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4798 Molecular Mechanisms Promoting Long-Term Cytopenia after BCMA CAR-T Therapy in Multiple Myeloma

Program: Oral and Poster Abstracts
Session: 702. CAR-T Cell Therapies: Basic and Translational: Poster III
Hematology Disease Topics & Pathways:
Research, Translational Research
Monday, December 9, 2024, 6:00 PM-8:00 PM

Maria Luisa Palacios-Berraquero1*, Paula Rodriguez-Marquez2*, Maria Erendira Calleja-Cervantes2,3*, Nerea Berastegui2*, Aintzane Zabaleta, PhD2*, Leire Burgos, PhD2*, Diego Alignani, PhD2*, Patxi San-Martin2*, Amaia Vilas-Zornoza1*, Saray Rodríguez Diaz2*, Susana Inogés, MD, PhD1,4,5*, Ascensión López-Díaz De Cerio1,4,5*, Sofia Huerga1,6*, Luis-Esteban Tamariz-Amador, MD1,6*, Jose J. Rifon, M.D.1,5,6*, Ana Alfonso Pierola, MD, PhD1,5,7*, Juan J Lasarte, PhD8*, Bruno Paiva, PhD2,5,6*, Mikel Hernaez, PhD3,5,9*, Paula Rodríguez-Otero, MD, PhD1,5,6*, Jesús F. San-Miguel, MD, PhD1,2,5,10, Teresa Ezponda2,11*, Juan Roberto Rodriguez-Madoz, PhD2,6,12* and Felipe Prosper, MD, PhD2,6,13,14

1Hematology and Cell Therapy Department. Clinica Universidad de Navarra. IdiSNA., Pamplona, Spain
2Hemato-Oncology Program. Cima Universidad de Navarra. IdiSNA., Pamplona, Spain
3Computational Biology Program. Cima Universidad de Navarra. IdiSNA., Pamplona, Spain
4Immunology and Immunotherapy Department. Clinica Universidad de Navarra., Pamplona, Spain
5Centro de Investigación Biomedica en Red de Cancer (CIBERONC)., Madrid, Spain
6Cancer Center Clinica Universidad de Navarra (CCUN)., Pamplona, Spain
7Cancer Center Clinica Universidad de Navarra (CCUN), Pamplona, Spain
8Immunology and Immunotherapy Program. Cima Universidad de Navarra. IdiSNA., Pamplona, Spain
9Center for Data Science and Artificial Intelligence Institute (DATAI), Universidad de Navarra, Pamplona, Pamplona, Spain
10Cancer Center Clinica Universidad de Navarra, CIMA, IDISNA, Pamplona, Spain
11Centro de Investigación Biomédica en Red de Cáncer, CIBERONC, Madrid, Spain
12Centro de Investigacion Biomedica en Red de Cancer (CIBERONC)., Madrid, Spain
13Hematology and Cell Therapy Department, Clinica Universidad de Navarra, IdiSNA, Pamplona, Spain
14Centro de Investigación Biomedica en Red de Cancer (CIBERONC), Madrid, Spain

Hematologic toxicity, specifically prolonged cytopenia, is a common side effect associated with CAR-T therapies, being particularly severe in patients with relapsed/refractory Multiple Myeloma (MM) (1). However, to date, its etiology is poorly understood. Thus, the main objective of this study was to identify and understand the underlying mechanisms of prolonged cytopenias associated with CAR-T therapy targeting BCMA.

In our work, we combined the retrospective analysis of a cohort of 48 patients treated with BCMA CAR-T cells at Clinica Universidad de Navarra, with ex vivo transcriptomic analysis using single-cell RNA sequencing, to characterize the kinetics of cytopenia, identify predictive factors and determine potential mechanism underlying these toxicities. The overall incidence of cytopenia was 95.7%, and grade>3 thrombocytopenia and neutropenia one month after infusion, was observed in 57% and 53% of the patients, being still present after one year in 4 and 3 patients respectively. Presence of cytopenia at baseline and high peak inflammatory markers highly correlated with cytopenia persisting up to three months.

To determine potential mechanisms underpinning cytopenias, we evaluated the paracrine effect of BCMA CAR-T cells on HSPCs differentiation, using an ex vivo myeloid differentiation model (2). Thus, supernatants from CAR-T cells or control T lymphocytes, co-cultured with MM tumor cell line U266, were added to the differentiation process. Phenotypic analysis by flow cytometry showed that supernatants from activated CAR-T cells (spCAR) halted HSPCs differentiation, promoting more immature phenotypes, with reduced expression of granulocytic, monocytic, and erythroid maturation markers, suggesting a paracrine effect related to CAR-T cells activation. Interestingly, this immature phenotype could be prevented by a combination of IFNγ, TNFα/β, TGFβ, IL-6, and IL-17 inhibitors, indicating the essential role of these cytokines and providing potential targets to prevent these cytopenias. Transcriptomic analyses at single cell level (scRNA-seq), performed using 10X Genomics technology, revealed that cells differentiated in the presence of spCAR were enriched in transcription factors related to early stages of hematopoietic differentiation, such as GATA2 RUNX1 and CEBPA. Additionally, gene regulatory network (GRN) analysis performed with SimiC (3), identified decreased activity of key regulons involved in neutrophil and monocytic maturation, such as MAFB and ID2, providing insights into the molecular mechanisms responsible for the observed phenotypic changes.

Our results suggest that CAR-T cell activation negatively influences hematopoietic differentiation through paracrine effects inducing HSPCs maturation arrest. Moreover, our study contributes to the understanding of mechanisms promoting severe cytopenia observed after CAR-T therapy in MM and provides potential targets to prevent or decrease its severity.

References:

  1. Brudno JN., et al. Nature reviews. (2024).
  2. Berastegui N., et al. Nature Communications (2022).
  3. Peng J., et al. Communications Biology (2022).

    Disclosures: Palacios-Berraquero: Gilead-Kite: Speakers Bureau. Rifon: Amgen: Honoraria, Other: Travel grant. Pierola: AstraZeneca: Research Funding; Abbvie, BMS, Jazz Pharma, Novartis, Syros: Speakers Bureau; Astellas, BMS, Jazz Pharma, Syros: Consultancy. Paiva: Bristol Myers Squibb/Celgene, Janssen, Sanofi, and Takeda: Consultancy; Aztra Zeneca, Bristol Myers Squibb/Celgene, EngMab, Roche, Sanofi, and Takeda: Research Funding; Adaptive, Amgen, Becton Dickinson, Bristol Myers Squibb/Celgene, Janssen, Merck, Novartis, Roche, Sanofi and Takeda: Honoraria. Rodríguez-Otero: Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants; Sanofi: Membership on an entity's Board of Directors or advisory committees, Other: Honoraria for lectures; Johnson & Johnson - Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Honoraria for lectures; Roche: Consultancy; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Honoraria for lectures; Amgen: Other: Honoraria for lectures; Regeneron: Other: Honoraria for lectures; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees, Other: Honoraria for lectures; GSK: Membership on an entity's Board of Directors or advisory committees, Other: Honoraria for lectures; Kite Pharma: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees. San-Miguel: Karyopharm: Other: Advisory board; Bristol Myers Squibb: Other: Advisory board; Amgen: Consultancy, Other: Advisory Board ; Celgene: Other: Advisory board; GlaxoSmithKline: Other: Advisory board; Abbvie: Consultancy, Other: Advisory Board; Novartis: Other; Takeda: Other: Advisory board; Haemalogix: Other: Advisory board; Janssen-Cilag: Other: Advisory board; MSD: Other: Advisory board; Regeneron: Other: Advisory board; Roche: Other: Advisory board; Sanofi: Other: Advisory board; SecuraBio: Other: Advisory board.

    *signifies non-member of ASH