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2051 Deciphering the Impact of Immunological Aging in CLL CAR T Cell Therapy

Program: Oral and Poster Abstracts
Session: 702. CAR-T Cell Therapies: Basic and Translational: Poster I
Hematology Disease Topics & Pathways:
Research, Translational Research, Chimeric Antigen Receptor (CAR)-T Cell Therapies, Treatment Considerations, Biological therapies, Immunology, Biological Processes
Saturday, December 7, 2024, 5:30 PM-7:30 PM

Julia Han Noll1,2,3*, Humza Hemani4*, Jiang Yuan Li, PhD4*, Kevin R. Amses, PhD3,5,6*, Bruce Levine, PhD2,3,7, Carl H. June, MD2,3,8,9, Nan-Ping Weng, PhD4* and Joseph A. Fraietta, PhD3,5,10*

1Department of Microbiology, University of Pennsylvania, Perelman School of Medicine, Philadelphia, PA
2Center for Cellular Immunotherapies, University of Pennsylvania, Philadelphia, PA
3Parker Institute for Cancer Immunotherapy, University of Pennsylvania, Perelman School of Medicine, Philadelphia, PA
4National Institute On Aging, National Institutes of Health, Baltimore, MD
5Center for Cellular Immunotherapies, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA
6Department of Microbiology, University of Pennsylvania, Philadelphia, PA
7Department of Pathology and Laboratory Medicine, University of Pennsylvania, Perelman School of Medicine, Philadelphia, PA
8Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA
9Abramson Cancer Center, Department of Medicine, Division of Hematology-Oncology, University of Pennsylvania, Philadelphia, PA
10Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA

Chronic Lymphocytic Leukemia (CLL) primarily affects older adults, with a median diagnosis age range of 65-72 years. CAR T cell therapy shows high complete response (CR) rates >80% in pediatric acute lymphoid leukemia, yet only under 30% in CLL. CAR T cells in non-responding (NR) CLL patients upregulate pathways for effector differentiation and exhaustion. However, immune checkpoint blockade doesn't change CLL progression, suggesting exhaustion alone doesn't explain CLL T cell deficiency. We hypothesize that immunosenescence also contributes to CAR T cell defects in CLL.

19CAR-BBz T cells were generated from 11 treatment naïve CLL patients and age- and gender-matched healthy controls with a median age of 67. Immunophenotyping was performed with flow cytometric panels including 25 markers. We conducted a chronic antigen stress test: every five days, 1E6 CAR T-cells were counted, phenotyped, and stimulated with fresh CD19-expressing K562 cells at a 1:1 ratio. 24 hours after co-culture, supernatant was collected for cytokine analysis using LEGENDplex™ kits. Senescence signatures and clinical characteristics in CLL patients enrolled in CAR T cell trials were contrasted using ssGSEA analysis of bulk RNA sequencing data.

CAR T cells derived from treatment-naïve CLL patients exhibit a decrease in co-stimulatory receptors CD27 and CD28 expression alongside heightened levels of senescence markers p16, p21, p53, β-galactosidase and γ-H2AX. The ex vivo manufacturing process solidifies senescent trends seen in baseline CLL T cells compared to healthy control. CD28 is instrumental in predicting CLL from control in a random forest prediction model. In functional assays, the proliferative capacity of CLL patient derived CAR T cells was attenuated and production of IFN-γ was decreased. However, CLL-derived CAR T cells showed signatures of inflamm-aging such as increased expression of Granzyme A, B and perforin and production of pro-inflammatory factors such as sFas and IL-1β. In the clinical trial subjects, several hallmarks of immunosenescence were found in T cells from NR CLL patients at leukapheresis and ex vivo manufacturing harvest such as the loss of co-stimulatory receptors CD27/CD28, KLRG1 upregulation and decrease in TCR diversity. Enrichment of senescence pathways is also able to differentiate therapeutic outcome, and correlates negatively with expansion during CAR T cell manufacturing, maximum in vivo CAR T cell expansion and overall survival of patients. Furthermore, a machine learning model developed by Lu et al. (Nature Communications, 2022) predicted higher age in NR CLL patients compared to CR CLL patients based on their CAR T cell transcriptome.

Our study underscores the role of immunosenescence in reducing CAR T cell efficacy in CLL. Senescence signatures such as CD27/CD28 loss and enrichment in senescent pathways should be considered for patient stratification to enhance therapeutic success. Addressing immunosenescence is critical for improving CAR T cell therapy in CLL.

Disclosures: Levine: In8bio: Membership on an entity's Board of Directors or advisory committees; Ori Biotech: Membership on an entity's Board of Directors or advisory committees; Oxford Biomedica: Membership on an entity's Board of Directors or advisory committees; ThermoFisher Pharma Services: Membership on an entity's Board of Directors or advisory committees; UTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; Tmunity Therapeutics: Current equity holder in publicly-traded company; Capstan Therapeutics: Current equity holder in private company; Immusoft: Membership on an entity's Board of Directors or advisory committees; Immuneel: Membership on an entity's Board of Directors or advisory committees; Avectas: Membership on an entity's Board of Directors or advisory committees. Fraietta: Tceleron Therapeutics, Inc: Membership on an entity's Board of Directors or advisory committees; OverT Bio, Inc: Membership on an entity's Board of Directors or advisory committees; CellFe Biotech: Membership on an entity's Board of Directors or advisory committees; Shennon Biotechnologies Inc.: Membership on an entity's Board of Directors or advisory committees; Cartography Bio: Membership on an entity's Board of Directors or advisory committees; Retro Biosciences: Consultancy; Tmunity Therapeutics: Research Funding; Danaher Corporation: Research Funding.

*signifies non-member of ASH