Delayed Neutrophil Recovery Following BCMA-Directed CAR-T Correlates with Poor Outcomes and Is Associated with a Cytokine Signature of Persistent Inflammation and T Cell Checkpoint Activation
Authors: Avigan Zachary M , Bodnar Saoirse , Pan Darren , Aleman Adolfo , Richter Joshua , Sanchez Larysa J , Rodriguez Cesar , Rossi Adriana , Richard Shambavi , Cho Hearn Jay , Jagannath Sundar , Parekh Samir , Thibaud Santiago 
Published: December 2024

Introduction: Chimeric antigen receptor T cells (CAR-T) targeting B-cell maturation antigen (BCMA) have remarkable efficacy in multiple myeloma (MM) and were recently approved for commercial use as early as first relapse. However, a subset of patients develop prolonged cytopenias and delayed absolute neutrophil count (ANC) recovery as a significant source of morbidity. While elevated inflammatory markers are a risk factor for prolonged post-CAR-T neutropenia, specific inflammatory pathways associated with delayed recovery have yet to be fully elucidated.

Methods: We reviewed MM patients at a single academic center receiving BCMA-directed CAR-T with at least 100 days of relapse-free follow-up between 2017 and 2024. Lab values were recorded from lymphodepletion until disease progression. ANC recovery was defined as the first of a consecutive 30-day period with ANC ≥1000 cells/μL without G-CSF use, and platelet (PLT) recovery was defined similarly for PLT ≥50k/μL. Patients were grouped by two timepoints of interest: early (≤30 days) and delayed ANC recovery (&gt;30 days) based on published consensus guidelines of early and late post-CAR-T hematotoxicity (Rejeski et al., Blood 2023), as well as patients who did or did not achieve ANC recovery by day 100 at the time of clinical restaging after cell therapy. Available patient biorepository samples were assayed for serum cytokine measurements using the Olink Target 96 system and compared at serial timepoints.

Results: A total of 160 patients were included with a median of 34 days to ANC recovery. At day 100 post-infusion, 20 patients (13%) had not met ANC recovery criteria. Non-recovered patients were more heavily pretreated (≥4 prior lines: 100% vs 77%, p=.01) and more frequently treatment refractory (triple-class: 95% vs 71%, p=.03; penta-drug: 65% vs 25%, p=.001), showed a trend toward more ISS stage II-III disease (59% vs 33%, p=.057), and had increased incidence of high CAR-HEMATOTOX scores (80% vs 34%, p=.0001). Patients without ANC recovery by 100 days also had shorter median progression-free survival (29 vs 12 months, p=.01) and overall survival (NR vs NR, p=.02). Additionally, 24 patients (15%) did not achieve PLT recovery by day 100, including 15/20 (75%) of those without ANC recovery in the same interval. ANC and PLT recovery times were closely correlated, with a slope of 0.81 for the best-fitting line (R2=.44, p&lt;.0001).

We next examined 654 banked cytokine samples across 47 patients, with a median of 14 measurements per patient from pre-CAR-T screening to day 14. Prior to CAR-T infusion, patients with delayed ANC recovery beyond 30 days (n=26) had increased mean TNFα (p=.01), Granzymes A (p=.03), B (p=.02) and H (p=.01), and T cell chemotactic factors such as CXCL11 (p=.04) and CCL17 (p=.04); a similar inflammatory profile persisted through days 7 and 14. Notably, those with delayed recovery greater than 30 days exhibited a steadily increasing T cell inhibitory profile at sequential timepoints, with elevated soluble checkpoint receptors PD-1 (p=.046) and LAG3 (p=.01) and checkpoint ligands PD-L1 (p=.03), PD-L2 (p=.047), and Galectin-9 (p=.009) by day 14. Additionally, these patients showed progressive markers of endothelial dysfunction with downregulation of ANGPT1 (p=.001), PDGFB (p=.0003), and EGF (p=.002) coupled with increased ANGPT2 (p=.04) and ANGPT2:ANGPT1 ratio (p=.0006), which have previously been associated with prolonged cytopenias, severity of cytokine release syndrome, and coagulopathy (Rejeski et al., Sci Adv 2023; Yang et al., Nat Commun 2024). Finally, we performed an exploratory analysis of patients without ANC recovery by day 100 with available cytokine data (n=8) and showed sustained IL-1α elevation through day 7 (p=.04). As a surrogate for IL-1 in the larger cohort, we evaluated serial ferritin values and showed that non-recovered patients by day 100 had elevated ferritin at baseline (p=.02) that persisted through days 14 (p=.047), 30 (p=.02), 60 (p=.045), and 100 (p=.06).

Conclusions: Prolonged neutropenia following BCMA CAR-T is more prevalent in heavily pretreated patients and correlates with decreased survival. Patients with delayed ANC recovery showed a characteristic cytokine profile of persistent inflammation culminating in increased T cell inhibitory checkpoint markers, endothelial dysfunction, and prolonged IL-1α and ferritin elevation, highlighting possible targets for therapeutic investigation.