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[Preprint]. 2025 Oct 30:2025.10.28.25338924.
doi: 10.1101/2025.10.28.25338924.

Robust CD4+ CAR T cell Expansion Is Associated with Non-ICANS Neurotoxicities Following Ciltacabtagene Autoleucel

Eric M Jurgens  1   2 , Sneha Mitra  3 , Kevin Herrera  4 , David Nemirovsky  5 , Brenden Bready  5 , Andriy Derkach  5 , Kinga Hosszu  6 , Devin McAvoy  6 , Ross S Firestone  7 , Sridevi Rajeeve  2   7 , Alexander M Lesokhin  2   7 , Neha Korde  7 , Carlyn R Tan  7 , Hamza Hashmi  2   7 , Hani Hassoun  2   7 , Kylee Maclachlan  7 , Urvi A Shah  7 , Malin Hultcrantz  7 , Maximilian Merz  2   7 , Francesco Maura  7 , Sergio A Giralt  1   2 , Gunjan Shah  1   2 , Heather J Landau  1   2 , Michael Scordo  1   2 , Bianca D Santomasso  5 , Jae Park  2 , Christina Leslie  3 , Saad Z Usmani  1   2   7 , Karlo Perica  2 , Sham Mailankody  2   7
Affiliations

Robust CD4+ CAR T cell Expansion Is Associated with Non-ICANS Neurotoxicities Following Ciltacabtagene Autoleucel

Eric M Jurgens et al. medRxiv. .

Abstract

Non-ICANS neurotoxicities (NINTs) are serious, atypical toxicities associated with ciltacabtagene autoleucel, a commercial chimeric antigen receptor (CAR) T cell therapy approved for relapsed/refractory multiple myeloma. Risk factors contributing to the development of NINTs are poorly understood. In a cohort of 109 patients, we identify predisposing risk factors and propose strategies to mitigate NINTs. We show that high peak absolute lymphocyte count is a strong NINT predictor which directly correlates with flow cytometry-based peripheral blood CAR T cell quantitation. The observed CAR lymphocytosis was polyclonal with a bias towards CD4+ CAR T cells rich in memory marker expression. We then identified CAR lymphocytosis associated CD4+ CAR T cell populations which exhibited increased inflammatory pathway gene expression. Finally, we characterize NINT associated CD4+ CAR T cell populations which are potential therapeutic targets for future exploration.

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Conflict of interest statement

T.S - Honoraria: Roche-Genentech. R.S.F. – Grants: ASCO YIA and the IMS. A.M.L. – Grants: Novartis, BMS, Trillium Therapeutics, Pfizer, Janssen; Personal fees: Trillium Therapeutics, Janssen; Patent (number US20150037346A1) with royalties paid. N.K. - Research funding: Amgen, Janssen, Epizyme, AbbVie; Consultancy: Clinical Care Options, OncLive, and Intellisphere Remedy Health; Advisory board: Janssen, MedImmune. C.R.T. – Research funding: Janssen, Takeda. Personal fees: Physician Educations Resource and MJH Life Sciences; Advisory boards: Janssen and Sanofi. H.H. – Grants: Celgene, Takeda, and Janssen. H.H. - Advisory boards: Sanofi, BMS, and Janssen. K.M. – Funding: Sebia, Binding site, and Siemens. U.A.S. - Research support: Celgene/BMS and Janssen; Personal fees: MashUp MD, Janssen Biotech, Sanofi, BMS, MJH Life Sciences, Intellisphere, Phillips Gilmore Oncology Communications, i3 Health, and RedMedEd. M.H. - Research funding: Daiichi Sankyo, Cosette Pharmaceuticals, GlaxoSmithKline, Abbvie, Beigene; Honoraria for consultancy/participated in advisory boards for Curio Science LLC, Projects in Knowledge, Intellisphere LLC, Bristol Myers Squibb, Janssen, and GlaxoSmithKline. S.A.G. - Personal fees and advisory board: Actinium, Celgene, BMS, Sanofi, Amgen, Pfizer, GSK, Jazz, Janssen, Omeros, Takeda, and Kite. G.L.S. - Research funding: Janssen, Amgen, BMS, Beyond Spring; serves on the data safety monitoring board (DSMB) for ArcellX; and receives research funding to the institution from Janssen, Amgen, BMS, Beyond Spring, and GPCR. H.J.L. – Consultancy: Takeda, Genzyme, Janssen, Karyopharm, Pfizer, Celgene, Caelum Biosciences; Research support: Takeda. M.S. – Consultancy: McKinsey & Company, Angiocrine Bioscience, Inc, and Omeros Corporation; Research funding: Angiocrine Bioscience, Inc, Omeros Corporation, and Amgen, Inc; Advisory boards: Kite, a Gilead company; Honoraria: i3 Health, Medscape, and CancerNetwork for CME-related activity. S.Z.U. - Grants and personal fees: AbbVie, 404 Amgen, BMS, Celgene, GSK, Janssen, Merck, Mundipharma, Oncopeptides, 405 Pharmacyclics, Sanofi, Seattle Genetics, SkylineDX, and Takeda. K.P. - Intellectual Property Rights: NexImmune; Professional Services and Activities: Pfizer, Inc.; RxCure LLC S.M. - Consultancy: Evicore, Optum, BioAscend, Janssen Oncology, BMS, AbbVie, HMP Education, and Legend Biotech; Honoraria: OncLive, Physician Education Resource, MJH Life Sciences, and Plexus Communications. Research funding: Janssen Oncology, BMS, Allogene Therapeutics, Fate Therapeutics, Caribou Therapeutics, and Takeda Oncology.

Figures

Fig. 1.
Fig. 1.. Distribution of NINTs amongst patients treated with cilta-cel (n=109).
(A) Number of patients experiencing one or multiple NINTs. (B) Total events observed for each specific NINT amongst affected patients (n=12). ICANS included delayed presentation (n=1) and abnormally prolonged course (n=1). (C) Estimated cumulative incidence of first NINT within 30 days of cilta-cel infusion. Red dotted line indicates the median time to peak ALC (12 days) (D) Swimmer plot depicting the onset and duration of individual NINTs (colored coded on bottom) for each affected patient relative to cilta-cel infusion and duration of response (gray bar). NINT = non-ICANs neurotoxicity; CNP = cranial nerve palsy; GBS = Guillain-Barré Syndrome; ICANS = Immune effector cell-associated neurotoxicity syndrome; PN = peripheral neuropathy; MNT = movement and neurocognitive toxicities; CRS = cytokine release syndrome; CAR = chimeric antigen receptor.
Fig. 2.
Fig. 2.. Peak ALC impacts progression free survival.
(A) PFS of peak ALC groups separated by the identified NINT risk cutoff (3.2×103/μL). (B) PFS of peak ALC groups separated by the identified optimal PFS cutoff (1.6×103/μL). PFS = progression free survival; ALC = absolute lymphocyte count
Fig. 3.
Fig. 3.. ALC correlates with absolute CAR T cells in patients with CL which exhibit bias towards CD4+ CAR T cells expressing memory markers.
(A) Flow cytometry quantitation of absolute CAR+ T cells in CL patient peripheral blood mononuclear cells (PBMC) samples plotted against absolute lymphocyte count (ALC) at corresponding timepoints within 30 days of cilta-cel infusion (n=36). Subplot depicts ALC vs Absolute CAR+ T cells on log-log scale. (B) Box plot comparison of total CAR T cells (CAR+ CD3+ cells) in PBMC samples from non-CL (blue) vs CL patients (red) within Days 7–14 (nnon-CL=34, nCL=22), Days 21–35 (nnon-CL=31, nCL=17), Days 60–90 (nnon-CL=15, nCL=9). (C-H) Box plot comparison of key CAR T cell populations in CL vs non-CL patients as a percentage of total CAR T cells within Days 7–14 (nnon-CL=30, nCL=21), Days 21–35 (nnon-CL=30, nCL=17), Days 60–90 (nnon-CL=14, nCL=9) including (C) CD4+ CAR T cells, (D) CD8+ CAR T cells, (E) CD4+ CD27+ CD28+ CAR T cells, (F) CD8+ CD27+ CD28+ CAR T cells, (G) CD4+ CD57+ CAR T cells, and (H) CD8+ CD57+ CAR T cells. P values were calculated using Mann-Whitney U test and depicted above boxes. CAR = chimeric antigen receptor; ALC = absolute lymphocyte count; CL = CAR lymphocytosis
Figure 4:
Figure 4:. Defining Genetic Determinants of CAR Lymphocytosis.
(A) UMAP projection of scRNA-seq data from CAR T cells from patients (n=16) treated with cilta-cel including patients without CAR lymphocytosis (control; n=7), CAR lymphocytosis with NINT (n=5), and CAR lymphocytosis without NINT (n=4) collected during three time windows; D7–14, D21–30, D70–90. Individual cells are color coded identifying CAR T cells from control vs CAR lymphocytosis patients. (B) T cell receptor (TCR) sequencing ratio to identify frequency and size of CAR T cell clones. (C) 10 distinct clusters were identified, color coded, and labeled according to defining features. (D) Abundance testing using Milo identifies clusters enriched in CAR T cells from patients with CAR lymphocytosis (red) vs control (blue). (E) Dot plot depicting CD8+ and (F) CD4+ CAR T cell gene expression by cluster and grouped by T cell phenotype or function. (G) Surface protein expression by CAR T cell cluster. CAR = chimeric antigen receptor; pALC = peak absolute lymphocyte count; NINT = non-ICANS neurotoxicity; UMAP = Uniform Manifold Approximation and Projection
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References

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