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85 Spatial Dissection of the Bone Marrow Microenvironment in Multiple Myeloma By High Dimensional Multiplex Tissue Imaging

Program: Oral and Poster Abstracts
Type: Oral
Session: 651. Multiple Myeloma and Plasma Cell Dyscrasias: Basic and Translational: Spatial Dissection and Multiomics Analysis of the Multiple Myeloma Tumor and Immune Microenvironment
Hematology Disease Topics & Pathways:
Plasma Cell Disorders, Diseases, Lymphoid Malignancies, emerging technologies, Technology and Procedures
Saturday, December 9, 2023: 9:30 AM

Marc-Andrea Baertsch, MD1,2,3*, Alexander Brobeil, MD4*, John Hickey, PhD3*, Maximilian Haist, MD3*, Alexandra Maria Poos, PhD5*, Guolan Lu, PhD3*, Wilson Kuswanto, MD, PhD3*, Christian Schuerch, MD, PhD6*, Harald Voehringer, PhD7*, Wolfgang Huber8*, Gunhild Mechtersheimer, MD9*, Carsten Mueller-Tidow, MD10*, Peter Schirmacher, MD4*, Katja Weisel, MD11, Roland Fenk, MD, PhD12*, Hartmut Goldschmidt13,14, Yury Goltsev, PhD15*, Marc S. Raab10,14,16*, Niels Weinhold, PhD10* and Garry P. Nolan, PhD17*

1Department of Hematology, Oncology and Rheumatology, University Hospital Heidelberg, Heidelberg, Germany
2Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), Heidelberg, Germany
3Department of Microbiology and Immunology, Stanford University, Stanford, CA
4Department of Pathology, Heidelberg University Hospital, Heidelberg, Germany
5Heidelberg Myeloma Center, Department of Internal Medicine V, University Hospital Heidelberg, Heidelberg, Germany
6Department of Pathology and Neuropathology, Tuebingen University Hospital, Tuebingen, Germany
7European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
8Genome Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany
9Institute of Pathology, Heidelberg, Germany
10Department of Hematology, Oncology and Rheumatology, Heidelberg University Hospital, Heidelberg, Germany
11Department of Oncology, Hematology and Bone Marrow Transplantation with Section of Pneumology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany, Hamburg, Germany
12Department of Hematology, Oncology and Clinical Immunology,, University Hospital Düsseldorf, Duesseldorf, Germany
13Internal Medicine V, GMMG-Study Group, University Hospital Heidelberg, Heidelberg, Germany
14National Center for Tumor Diseases (NCT), Heidelberg University Hospital, Heidelberg, Germany
15Stanford Medical School, Stanford, CA
16Clinical Cooperation Unit (CCU) Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), Heidelberg, Germany
17Department of Pathology, Stanford University, Stanford, CA

Introduction

Multiple myeloma (MM) is shaped by interactions between immune, stromal and tumor cells in the bone marrow (BM) microenvironment (BMME). Our understanding of these processes is based on high parametric flow cytometry and single cell transcriptomics, which lack spatial resolution. Thus, crucial aspects of the in situ tumor ecosystem, including cell-cell interactions, can only be predicted. Novel high dimensional, multiplex tissue imaging methods are able to capture similar granularity while also preserving spatial organization. By optimizing antibody selection, as well as staining, imaging and data processing protocols we have established a BMME-targeted CODEX workflow based on a 60 marker panel. Here, we conducted the first comprehensive multiplex protein imaging survey of the BMME across the spectrum of MM and precursor stages and leveraged paired FISH and whole genome sequencing (WGS) to elucidate the interplay of tumor intrinsic features and the BMME.

Methods

We imaged 489 BM trephine biopsies in tissue microarray format on the CODEX (Akoya Biosciences) platform. After quality control (≥750 cells per core, <25% unclassifiable cells) we retained data from 12 non-malignant controls, 13 MGUS, 20 smoldering MM, 349 uniformly treated newly diagnosed MM (NDMM) patients from a phase III clinical trial, as well as 22 relapsed MM samples with a median of 3 (range 1-6) tissue cores per sample, comprising a total of 3.1 million single cells. CODEX data was processed and analyzed using custom scripts. Cellular neighborhoods were computed by clustering the cell type composition of sliding windows (n=10 nearest neighboring cells) across the tissue. WGS (n=212) and FISH (n=309) were performed on CD138-purified BM mononuclear cells and analyzed using in-house pipelines.

Results

We identified 40 cell types, encompassing the relevant lineages (myeloid, lymphoid, stromal, endothelial, tumor), subtypes (e.g. T, B, NK cells) and functional states (e.g GRZB, Ki67, and PD1) of the BMME. The degree of plasma cell infiltration determined by CODEX correlated with the clinical pathologist’s quantification based on classical immunohistology (r=0.77, p<0.0001), and the kappa/lambda status of MM cells matched the reported light chain restriction, supporting the validity of our data.

At the compositional level, we observed shifts in relative cell type abundance from precursor to MM stages. Major changes in the BMME included enriched stromal, endothelial and CD8+ T cells as well as depleted polymorphonuclear (PMN), erythropoietic and CD4+ T cells in advanced stages. The highest proportion of monocytoid dendritic cells, monocytes, mast cells and cytotoxic CD8+ T cells was seen in relapsed MM.

Leveraging the spatial resolution of the CODEX data, we identified multiple distinct cellular neighborhoods (CNs) defined by recurrent local cell type composition. CNs enriched for plasma cells in combination with different immune cell subsets including exhausted T cells increased towards the MM stage, while CNs dominated by PMNs, erythropoietic cells and CD206 negative macrophages decreased.

To investigate links between MM subtypes and BMME patterns, we correlated CODEX data with MM-initiating and driver events. While patients with t(4;14) showed a significant increase in eosinophils, patients with gain1q had higher levels of endothelial and stromal subsets colocalizing in the same CN. Distinct types of stromal cells were also enriched in patients with double hits (≥2 FISH high risk features). RAS mutations and biallelic inactivation of tumor suppressor genes were associated with altered T cell subset composition.

Further corroborating the importance of BMME architecture, a CN with colocalization of plasma and exhausted CD8 T cells was associated with unfavorable prognosis in NDMM patients.

Conclusions

High dimensional, multiplex tissue imaging of the BMME enables interrogation of cellular interactions and tissue architecture in situ in clinical samples at single cell resolution. To our knowledge this is the first highly multiplexed spatial study of the human BMME in a large clinical cohort and the first such study in multiple myeloma. We track characteristic shifts in tissue composition associated with disease progression, reveal genotype-phenotype associations in the BMME as a correlate of tumor-microenvironment co-evolution and detect prognostically relevant tissue architectural features.

Disclosures: Schuerch: AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Enable Medicine: Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees, Research Funding. Mueller-Tidow: Astellas: Other: Lecture fees; Pfizer, Janssen: Membership on an entity's Board of Directors or advisory committees; Pfizer, Daiichi Sankyo, BiolineRx, Bayer: Research Funding. Weisel: Bristol Myers Squibb/Celgene: Consultancy, Honoraria, Other: Research grant to institution; GlaxoSmithKline: Consultancy, Honoraria, Other: Research grant to institution; Takeda: Consultancy, Honoraria, Other: Research grant; BeiGene: Consultancy, Honoraria; Stemline: Honoraria; Sanofi: Consultancy, Honoraria, Other: Research grant to institution; Roche Pharma: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; AstraZeneca: Honoraria; Karyopharm: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Other: Research grant to institution; Oncopeptides: Consultancy, Honoraria; Novartis: Honoraria; Adaptive Biotech: Consultancy, Honoraria; Amgen: Consultancy, Honoraria, Other: Research grant to institution; AbbVie: Consultancy, Honoraria, Other: Research grant to institution; Menarini: Consultancy, Honoraria. Fenk: Janssen, Amgen, GSK, Takeda, BMS: Honoraria. Goldschmidt: Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Research Funding; Array Biopharma: Research Funding; BMS/Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Research Funding; Chugai: Honoraria, Patents & Royalties, Research Funding; Dietmar-Hopp-Foundation: Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Research Funding; Johns Hopkins University: Research Funding; Mundipharma: Research Funding; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Research Funding; Glycomimetics: Research Funding; GSK: Honoraria, Other: Travel Support, Research Funding; Heidelberg Pharma: Research Funding; Hoffman- La Roche: Research Funding; KaryoPharm: Research Funding; Incyte: Research Funding; Millenium Pharmaceuticals: Research Funding; Molecular Partners: Research Funding; MSD: Research Funding; Morphosys AG: Research Funding; Pfizer: Honoraria, Patents & Royalties: Travel Support, Research Funding; Takeda: Research Funding; Novartis: Honoraria, Other: Travel Support, Research Funding; Adaptive Biotechnology: Membership on an entity's Board of Directors or advisory committees. Goltsev: Akoya Biosciences: Current equity holder in publicly-traded company. Raab: AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Heidelberg Pharma: Research Funding; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Heidelberg University Hospital: Current Employment; GSK: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Nolan: Akoya Biosciences: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Pfizer, Inc., Vaxart, Inc., Celgene, Inc. and Juno Therapeutics, Inc.: Research Funding.

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