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645 Cytokine-Linked Remodeling of the Immune Microenvironment Is a Hallmark of Transformation in Follicular Lymphoma

Program: Oral and Poster Abstracts
Type: Oral
Session: 622. Lymphomas: Translational - Non-Genetic: Demystifying the Complexity of the Lymphoma Tumor Microenvironment and Immune Responses
Hematology Disease Topics & Pathways:
Research, Translational Research, Lymphomas, Non-Hodgkin lymphoma, B Cell lymphoma, Diseases, Indolent lymphoma, Aggressive lymphoma, Lymphoid Malignancies, Computational biology, Biological Processes, Technology and Procedures, Imaging, Omics technologies
Sunday, December 8, 2024: 5:00 PM

Nicholas J. Haradhvala, PhD1*, Stephanie Yiu, PhD2,3*, Sam Sadigh, MD4,5*, Laura Beckmann, MD6,7*, Amber Pospistle1*, Stephanie Deng6*, Santiago Rivero6*, Yao Yu Yeo2,8*, Katie Maurer, MD, PhD1,5,6, Vignesh Shanmugam, MD1,4,5, McKayla Van Orden9*, Wesley S Lu, BS9*, Shuqiang Li, PhD1,9*, Kenneth J. Livak, PhD9*, Huaying Qiu10*, Yuzhou Chang10,11*, Ankit Basak1,12,13*, Alex K Shalek, PhD1,13,14*, Mikaela M. McDonough15*, Robert A. Redd, MS16*, Scott J. Rodig, MD, PhD4,5,17, Satyen Gohil, MD PhD18*, Donna S. Neuberg, ScD16, Catherine J. Wu, MD1,5,6*, Gad Getz, PhD1,5,19,20*, Sizun Jiang, PhD1,2,5,21 and Erin M. Parry, MD, PhD1,5,6

1Broad Institute of MIT and Harvard, Cambridge, MA
2Center for Virology and Vaccine Research, Beth Israel Deaconess Medical Center, Boston, MA
3Wyss Institute at Harvard, Boston, MA
4Department of Pathology, Brigham and Women's Hospital, Boston, MA
5Harvard Medical School, Boston, MA
6Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA
7Faculty of Medicine and Cologne University Hospital, Department I of Internal Medicine, Center for Integrated Oncology Aachen Bonn Cologne Duesseldorf, University of Cologne, Cologne, Germany, Cologne, Germany
8Program in Virology, Harvard Medical School, Boston
9Translational Immunogenomics Lab, Dana-Farber Cancer Institute, Boston, MA
10Center for Virology and Vaccine Research, Beth Israel Deaconess Medical Center, Boston
11The Ohio State University, Columbus
12Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge
13Ragon Institute of Mass General, MIT and Harvard, Boston, MA
14Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA
15Dana-Farber Cancer Institute, Boston, MA
16Department of Data Science, Dana-Farber Cancer Institute, Boston, MA
17Center for Immuno-Oncology, Dana-Farber Cancer Institute, Boston, MA
18University College London Hospitals NHS Foundation Trust, London, United Kingdom
19Department of Pathology, Massachusetts General Hospital, Boston, MA
20Krantz Family Center for Cancer Research and Dept. of Pathology, Massachusetts General Hospital, Boston, MA
21Department of Pathology, Dana-Farber Cancer Institute, Boston, MA

Histologic transformation of follicular lymphoma (FL) to an aggressive lymphoma (transformed FL [tFL]) is a common disease complication associated with poor clinical outcomes. However, understanding of the mechanisms that drive FL transformation and the associated lymph node (LN) tumor microenvironment (TME) reorganization remains limited. To define the remodeling that occurs at tFL, we performed single-cell and spatial dissection of clinical samples over the process of transformation.

To identify immune cell populations and malignant B cells, single-cell RNA sequencing (scRNA-seq) was applied to >47,300 malignant and 24,000 non-malignant cells across 10 LN biopsies (4 tFL, 6 FL). tFL samples displayed reduced plasmacytoid (pDCs) and follicular dendritic cells (FDCs) (FDR-corrected Wilcoxon rank sum q=0.13). Bulk RNA-seq analysis of biopsy samples from 18 independent patients with relapsed/refractory disease (5 FL, 13 tFL) confirmed FL and tFL transcriptomes were distinct by principal components (PC) analysis (Hotelling t2 test on first two PCs p=0.0018), with high expression of myeloid genes (CD163, FCGR3A, CES1) and low expression of the FDC gene FDCSP as the top tFL-enriched PC. Decreased FDC and pDCs in tFL, as well as increased fibroblastic reticular cells (FRCs) and monocytes (FDR-corrected Wilcoxon rank sum q<0.1), were detected by CIBERSORTx deconvolution.

As orthogonal validation of our findings in a larger independent cohort, we performed spatial transcriptomics (GeoMx) and proteomics (CODEX, Co-detection by Indexing) across 2 tissue microarrays (TMAs). We localized cell populations from our discovery analysis in the LN TME through acquiring 70 unique GeoMx regions of interest derived from 52 biopsies (33 FL,19 tFL; 9 were paired FL-tFL). Each was subjected to 4 samplings (CD3+ T cells, PAX5+ tumor cells, CD68+ macrophages and other cells). tFL was confirmed to display alterations in LN cell types that establish follicle organization through cell-interactions and chemokine secretion. Consistent with our discovery findings and the expected loss of FDC meshworks with transformation, tFL demonstrated reduced expression of FDC-associated genes and the FDC-secreted CXCL13 (log2 fold-change [L2FC] tFL/FL=-0.87, FDR-corrected Limma moderated t-test q=0.011) compared to FL. While FRC gene expression (e.g. COL1A1 L2FC=1.1, q=1.5x10-5) and expression of matrix remodeling genes (e.g. POSTN and TIMP1, q<0.01) were increased in tFL, we observed reduced expression of secreted chemokines CCL19 and CCL21 (L2FC=-0.82 and q=0.0035, L2FC=-1.3 and q=2.2x10-5, respectively).

From our TMA analyses, we identified differences in monocyte subpopulations between FL and tFL. Our scRNA-seq data identified 3 subsets of monocytes with distinct chemokine expression. One subset was IL1-Bhigh, while another was APOEhighGPNMBhigh; the latter were enriched in TMA samplings of tFL to FL (APOE L2FC=0.89, q=1.2x10-6, GPNMB L2FC=1.2, q=3.3x10-9). A final subset was MRC1(CD206)highCXCL9high, which were found to co-localize with cytotoxic T lymphocytes (CTLs); CXCL9 expression correlated with CTL genes CXCR3, NKG7 and GZMB (p<0.007) across TMA samples. Moreover, high expression of the CXCL9-receptor CXCR3 was identified by receptor-ligand inference in CD8+ CTLs. Spatial analysis of cases with co-existing FL and tFL histology (Visium HD and CODEX) demonstrated that MRC1(CD206)highCXCL9high cells were prominent at boundary regions separating tFL from FL.

Regarding the malignant B cells, we applied non-negative matrix factorization to our scRNA-seq data to identify signatures shared by FL cells across patients, including plasmablast-like (XPB1, MZB1, PRDM1), memory (KLF2, CD69) and CXCR4high. Analysis of TMA samplings identified memory B cell genes depleted in tFL compared to FL (e.g. KLF2 log2 FC=-1.1, q=9.2x10-14), whereas cell cycle gene signature scores increased in tFL (t-test p=8.0x10-5).

Altogether, our data suggests the chemokine and stromal cell driven spatial organization of indolent FL is disrupted upon transformation, associated with a combination of FDC loss, altered FRC function and increased monocytes. Ongoing work is focused on integrative analysis of spatial transcriptomic and proteomic datasets and investigation of the predictive potential of TME evolution for tFL risk.

Disclosures: Haradhvala: MorphoSys: Consultancy. Livak: MBQ Pharma Inc.: Membership on an entity's Board of Directors or advisory committees. Shalek: Honeycomb Biotechnologies, Cellarity, Ochre Bio, Bio-Rad Laboratories, Relation Therapeutics, IntrECate biotherapeutics, Fog Pharma, Passkey Therapeutics, and Dahlia Biosciences: Consultancy, Membership on an entity's Board of Directors or advisory committees. Rodig: Immunitas Therapeutics: Membership on an entity's Board of Directors or advisory committees; Bristol Myers-Squibb, KITE/Gilead, Surface Oncology: Research Funding. Gohil: Novalgen​: Consultancy, Patents & Royalties; EUSA/Recordati: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel support; Gilead: Speakers Bureau; Beigene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel support; Abbvie: Honoraria, Other: travel support; Electra Pharma: Membership on an entity's Board of Directors or advisory committees; UCL Business: Patents & Royalties; Takeda: Other: Travel and conference support; Janssen: Speakers Bureau. Neuberg: Madrigal Pharmaceutical: Current equity holder in publicly-traded company. Wu: BioNtech, Inc: Current equity holder in publicly-traded company; Pharmacyclics: Research Funding; Repertoire: Membership on an entity's Board of Directors or advisory committees; Aethon Therapeutics: Membership on an entity's Board of Directors or advisory committees; Adventris: Membership on an entity's Board of Directors or advisory committees. Getz: Broad Institute: Patents & Royalties: MSMuTect, MSMutSig, POLYSOLVER, SignatureAnalyzer-GPU, MSEye, and MinimuMM-seq; PreDICTA Biosciences: Consultancy, Current equity holder in private company, Other: Founder; IBM, Pharmacyclics/Abbvie, Bayer, Genentech, Calico, and Ultima Genomics: Research Funding; Scorpion Therapeutics: Consultancy, Current equity holder in private company, Other: Founder. Jiang: Elucidate Bio: Membership on an entity's Board of Directors or advisory committees, Other: Founder, board member and scientific advisor; Roche and Sanofi.: Research Funding.

*signifies non-member of ASH