Chimeric antigen receptor (CAR) T-cell therapy holds great promise for patients with cancer, and the identification of predictive biomarkers is crucial in finding new ways to guide therapy. Major challenges to the application of informatics and machine learning in CAR T-cell therapy include limited sample sizes and non-uniformity in data generation across cancer indications and trials. Here we took a global, pan-haematologic cancer approach, analysing 256 patients across 5 cancer types and 13 clinical trials. We generated data using a framework that included pre-infusion clinical features, over 2 million apheresis T cells analysed by flow cytometry using 17 unique markers, ex vivo T-cell expansion during CAR T-cell manufacture, more than 90,000 measurements of 30 serum markers and serial tracking of circulating CAR T cells using qPCR. From this data resource, we demonstrate the potential of pan-cancer predictive biomarkers that capture generalizable characteristics of treatment response and non-response in CAR T-cell therapy.

HLA sensitization poses a major challenge to kidney transplantation for patients with end-stage kidney disease, especially for highly sensitized candidates. Attempts at antibody elimination (desensitization) have had inconsistent efficacy and have often failed to produce sustained reductions in anti-HLA antibodies in patients with the highest level of sensitization (calculated panel-reactive antibody score, ≥99.9%). We now report the results for the safety run-in cohort of a multicenter phase 1 clinical study evaluating the safety and efficacy of combined CD19-targeted and B-cell maturation antigen (BCMA)-targeted chimeric antigen receptor (CAR) T cells in eliminating the cellular sources of preformed anti-HLA antibodies (ClinicalTrials.gov number, NCT06056102). Kidney transplantation was performed in two highly sensitized candidates after desensitization with the use of dual CAR T-cell therapy.

BACKGROUND:Teclistamab is the first approved bispecific antibody targeting B-cell maturation antigen. It has demonstrated rapid, deep, durable responses with manageable safety in patients with triple-class exposed relapsed/refractory multiple myeloma (TCE RRMM). METHODS:The authors report results from three cohorts: Global Trial cohort consisting of 217 patients from three registrational studies (pivotal MajesTEC-1 study [n = 165], China cohort of MajesTEC-1 [n = 26], and the Japan MMY1002 study [n = 26]); the subset of 52 of 217 patients formed the Asian Trial cohort, and 42 patients treated outside of trials in the pre-approval access (PAA) program formed the Asian PAA cohort. RESULTS:In the Global Trial cohort, median age was 65 years, weight was 69 kg, and prior lines-of-therapy was five; 29.0% had high-risk cytogenetics and 18.9% had extramedullary disease. With 29.5 months median follow-up, overall response rate (ORR)/≥complete response (CR) was 66.4%/50.2%, median duration of response (DOR) was not reached; progression-free survival (PFS) and overall survival (OS) were 15.6, and 29.1 months, respectively. In the subset of Asian Trial cohort, baseline features were similar except for lower weight (median, 58 kg); median follow-up was 26.3 months. ORR/≥CR was 76.9%/63.5%, 24-month DOR, PFS, and OS rates were 67.5%, 59.5%, 71.4%, respectively, with medians not yet reached. Efficacy was consistent in the Asian PAA cohort with ORR/≥CR of 66.7%/40.5%. Most common adverse events were cytopenias, cytokine release syndrome, and infections. Infection management improved over time, supported by increased immunoglobulin use in later-enrolling Asian studies, aligned with guideline adoption. CONCLUSION:Teclistamab demonstrated clinically meaningful benefits across diverse patients, encompassing various weight categories and geographies, reinforcing its potential as a standard of care for TCE RRMM. TRIAL REGISTRATION:ClinicalTrials.gov MajesTEC-1 (NCT03145181 and NCT04557098) and Japan MMY1002 study (NCT04696809).

INTRODUCTION/AIMS:Chimeric antigen receptor T-cell (CAR-T) therapy has transformed the management of refractory hematologic malignancies, but neurologic complications remain a major concern. While immune effector cell-associated neurotoxicity syndrome is well recognized, peripheral neuropathies such as neuralgic amyotrophy have rarely been described. METHODS:We characterize the clinical, electrophysiologic, and imaging features of neuralgic amyotrophy after B-cell maturation antigen (BCMA)-directed CAR-T therapy at a single tertiary care center. RESULTS:We identified four patients with neuralgic amyotrophy following BCMA-directed CAR-T therapy, corresponding to 1.5% of total patients treated in that time. Patients developed acute bilateral shoulder pain followed by weakness 8-12 days after CAR-T infusion. Manifestations included scapular winging and arm weakness, with electromyography showing denervation most prominently in the muscles supplied by the long thoracic and suprascapular nerves. Corticosteroid use appeared to be associated with symptomatic improvement, although residual weakness persisted in most. Diagnosis was delayed in three patients due to misattribution of symptoms to other causes. DISCUSSION:Neuralgic amyotrophy may represent an underrecognized complication of BCMA-directed CAR-T therapy. The consistent timing of onset, bilateral involvement, and responsiveness to corticosteroids support an inflammatory mechanism. Clinicians should maintain vigilance for this complication, as prompt recognition and treatment may mitigate long-term neurological morbidity.

Lentiviral vectors are commonly used to introduce chimeric antigen receptor transgenes into T cells, but routine assays quantify vector copy number or integration sites without sequencing full-length integrated vectors. HIV-1 proviruses often acquire large deletions and cytidine deaminase-driven hypermutation; whether similar variation occurs in therapeutic lentiviral vectors is unclear. We adapted a novel long-read capture approach to enrich long fragments spanning vector DNA and adjacent human sequence, enabling simultaneous integration-site mapping and proviral integrity analysis with single-molecule resolution. In research-grade CAR T cells produced with an experimental, transient-transfection lentiviral vector workflow, 40% of integrated vectors carried recurrent deletions that removed the internal promoter or parts of the chimeric antigen receptor cassette. The dominant promoter deletion was present in the viral stock. In clinical chimeric antigen receptor T cell products, promoter deletions were less frequent, but detectable pre-infusion and post-infusion. Across datasets we observed widespread G-to-A substitutions consistent with restriction factor editing, including changes predicted to introduce premature stop codons within the transgene open reading frame. Our method reveals proviral variants invisible to standard quality-control assays and provides a framework to improve vector production and monitor transgene integrity in clinical products.

Abstract Multiple myeloma remains a fatal, incurable disease. Most therapies are targeted to the cancer cell or T cell engagement. Little is known about the supporting myeloma microenvironment and its contribution to tumor fitness. Here, we expand upon the observation of human mast cells in the NSG-hIL6 myeloma patient derived xenograft mouse model to show mast cells decrease time to engraftment, promote increased myeloma engraftment and cause myeloma bone disease. We identify 10 mast cell secreted factors that together improve the survival of patient myeloma cells in vitro . Our results highlight the versatility of the NSG-hIL6 model to study microenvironmental interactions between human bone marrow cells and myeloma and confirm prior suggestions that clinical signs of disease, such as osteolytic lesions, may at least partially be related to non-malignant bone marrow microenvironmental cells, such as mast cells.

CD19-directed CAR T-cells (CTL019) can produce durable remissions in chronic lymphocytic leukemia (CLL), but therapeutic success depends on whether autologous T-cells expand, persist, and retain cytotoxic function after manufacturing. Failure of CLL T-cells is often attributed to exhaustion, although many dysfunctional CLL T-cells retain inflammatory cytokine production. We tested whether this paradox reflects immunosenescence, an aging-like program defined by costimulatory loss, DNA damage, inflammatory secretion, repertoire restriction, and proliferative arrest. In response-linked preinfusion CTL019 products, nonresponders and short partial responders had higher senescence and senescence-associated secretory phenotype (SASP) programs than the functional-responder group, comprising complete responders and partial responders with transformed disease. These programs were detectable before infusion and became more prominent during manufacture in susceptible products. They tracked with weak in vivo CAR T-cell expansion and were associated with inferior survival. Serum proteomics revealed a circulating, CLL-associated SASP-like inflammatory milieu in nonresponders. Flow cytometry and T-cell receptor (TCR) profiling showed that poor responders carried CD27⁻CD28⁻ and KLRG1⁺ CD8 T-cells at apheresis, lower product CD27, and reduced product TCR diversity. Research-manufactured CAR T-cells from treatment-naïve CLL samples displayed the same state, most prominently in CD4 CAR T-cells, with SA-β-gal, p16, p53, DNA damage, depletion of less-differentiated states, and senescence-enriched transcriptomes. Under repeated CD19 stimulation, CLL-derived products reached a proliferative ceiling, and p53-high products lost cytotoxic reserve. Ibrutinib improved proliferative fitness, attenuated senescence-associated features and SASP output in paired patient and direct-exposure assays, and enhanced CAR T-cell expansion in an ibrutinib-resistant CLL model. Together, these data identify immunosenescence as a measurable and functionally consequential barrier to CAR T-cell efficacy in CLL and a candidate for therapeutic modulation.

We show continuous tumor exposure results in a loss of chimeric antigen receptor (CAR) T cell (CART) endocytic activity due to downregulation of Rab5. Loss of endocytic activity exacerbates the effects of trogocytosis, the bidirectional transfer of tumor target antigens and CARs between malignant cells and CARTs, resulting in CART dysfunction and fratricide. Constitutive expression of Rab5 within the CARTs reduced fratricide by reducing the amount of trogocytosed antigens on the cell surface, while simultaneously enhancing CAR availability through dissociation of CAR from target, recycling unbound CAR back to the plasma membrane, and limiting CAR capture by tumor cells. Rab5-expressing CARTs exhibited superior antitumor activity in both BCMA-CARTs isolated from the bone marrow of treated patients and mesothelin-specific CARTs in a solid tumor model. These studies uncover an unexpected relationship between endocytosis and CART function and suggest that pairing Rab5 with CAR expression could improve the clinical efficacy of CART therapy.

Teclistamab is a B cell maturation antigen × CD3 bispecific antibody approved for relapsed/refractory multiple myeloma. Two step-up doses (SUDs) are used to mitigate the risk of cytokine release syndrome (CRS). For patients who experience dose delays, it is uncertain what length of delay necessitates repeat SUDs. We used modeling simulations and retrospective analysis of the phase 1/2 MajesTEC-1 study to optimize recommendations for repeat SUDs after teclistamab dose delay. Population pharmacokinetic modeling was used to simulate teclistamab serum concentrations after dose delays to assess the duration required to achieve levels comparable to estimated trough concentrations (Ctrough) following SUDs. Quantitative systems pharmacology modeling was used to simulate cytokine dynamics. Modeling-informed time windows were applied to a retrospective analysis of CRS data from MajesTEC-1 recommended phase 2 dose cohorts to further evaluate CRS incidence with prolonged dose delays (> 28 days). Median teclistamab serum concentrations were estimated to drop to levels comparable to the simulated SUD 2 median Ctrough after 62 days and SUD 1 median Ctrough after 111 days. Simulated cytokine peaks at treatment restart during weekly or biweekly dosing at these intervals were lower than those following the initial SUD. Retrospective analysis of clinical data revealed a low incidence of CRS (grade 1–2; 2/61 [3.3

B cell maturation antigen (BCMA)-targeted chimeric antigen receptor (CAR) T cell therapy has revolutionized the treatment of multiple myeloma but can cause unique toxicities, including cranial nerve palsy, parkinsonism and enterocolitis, which we refer to collectively as CAR T cell therapy-associated immune-related adverse events (CirAEs). Among 198 patients treated with ciltacabtagene autoleucel or idecabtagene vicleucel (June 2021–December 2024), 27 (13.6

Supplemental tables: (1) Subject characteristics. (2) Cytogenetic profiles and high-risk features. (3) Prior treatment exposures and refractoriness. (4) CAR T cell product characteristics. (5) Products that did not meet target dose. (6) Adverse events of grade 3-4. (7) Cytokine release syndrome and ICANS. (8). Maintenance therapy. Supplemental Figures: (1) Study schematic and subject disposition, (2) Correlates of manufacturing success, (3) Hematopoietic recovery, (4) Post-infusion T cell phenotypes, (5) Correlates of in vivo expansion and manufacturing success, (6) Late post-infusion CAR T cell re-expansion, (7) Soluble BCMA, (8) Late-onset clinical responses, (9) MM cell BCMA expression, (10) Pre- and post-treatment Sox2-specific T cell responses in CART-BCMA monotherapy patients, (11) Pre- and post-treatment Sox2-specific T cell responses in CART-BCMA + huCART19 combination therapy patients, (12) Sustained post-treatment SOX2-specific T-cell responses.

7518 Background: The two approved BCMA-targeted CAR-T products, cilta-cel and ide-cel, have significant efficacy in RRMM, but are not considered curative. Initial studies in ≥4 th line RRMM required progressive disease (PD) at time of enrollment and T cell apheresis. We hypothesized that using CAR-T cells as a planned consolidation strategy (i.e. in patients (pts) with stable or responsive disease on their current therapy) may lead to lower toxicity and better long-term disease control. Methods: We conducted a retrospective review of all RRMM pts receiving commercial CAR-T cells at the University of Pennsylvania from 6/1/21 to 4/30/24, with at least 6 months of follow-up. Intent for consolidation was retroactively assigned by chart review. Kaplan-Meier methodology was used to determine PFS and OS. Results: We identified 149 pts for analysis, with a median follow-up of 14.4 months (mos). Median prior lines was 6 and 81% of pts were triple class-refractory; 46% had high-risk cytogenetics, 26% had extramedullary disease, and 17% had prior BCMA-directed therapy. Pts received either cilta-cel (54%) or ide-cel (46%), and 95% received bridging therapy. CAR-T cells were intended as planned consolidation in 51 pts (34%); of these, 36 (71%) had ≥PR at time of apheresis. For consolidation vs non-consolidation groups, this translated into greater depth of response post-CAR-T cells (≥VGPR, 86% vs. 66%, p=0.01), lower rates of ≥grade 3 CRS (1.9% vs. 9.1%, p=0.16), and longer PFS (median not reached vs. 10 mos, p=0.001), respectively. The PFS improvement was seen for both cilta-cel (p=0.01) and ide-cel (p=0.04). No differences in neurotoxicity were noted. We also performed analyses based on response at apheresis, regardless of intent (8% ≥VGPR, 23% PR, 27% stable disease (SD), and 42% PD). PFS at 20 mos was 88%, 47%, 55%, and 31% for ≥VGPR, PR, SD, and PD at apheresis, respectively (p=0.015). Median PFS of pts with at least SD (≥SD) at apheresis was not reached vs. 9.4 mos in those with PD (p= 0.003), with 20-month OS of 87% in the ≥SD group and 68% in the PD group (p=0.015). Subgroup analysis confirmed this PFS difference for both cilta-cel and ide-cel, while the OS impact was only seen for cilta-cel. On multivariate analysis, having ≥SD at apheresis was an independent predictor for PFS. No statistically significant differences in CRS and ICANS were observed based on response at apheresis. Pts with ≥SD at apheresis had higher absolute lymphocyte counts at days 7 and 14 post-CAR-T infusion than those with PD, indicating disease status at apheresis may be associated with CAR-T product quality. Conclusions: Our data suggest that disease control (≥SD) at time of T-cell collection is associated with more durable responses, supporting use of CAR-T cells as a consolidation strategy in RRMM. We cannot conclude these associations are causal. Further analyses of apheresed T cell characteristics are planned.

Supplemental Table 1: Reasons for MajesTEC-1 ineligibility

CONTEXT:Caregivers are essential in the care of CAR T-cell patients, especially immediately before and after CAR T-cell therapy. However, the long-term CAR T-cell therapy caregiving implications are understudied. OBJECTIVES:We aimed to characterize long-term caregiver health-related quality of life (HRQoL) and caregiving burden and understand the relationship between long-term caregiving burden and patient-reported HRQoL, cognitive function, and symptom burden. METHODS:This cross-sectional study included patients (≥18 years of age) who had received CAR T-cell therapy for B-cell lymphoma or multiple myeloma in the past one to five years and were in remission. Participating patients identified their current caregiver or the individual who was their caregiver at the time of infusion. Patients completed a survey with measures of HRQoL, cognitive function and symptom burden, while caregivers completed measures of HRQoL and caregiving burden. Linear regression was used to identify predictors of caregiving burden. RESULTS:In total, 58 patients and 31 caregivers participated. Caregiver HRQoL was similar to population norms, and 26% of caregivers reported mild to moderate caregiving burden. Caregiver age, and patient mental HRQoL, cognitive function, and symptom burden were significantly associated with caregiving burden in bivariate analyses (P < .05). Caregiver age was a significant predictor of caregiving burden in the linear regression analysis. CONCLUSION:Caregivers of patients in sustained remission 1 to 5 years after CAR T-cell therapy report good HRQoL and minimal caregiving burden. However, clinicians should be aware that younger caregivers and caregivers of patients with worse symptom burden, cognitive function and HRQoL may experience increased caregiving burden.

Supplemental Table S8: Multivariable regression for overall response

Data describing outcomes of teclistamab in multiple myeloma patients with prior exposure to BCMA-directed therapy (BCMA-DT) are limited. The goal of this multicenter retrospective analysis was to report the efficacy and safety of standard-of-care teclistamab in patients with prior BCMA-DT. A total of 385 patients were included, of whom 193 (50%) had received prior BCMA-DT, including 47 (24%) patients with prior antibody-drug conjugate (ADC)-only, 99 (51%) with chimeric antigen receptor T-cell therapy (CAR T)-only, 36 (19%) with both ADC and CAR T, 6 (3%) with bispecific antibody-only, and 5 (3%) with other combinations. Most safety parameters between cohorts were comparable. The prior BCMA-DT cohort had a lower overall response rate (ORR: 48.7% versus 61.5%; p = 0.012), and median progression-free survival (PFS: 4.6 versus 8.2 months; p = 0.017) compared to the cohort without prior BCMA-DT. However, in multivariable analysis, despite a clear trend, ultimately receipt of a prior BCMA-DT was not independently associated with ORR or PFS (p = 0.057 and p = 0.1, respectively). No significant differences in PFS were noted when stratifying patients by number of prior BCMA-DTs, types of all prior BCMA-DTs received, type of most recent prior BCMA-DT, or depth of response to most recent BCMA-DT. Using the maximally selected rank statistics method, the optimal cut-off for time from the last BCMA-DT exposure to teclistamab initiation was identified as 8.7 months. Patients with >8.7 months between their last exposure to prior BCMA-DT and teclistamab initiation had a significantly improved median PFS with teclistamab (8.1 months, 95% CI: 4.6-11.7) compared to patients with <8.7 months (2.5 months, 95% CI: 1.1-5.7), p = 0.001. Altogether, our findings support the use of teclistamab as a viable treatment option in patients previously exposed to BCMA-DT.