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453 Single Cell Multi-Omic Dissection of Response and Resistance to Chimeric Antigen Receptor T Cells Against BCMA in Relapsed Multiple Myeloma

Program: Oral and Poster Abstracts
Type: Oral
Session: 651. Multiple Myeloma and Plasma Cell Dyscrasias: Basic and Translational: Characterizing Response and Resistance to CAR-T and TCEs
Sunday, December 10, 2023: 10:00 AM

Maximilian Merz, MD1, Luise Fischer2*, Jaren Sia3*, Patrick Born4*, Ronald Weiss5*, Nora Grieb1*, Michael Rade6*, Andreas Boldt7*, Stephan Fricke8*, Paul Franz9*, Jonathan Scolnick, PhD10*, Lakshmi Venkatraman3*, Stacy Xu, PhD11*, Christina Kloetzer12*, Simone Heyn12*, Anne Sophie Kubasch, MD13*, Ronny Baber14*, Song Yau Wang5*, Enrica Bach12*, Sandra Hoffmann12*, Jule Ussmann4*, Klaus H Metzeler, MD15, Marco Herling12*, Carmen Herling, MD12*, Madlen Jentzsch12*, Georg-Nikolaus Franke12*, Ulrich Sack7*, Ulrike Koehl16*, Kristin Reiche6*, Uwe Platzbecker, MD17 and Vladan Vucinic, M.D.18*

1Department of Hematology, Cellular Therapy, Hemostaseology and Infectious Diseases, University Leipzig Medical Center, Leipzig, Germany
2University of Leipzig, Leipzig, Saxony, Germany
3Singleron Biotechnologies, Singapore, Singapore
4Department for Hematology, Cell Therapy, Hemostaseology and Infectious Diseases, University of Leipzig Medical Center, Leipzig, Germany
5University of Leipzig, Leipzig, Germany
6Fraunhofer IZI, Leipzig, Germany
7University of Leipzig, Leipzig, DEU
8Fraunhofer Institute for Cell Therapy and Immunology (IZI), Leipzig, Germany
9Fraunhofer IZI, Leipzig, DEU
10Singleron Biothechnologies Pte Ltd, Singapore, Singapore
11Singleron Biotechnologies Pte LtD, Singapore, Singapore
12Department of Hematology, Celltherapy, Hemostaseology and Infectious Diseases, University Leipzig Medical Center, Leipzig, Germany
13Department of Hematology, University Hospital of Leipzig, Dresden, Germany
14Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University of Leipzig Medical Center, Leipzig, Germany
15Department of Hematology, Cellular Therapy, Hemostaseology and Infectious Diseases, University Leipzig Medical Center, Leipzig, NA, Germany
16Fraunhofer IZI, Leipzig, Saxony, DEU
17Department of Hematology, University Hospital of Leipzig, Leipzig, Germany
18Department of Hematology, Cellular Therapy, Hemostaseology and Infectious Diseases, University of Leipzig, Leipzig, Germany

Background: Chimeric antigen receptor (CAR) T cell therapy has revolutionized treatment of relapsed/refractory multiple myeloma (RRMM). Robust variables that predict long-term response are currently missing. Limited data are especially available on the impact of bridging therapies on manufacturing and outcome. We conducted a longitudinal single-cell multi-omics study to identify factors that predict response to BCMA-directed CAR T cells. Changes in the immune microenvironment associated with response were analyzed as well as the impact of prior bridging therapy with bispecific antibodies on subsequent CAR T cell manufacturing and outcome.

Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from 29 consecutive MM patients treated with commercially available anti-BCMA CAR T cells on the day of leukapheresis as well as days 30 and 100 after CAR T cell infusion. PBMCs were subjected to single cell RNA, T-cell receptor (TCR) and B-cell receptor (BCR) sequencing. A custom panel of 57 oligonucleotide-coupled antibodies was used to study surface proteomics. Downstream analyses were performed with Seurat. Differences in cellular compositions at all three time points were analyzed with scCODA. To analyze CAR T cell functionality, CAR T cells from peripheral blood were isolated 7 days after infusion and subjected to an in vitro cytotoxicity assay after expansion and stimulation. Patients were grouped based on their best response following CAR T cell infusion (CR: n=12, non CR: n=17).

Results: In total, 375,338 cells were sequenced (median 7246 cells/sample, range 1,569-10,972 cells) and 354,878 cells (94.5%) passed quality assessment. Quantitative and qualitative differences in the cellular composition of peripheral blood between CR and non CR patients were detected at the time of leukapheresis as well as on days 30 and 100 following infusion. CR patients harbored significantly more CD8+ effector memory T cells (TEM) at leukapheresis and less NK cells on day 30 after therapy compared to non CR patients. Regulatory T cells isolated at the time of leukapheresis from non CR patients exhibited significantly higher surface protein levels of CXCR3, CD40, CD95 and KLRG1 (p<0.015, respectively) that have been associated with T cell senescence and impaired tumor immunity. No significant differences in cell numbers between CR and non CR were detected on day 100 after CAR T cell infusion. However, single cell TCR analysis revealed an increasing diversity in the TCR repertoire over time in patients with CR, while Shannon diversity decreased from leukapheresis over day 30 to day 100 in non CR patients (p=0.004). The prior administration of the bispecific antibody teclistamab had no significant impact on the quantitative cellular composition at the time of leukapheresis. However, termination of manufacturing in the first attempt occurred in all patients with a close proximity of teclistimab administration and apheresis. Differential gene expression analysis showed that the application of teclistamab was associated with impaired T cell activation and exhaustion indicated by upregulation of e.g. CTLA4, TIGIT, LAG3 and GZMK. After discontinuation of teclistamab (median 4 weeks) and successful manufacturing of CAR T cells, we found no significant differences for in vitro cytotoxicity and in vivo expansion of CAR T cells. CAR T cells isolated at day 7 post-infusion from patients in CR, non CR or with prior teclistamab exposure, effectively eliminated MM cells (U-266). Tracking of single CAR T cells over time showed that the majority of CAR+ cells were CD8+ TEMs regardless of remission achievement or prior teclistamab exposure.

Conclusion: We demonstrate that differences between MM patients achieving a CR and patients with suboptimal response upon anti-BCMA CAR T cell therapy can already be identified at the time of leukapheresis. Long-term changes associated with CR include a diversification of the TCR repertoire. Successful CAR T cell manufacturing is hampered by exposure to bispecific antibodies but can be successfully achieved by allowing for a wash-out phase of ca. 4 weeks.

Disclosures: Merz: AMGEN, TAKEDA, BMS, JANSSEN, STEMLINE, ROCHE: Honoraria. Fricke: Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V.: Patents & Royalties, Research Funding; MSGO: Consultancy, Honoraria; AstraZeneca: Consultancy, Honoraria; Vertex Pharmaceuticals: Consultancy, Honoraria; Kite/Gilead Sciences: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Janssen-Cilag: Consultancy, Honoraria. Scolnick: Singleron Biotechnologies: Current Employment. Xu: Singleron Biotechnologies: Current Employment. Kloetzer: Janssen: Honoraria. Heyn: Janssen: Honoraria. Ussmann: Sanofi: Other: former travel support. Metzeler: BMS: Consultancy, Honoraria; AbbVie: Honoraria, Research Funding; Pfizer: Honoraria; Otsuka: Honoraria; Janssen: Honoraria; Novartis: Consultancy. Herling: Abbvie, Beigene, Janssen, Stemline, Takeda: Consultancy; Mundipharma EDO, Janpix, Novartis, Roche: Research Funding. Jentzsch: Blueprint Medicine: Honoraria; Pfizer: Honoraria; Novartis: Honoraria; BMS: Honoraria; Amgen: Honoraria; JAZZ: Honoraria. Platzbecker: Syros: Consultancy, Honoraria, Research Funding; MDS Foundation: Membership on an entity's Board of Directors or advisory committees; Fibrogen: Research Funding; Silence Therapeutics: Consultancy, Honoraria, Research Funding; Servier: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel support; medical writing support, Research Funding; BMS: Research Funding; BeiGene: Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Celgene: Honoraria; Jazz: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy; Curis: Consultancy, Research Funding; Merck: Research Funding; Janssen Biotech: Consultancy, Research Funding; Geron: Consultancy, Research Funding; Roche: Research Funding; Amgen: Consultancy, Research Funding; Novartis: Consultancy, Honoraria, Research Funding.

*signifies non-member of ASH