance in a prior CTCL study as well as in an in vitro ADCC assay. The results emphasize aberrant pathways implicated by genomic and clinical evaluation of two novel PTCL subtypes, as well as support the novel role of mutant CCR4 in enhancing T-cell lymphomagenesis and T-helper differentiation, highlighting multiple therapeutic targets.

Disclosures: Feldman: Zeno Pharmaceuticals: Patents & Royalties; Seattle Genetics: Research Funding. Vose: Novartis: Honoraria; Abbvie: Honoraria, Research Funding; Pfizer: Research Funding; GenMab: Honoraria, Research Funding. Rosenwald: Incyte: Other: Institutional research contract ; MorphoSys: Other: institutional research contract. Savage: Regeneron: Other: DSMC; AbbVie: Consultancy; Seagen: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding. Scott: Roche: Consultancy, Honoraria; AstraZenenca: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Genmab: Consultancy, Honoraria; Veracyte: Consultancy, Honoraria; Roche/Genentech: Research Funding; Nanostring: Patents & Royalties: use of gene expresssion to subtype aggressive lymphoma. Inghirami: Daiichi Sankyo: Consultancy.

<< Previous Abstract | Next Abstract
*signifies non-member of ASH