Single-cell analysis of anti-BCMA CAR T cell therapy in patients with central nervous system autoimmunity.

Chimeric antigen receptor (CAR) T cell immunotherapy for the treatment of neurological autoimmune diseases is promising, but CAR T cell kinetics and immune alterations after treatment are poorly understood. Here, we performed single-cell multi-omics sequencing of paired cerebrospinal fluid (CSF) and blood samples from patients with neuromyelitis optica spectrum disorder (NMOSD) treated with anti-B cell maturation antigen (BCMA) CAR T cells. Proliferating cytotoxic-like CD8+ CAR T cell clones were identified as the main effectors in autoimmunity. Anti-BCMA CAR T cells with enhanced features of chemotaxis efficiently crossed the blood-CSF barrier, eliminated plasmablasts and plasma cells in the CSF, and suppressed neuroinflammation. The CD44-expressing early memory phenotype in infusion products was potentially associated with CAR T cell persistence in autoimmunity. Moreover, CAR T cells from patients with NMOSD displayed distinctive features of suppressed cytotoxicity compared with those from hematological malignancies. Thus, we provide mechanistic insights into CAR T cell function in patients with neurological autoimmune disease.

B cell lineage reconstitution underlies CAR-T cell therapeutic efficacy in patients with refractory myasthenia gravis.

B-cell maturation antigen (BCMA), expressed in plasmablasts and plasma cells, could serve as a promising therapeutic target for autoimmune diseases. We reported here chimeric antigen receptor (CAR) T cells targeting BCMA in two patients with highly relapsed and refractory myasthenia gravis (one with AChR-IgG, and one with MuSk-IgG). Both patients exhibited favorable safety profiles and persistent clinical improvements over 18 months. Reconstitution of B-cell lineages with sustained reduced pathogenic autoantibodies might underlie the therapeutic efficacy. To identify the possible mechanisms underlying the therapeutic efficacy of CAR-T cells in these patients, longitudinal single-cell RNA and TCR sequencing was conducted on serial blood samples post infusion as well as their matching infusion products. By tracking the temporal evolution of CAR-T phenotypes, we demonstrated that proliferating cytotoxic-like CD8 clones were the main effectors in autoimmunity, whereas compromised cytotoxic and proliferation signature and profound mitochondrial dysfunction in CD8+ Te cells before infusion and subsequently defect CAR-T cells after manufacture might explain their characteristics in these patients. Our findings may guide future studies to improve CAR T-cell immunotherapy in autoimmune diseases.

Single-cell analysis of refractory anti-SRP necrotizing myopathy treated with anti-BCMA CAR-T cell therapy.

Immune-mediated necrotizing myopathy (IMNM) is an autoimmune disorder associated with the presence of autoantibodies, characterized by severe clinical presentation with rapidly progressive muscular weakness and elevated levels of creatine kinase, while traditional pharmacological approaches possess varying and often limited effects. Considering the pathogenic role of autoantibodies, chimeric antigen receptor (CAR)-T cells targeting B cell maturation antigen (BCMA) have emerged as a promising therapeutic strategy. We reported here a patient with anti-signal recognition particle IMNM refractory to multiple available therapies, who was treated with BCMA-targeting CAR-T cells, exhibited favorable safety profiles, sustained reduction in pathogenic autoantibodies, and persistent clinical improvements over 18 mo. Longitudinal single-cell RNA, B cell receptor, T cell receptor sequencing analysis presented the normalization of immune microenvironment after CAR-T cell infusion, including reconstitution of B cell lineages, replacement of T cell subclusters, and suppression of overactivated immune cells. Analysis on characteristics of CAR-T cells in IMNM demonstrated a more active expansion of CD8+ CAR-T cells, with a dynamic phenotype shifting pattern similar in CD4+ and CD8+ CAR-T cells. A comparison of CD8+ CAR-T cells in patients with IMNM and those with malignancies collected at different timepoints revealed a more NK-like phenotype with enhanced tendency of cell death and neuroinflammation and inhibited proliferating ability of CD8+ CAR-T cells in IMNM while neuroinflammation might be the distinct characteristics. Further studies are warranted to define the molecular features of CAR-T cells in autoimmunity and to seek higher efficiency and longer persistence of CAR-T cells in treating autoimmune disorders.

A novel two-step administration of XPO-1 inhibitor may enhance the effect of anti-BCMA CAR-T in relapsed/refractory extramedullary multiple myeloma.

BACKGROUND: Extramedullary disease usually implies a dismal outcome in relapsed/refractory multiple myeloma patients, and requires novel treatment approaches. We designed a trial using Selinexor, a nuclear export protein 1 inhibitor, together with anti-B cell maturation antigen (BCMA) chimeric antigen receptor (CAR)-T cell product CT103A to treat these patients, and describe the first two cases in this report. METHODS: Selinexor was administered with a novel two-step schedule in bridging therapy and in maintenance. The clinical responses and adverse events were recorded after CAR-T infusion and Selinexor administration. In vitro analysis of the influence of Selinexor on CAR-T cell function was performed using myeloma cell lines. RESULTS: After infusion, both patients achieved stringent complete remission (sCR), and were maintained in sCR at data-cutoff, with survival over 13 and 10 months, respectively. Neither immune effector cell-associated neurotoxicity syndrome nor over grade 2 cytokine release syndrome was observed. Meanwhile, the patients showed good tolerance to the combination. In addition, we demonstrated that low dose of Selinexor could upregulate the expression of BCMA on plasma cell lines and subsequently enhance the function of CAR-T cell in vitro. CONCLUSIONS: The combination of Selinexor and CT103A exerts preliminary synergistic effect, and can be developed as a promising strategy for relapsed/refractory extramedullary myeloma.

Anti-BCMA CAR T-cell therapy CT103A in relapsed or refractory AQP4-IgG seropositive neuromyelitis optica spectrum disorders: phase 1 trial interim results.

Chimeric antigen receptor (CAR) T-cell therapy that targets B-cell maturation antigen (BCMA) have great potentials in autoimmune diseases and could be novel therapeutics for relapsed/refractory neuromyelitis optica spectrum disorder (NMOSD). To evaluate the safety and efficacy of the CT103A, a self-developed BCMA-targeting CAR construct against BCMA, in patients with AQP4-IgG seropositive NMOSD, an ongoing, investigator-initiated, open-label, single-arm, phase 1 clinical trial is conducted at our center. In total, 12 patients were administered with a CAR-BCMA infusion. Ten of the 12 patients dosed were women (83.3%), with a median age of 49.5 years (range, 30-67). were The most common events of grade 3 or higher were hematologic toxic effects. Seven patients (58%) developed infections, but no grade 4 infections occurred. Cytokine release syndrome was reported in all patients with only events of grade 1 or 2 observed. During the follow-up of a median 5.5 months, 11 patients had no relapse; all patients generally reported improvement in disabilities and quality-of-life outcomes; 11 patients' AQP-4 antibodies in serum showed a downward trend by the cutoff date. CAR T-cell expansion was associated with responses, and persisted more than 6 months post-infusion in 17% of the patients. In summary, CAR T-cell therapy shows a manageable safety profile and therapeutic potentials for patients with relapsed/refractory AQP4-IgG seropositive NMOSD. Another expansion phase is currently underway to determine the safety and efficacy of CAR T-BCMA infusion in patients with other neuro-inflammatory diseases.

Delivery of adipose-derived mesenchymal stem cells attenuates airway responsiveness and inflammation in a mouse model of ovalbumin-induced asthma.

Adipose-derived mesenchymal stem cells (ADMSCs) possess immunomodulation property, yet their therapeutic potential in asthma is unclear. This study aimed to explore the effects of ADMSCs on airway hyperresponsiveness and inflammation in ovalbumin (OVA)-induced asthma models. The underlying mechanism(s) was also examined. BALB/c mice were sensitized with OVA on days 0, 7, and 14, followed by 8-week OVA challenge from day 22. ADMSCs were injected via tail vein on day 21. Animals were measured for airway responsiveness, lung pathology, IgE and cytokine levels in serum, cell composition in bronchoalveolar lavage fluid (BALF), gene expression in the lung, and regulatory T cells (Tregs). We found that delivery of ADMSCs decreased airway responsiveness and eosinophil counts in BALF and reduced infiltration of inflammatory cells and number of mucus-expressing goblet cells in the lung in OVA-challenged mice. OVA-evoked elevation of serum IgE levels and alteration of cytokine production in serum and BALF was significantly prevented by ADMSCs. In addition, administration of ADMSCs impaired the regulation of lung IL-10, Foxp3, IL-17, and RORγ expression by OVA challenge and restored the percentage of CD4+CD25+Foxp3+ Tregs in the spleen. In conclusion, ADMSCs confer protection against OVA-induced airway hyperresponsiveness and inflammation, which is associated with induction of Tregs and restoration of immune homeostasis. These findings suggest that ADMSCs may have therapeutic implications for allergic asthma.

[Medicated serum prepared with Chinese herbal medicine Zhizhen Recipe down-regulates activity of nuclear factor-κB and expression of P-glycoprotein in human colorectal cancer multidrug-resistant cell line HCT-8/VCR].

OBJECTIVE: To investigate the effects of medicated serum prepared with Chinese herbal medicine Zhizhen Recipe (ZZR) on activity of nuclear factor-κB (NF-κB) and expression and function of P-glycoprotein (P-gp) in human colorectal cancer multidrug-resistant cell line HCT-8/VCR. METHODS: The multidrug resistance of HCT-8/VCR cells was detected by cell counting kit-8 method, and the experimental concentrations of ZZR-medicated serum were determined by the same way. HCT-8 and HCT-8/VCR cells were treated with ZZR-medicated serum of medium dose for 24 h. The activity of NF-κB was determined by enzyme-linked immunosorbent assay. The intracellular distribution of P-gp was detected by laser scanning confocal microscopy, and the mean fluorescence intensity of rhodamine 123 was detected by flow cytometry. RESULTS: ZZR-medicated sera with volume fraction of 8%, 16% and 32% of medium dose were confirmed as the experimental sera. Compared with the untreated group, NF-κB activities of the ZZR-medicated serum groups (ZZR-medicated serum with volume fraction of 8%, 16% and 32% of medium dose) were obviously down-regulated (P<0.01), which had a negative correlation with the concentrations. After interfering HCT-8/VCR with ZZR-medicated serum of different concentrations for 24 h, P-gp in HCT-8/VCR transmitted gradually from cell membrane to cytoplasm and nuclei. Nuclei became pyknotic and cracking. Compared with the untreated group, the mean fluorescence intensities of ZZR-medicated serum groups declined with concentration gradients (P<0.01). The efflux of intracellular rhodamine 123 decreased, the wave crest shifted to right, and the intracellular fluorescence intensity strengthened (P<0.01). CONCLUSION: ZZR-medicated sera of experimental concentrations down-regulate activity of NF-κB and expression and function of P-gp in human colorectal cancer multidrug-resistant cell line HCT-8/VCR and the effect is related to the concentrations.