Challenges and strategies in clinical applications of CAR-T therapy for autoimmune diseases.

Introduction

Autoimmune diseases (ADs) represent a clinically heterogeneous group of disorders characterized by the breakdown of immunological tolerance, wherein aberrant activation of adaptive and innate immunity mediates tissue-specific or systemic damage [ 1 ]. Defective B-cell maturation and activation promote autoantibody generation [ 2 ], while dysfunctional T-cell responses and aberrant phagocyte (macrophage/neutrophil) activity exacerbate immune-mediated tissue injury [ 3 – 5 ]. ADs include both multisystem disorders [e.g., systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), multiple sclerosis (MS)] and organ-specific conditions such as type 1 diabetes mellitus (T1DM) among others [ 2 , 6 ]. Conventional therapeutic strategies primarily involve immunosuppression (e.g., corticosteroids, azathioprine, calcineurin inhibitors) and targeted immunomodulation blocking TNF-α, IL-6, or utilizing B-cell depletion therapies [ 7 , 8 ]. Although these immunosuppressive regimens effectively control disease activity, their prolonged use inevitably increases susceptibility to infectious complications [ 9 ].

Chimeric antigen receptor (CAR) T-cell therapy has produced remarkable results in the treatment of hematological malignancies [ 10 – 12 ]. Notably, adoptive immunotherapy using CD19-specific CAR-T cells has become an approved intervention in onco-hematology, with indications for relapsed and/or refractory B-cell malignancies [ 13 , 14 ]. The success of CAR-T cell therapy in B-cell malignancies has inspired its clinical translation to autoimmune diseases [ 15 , 16 ]. Recent studies indicate that CAR T-cell therapy induces profound biological alterations in autoimmune disease, including suppression of key immunological pathways—such as complement activation and type I interferon responses—accompanied by metabolic reprogramming [ 17 ].

This review synthesizes clinical progress in CAR-T cell therapy across diverse autoimmune diseases and comprehensively examines emerging strategies to enhance the therapeutic efficacy of CAR-T cells in autoimmunity. These strategies include the selection of specific T cell subsets, structural modifications, improved infiltration into the immunosuppressive microenvironment, and the application of genetic and metabolic interventions. The review also highlights advances in CAR-T cell manufacturing and the adoption of novel technologies and platforms to reduce time and cost. Furthermore, we explore the challenges encountered in translating CAR-T cell therapy to autoimmune diseases by analyzing relevant preclinical studies and clinical trials. Ultimately, this review aims to propose potential solutions to overcome these challenges and to enhance the efficacy and applicability of CAR-T cell therapies for the treatment of autoimmune diseases.

Autoimmune disease pathogenesis and the role of immune cells

Autoimmune diseases comprise a clinically heterogeneous group of disorders characterized by a breakdown of immunological tolerance, leading to dysregulated activation of both adaptive and innate immune responses that drive tissue-specific or systemic pathology [ 18 , 19 ]. Multiple immunological aberrations underlie the loss of self-tolerance and perpetuate autoreactive responses in autoimmune diseases. In particular, dysregulated innate immune responses initiate and amplify adaptive immune activation, fueling the inflammatory cascade [ 19 , 20 ]. Studies demonstrate that T cells, B cells, neutrophils, natural killer (NK) cells, and macrophages contribute differentially to autoimmune pathogenesis through dynamic intercellular crosstalk and cytokine-mediated immune modulation [ 18 , 19 ] (Figure. 1 ). A comprehensive understanding of the roles of distinct immune cell populations in autoimmune diseases pathogenesis may reveal novel cellular targets and therapeutic opportunities for CAR-T cell therapy in autoimmunity.

Abnormalities of B cells in autoimmune diseases

Under physiological conditions, immature B cells differentiate into naïve B cells (NBCs), which, upon antigen encounter, migrate into nascent germinal centers (GCs) for affinity maturation in the spleen. Supported by T follicular helper (Tfh) cells, high-affinity GC B cells subsequently differentiate into memory B cells or antibody-secreting plasma cells [ 21 ]. Notably, a subset of immature B cells can undergo extrafollicular activation—for instance, within T-cell zones or medullary cords—rapidly differentiating into short-lived plasma cells [ 22 ]. Under interferon-γ (IFNγ) exposure, B cells upregulate T-bet, with a fraction of these T-bet + cells persisting as splenic memory B cells [ 23 – 25 ].

Autoimmune pathogenesis involves the early emergence of autoreactive B-cell clones and the production of self-antigen-targeting autoantibodies, which may arise years before clinical onset. B cells drive disease progression through antigen presentation, cytokine secretion, and differentiation into antibody-producing plasmablasts and plasma cells [ 26 – 28 ]. The prevailing paradigm implicates autoantibody production as the central mechanism of B cell-driven autoimmune pathology [ 6 ]. Clinical diagnosis often relies on disease-specific autoantibody signatures, which reflect the activation of autoreactive B-cell populations [ 29 ]. The presence of Fab-glycosylated IgG antibodies is associated with several B-cell-mediated autoimmune diseases, including RA, SLE, anti-glomerular basement membrane glomerulonephritis, and Guillain-Barré syndrome [ 30 ]. Furthermore, immune complexes containing autoantibodies can mediate tissue damage via complement activation (CDC), antibody-dependent cellular cytotoxicity (ADCC), and synergistic FcR/TLR signaling, as demonstrated in RA and SLE [ 31 , 32 ].

B cell activation requires two critical signals: (1) antigen engagement by the B cell receptor (BCR) and (2) co-stimulation, typically provided by Tfh cells or pattern recognition receptors (PRRs). Immune checkpoints—comprising both activating and inhibitory receptors—fine-tune B cell responses, ensuring balanced immunity while preventing autoimmunity [ 27 ]. In SLE, systematic analysis of BCR repertoires reveals significant biases in VDJ gene usage, particularly in CD27 + IgD + unswitched memory B cells and plasmablasts. Autoreactive B cell clones are enriched within this unswitched memory compartment and correlate strongly with disease activity, suggesting direct involvement in SLE pathogenesis [ 33 ]. Similar BCR repertoire abnormalities have been observed in RA and other autoimmune diseases, implicating dysregulated B cell selection as a shared feature of autoimmunity [ 34 ].

B cell activation and effector functions are regulated by immune checkpoints that modulate B cell tolerance, activation, antigen presentation, T cell help, class switching, antibody production, and cytokine secretion [ 27 ]. B cells activate CD4 + T cells through MHC class II-mediated antigen presentation, while reciprocally receiving help from Tfh cells—particularly during dendritic cell (DC) priming within immune responses [ 35 ]. Profiling studies of RA synovial tissue and fluid have identified a prominent population of T peripheral helper (Tph) cells alongside Tfh cells, both facilitating B cell assistance via IL-21 secretion and CD40 ligand (CD40L) expression [ 36 ]. Receptor-ligand analyses have implicated IFNG and TNFRSF members as key mediators of synovial Tph–B cell interactions [ 37 ]. In SLE, the frequency of Tph cells correlates with clinical disease activity and the expansion of CD11c + B cells, while CRISPR-Cas9-mediated MAF knockout or IL-21 neutralization abrogates the capacity of Tph cells to drive memory B cell differentiation into plasmablasts [ 38 ]. Targeting SLE-associated SLAMF1 overexpression with blocking antibodies disrupts pathogenic T–B cell interactions, reducing immunoglobulin production and B cell activation [ 39 ].

B cells can also undergo extrafollicular maturation within inflamed tissues, generating autoantibody-secreting plasmablasts and short-lived plasma cells. This process, often driven by TLR activation independently of T cell help, is a key mechanism of T cell-independent B cell maturation [ 40 ]. In SLE, elevated circulating B-cell-activating factor (BAFF) levels positively correlate with anti-dsDNA antibody titers and increased risk of disease progression [ 41 ]. In addition to autoantibody production and antigen presentation, B cells differentiate into cytokine-producing effector subsets, secreting pro-inflammatory mediators (IFN-γ, IL-6, IL-2) to amplify immune responses or immunoregulatory cytokines (IL-10, IL-4) to enforce tolerance [ 42 , 43 ]. Understanding these mechanisms may facilitate the development of B cell-modulating therapies for autoimmune rheumatic diseases.

T cell dysregulation in autoimmune diseases

Autoimmune pathogenesis involves not only B cell-mediated mechanisms but also critically depends on T cell participation as a key immunological determinant. In ADs, CD4 + T cell homeostasis is disrupted, with pathological skewing toward pro-inflammatory effector lineages—including Th1, Th17, and Tfh cells—at the expense of regulatory subsets [ 44 ]. Tfh cells play a pivotal role in driving autoimmunity by providing cognate B cell help, promoting germinal center formation and pathogenic autoantibody production [ 45 , 46 ]. Recent evidence demonstrates that NLRP3 inflammasome activation in Tfh cells drives SLE pathogenesis by promoting germinal center responses and high-affinity autoantibody production [ 47 ]. Basophil-derived PD-L1 and IL-4 are required for the accumulation and function of pathogenic Tfh cells in secondary lymphoid organs, facilitating Tfh2 cell development and effector function in lupus [ 48 ].

Regulatory T cells (Tregs) are master regulators of immune homeostasis, enforcing peripheral tolerance via multifaceted suppressive mechanisms. Both quantitative and functional impairments in the Treg compartment represent fundamental breaches in self-tolerance that drive autoimmune pathogenesis [ 49 ]. In active SLE, antigen-presenting cells (APCs) induce Treg dysfunction via OX40L signaling, with OX40L + cells co-localizing with Foxp3 + Tregs in cutaneous lesions [ 50 ]. Single-cell transcriptomic and chromatin accessibility analyses have revealed a CCR7 low CD74 hi Treg subset exhibiting type I interferon-induced functional exhaustion in SLE patients [ 51 ]. In RA, patients in stable remission exhibit higher frequencies of circulating follicular regulatory T (Tfr) cells, which correlate with lower autoantibody levels [ 52 ]. We summarize these diseases and the associated T cell abnormality in the supplemental Table 1 .

Dendritic cells in autoimmune diseases

Dendritic cells (DCs), the most potent professional antigen-presenting cells, exhibit a dual role in immunity by maintaining immune tolerance while also initiating pathogenic autoimmune responses. Their heterogeneous populations include classical DCs (cDCs), plasmacytoid DCs (pDCs), monocyte-derived DCs (MoDCs), and tolerogenic DCs (tolDCs) [ 53 ]. In RA, thymus and activation-regulated chemokine (TARC), elevated in synovial fluid, promotes T-cell recruitment and drives pro-inflammatory cytokine secretion, a process critically dependent on CD1c-expressing cDCs [ 54 ]. pDCs, a distinct innate immune cell population, uniquely recognize microbial nucleic acids via endosomal Toll-like receptor (TLR) signaling, leading to type I interferon secretion and modulation of innate and adaptive immunity [ 55 ]. In SLE, extrafollicular generation of short-lived antibody-forming cells (AFCs) constitutes a major mechanism of anti-DNA autoantibody production, dependent on pDC-derived IFN-I and TLR7/9 activation, bypassing conventional germinal center pathways [ 56 ]. These findings provide mechanistic insights and potential therapeutic targets in autoimmune diseases pathogenesis.

Macrophages and neutrophils in autoimmune diseases

Macrophages are pivotal components of the innate immune system, playing essential roles in tissue homeostasis, host defense, tissue repair, metabolic regulation, and inflammation [ 57 , 58 ]. In lupus nephritis, monocyte-derived macrophages (MoMacs) exhibit robust upregulation of Fcγ receptor (FcγR) response genes, enhancing immune complex (IC) internalization, while tissue-resident macrophages (TrMacs) facilitate the formation of autoantibody-producing lymphoid aggregates via sustained secretion of B-cell niche factors [ 59 ]. Macrophage plasticity in SLE represents an attractive therapeutic target for rebalancing dysregulated immune responses through phenotype modulation [ 60 ]. Neutrophil activation and neutrophil extracellular trap (NET) release are key pathogenic drivers of systemic autoimmunity, promoting disease initiation, persistence, and end-organ damage [ 61 – 63 ]. The REDD1/autophagy axis promotes thromboinflammation and fibrosis in SLE by generating tissue factor (TF)- and interleukin-17 A (IL-17 A)-bearing NETs, which contribute to both disease onset and chronicity [ 64 ]. Distinct neutrophil subsets with mitochondrial dysfunction and excessive mitochondrial reactive oxygen species (mROS) generation have been identified as key drivers of NETosis-mediated renal injury in lupus [ 65 ]. Collectively, these findings establish macrophages and neutrophils as central mediators of autoimmune diseases progression and promising therapeutic targets.

NK and NKT cells

Although autoimmune diseases are primarily driven by dysregulated T and B lymphocyte responses, emerging evidence suggests that NK cells also contribute to the pathogenesis and modulation of peripheral tolerance [ 66 ]. Dysregulation of CD56 bright CD16 − NK cells and CD56 dim CD16 − NK cell homeostasis may influence SLE progression and serve as novel biomarkers for disease severity and renal complications [ 67 ]. Single-cell RNA sequencing studies in patients with autoimmune diseases have consistently identified NK cells as active participants in disease mechanisms [ 68 , 69 ], while preclinical data suggest that restoring NK cell homeostasis could represent a viable therapeutic strategy [ 70 ]. Immunohistochemical analysis reveals the presence of CD56 + /NKp46 + CD3 − NK cells and CD56 + /NKp46 + CD3 + NK-like T cells in lupus nephritis kidneys, predominantly accumulating in interstitial and peri-glomerular regions, suggesting a role in renal inflammation in SLE [ 71 ]. Invariant natural killer T (iNKT) cells, a subset of innate-like T lymphocytes that recognize lipid antigens [ 72 ], have been shown to promote pathogenic B-cell responses in SLE via CD1d and CD40L–CD40 signaling, highlighting these pathways as potential therapeutic targets [ 73 ]. While these findings imply NK and NKT cells in autoimmune pathogenesis, further mechanistic and preclinical investigation is warranted to clarify their roles and translational potential.

Current treatment and limitations of autoimmune diseases

Autoimmune diseases primarily arise from maladaptive crosstalk between adaptive (T/B cells) and innate immune cells (neutrophils, NK cells, macrophages), where dysregulated cell-cell signaling and cytokine networks drive loss of self-tolerance and chronic tissue inflammation. Current therapeutic strategies mainly focus on modulating T cell and B cell functions as key immunoregulatory targets. Conventional treatments include corticosteroids, non-steroidal anti-inflammatory drugs (NSAIDs), hydroxychloroquine, and immunosuppressive agents such as calcineurin inhibitors and azathioprine. Given the central role of T cells in mediating autoimmune responses, antibody-based immunotherapies are actively under investigation [ 74 ]. By targeting ligand-receptor interactions, these therapeutic antibodies direct cell-mediated cytotoxicity toward pathogenic cells, as exemplified by anti-CD3 monoclonal antibody (mAb) therapy.

In addition to mAbs, bispecific T cell engagers (BiTEs) comprise an antibody fragment targeting the TCR-CD3 complex and a disease-specific antigen-binding domain, bridging T cells and target cells to induce an artificial immune synapse [ 75 ]. This mechanism enables T cell-mediated cytotoxicity independent of endogenous TCR specificity. Moreover, targeted therapies against integrin receptors—including the α4 integrin antagonist natalizumab and the α4β7 integrin inhibitor vedolizumab—as well as sphingosine-1-phosphate (S1P) receptor modulators, have proven effective in treating inflammatory bowel disease (IBD) [ 76 ].

Emerging evidence also supports the use of therapeutic antibodies targeting co-inhibitory molecules, such as CTLA-4 and PD-1/PD-L1, as promising strategies for autoimmune diseases management [ 77 ]. Beyond direct antibody-based therapies, cytokine-targeted approaches such as small-molecule JAK inhibitors have demonstrated clinical efficacy in various autoimmune and inflammatory disorders [ 78 ]. Furthermore, owing to its capacity to selectively expand immunomodulatory Tregs, low-dose interleukin-2 (IL-2) therapy has emerged as a promising approach for autoimmune diseases [ 79 , 80 ].

Selective targeting of B cell activation and autoantibody production also represents a key therapeutic strategy. Rituximab, a monoclonal antibody directed against the CD20 antigen, has been extensively tested in numerous autoimmune diseases, including SLE, MS, RA, and lupus nephritis [ 29 ]. Additionally, anti-CD19 and anti-BAFF monoclonal antibodies have demonstrated therapeutic potential in clinical trials for various autoimmune indications [ 81 , 82 ]. However, the EXPLORER randomized controlled trial (Phase II/III) found that rituximab did not yield significant clinical improvement over placebo in moderate-to-severe extrarenal SLE, as measured by all predefined efficacy endpoints [ 83 ]. Moreover, relapses after rituximab are common, necessitating retreatment to maintain disease control—a pattern also observed with other therapeutic monoclonal antibodies.

B cell depletion reduces autoantibody levels by only approximately 30–70%, suggesting that other B cell functions are also critical in the pathogenesis of autoimmunity. These treatments do not fully eliminate B cells in secondary lymphoid organs or tissues, and antibody concentrations eventually decline, which may limit the clearance of pathogenic cells [ 84 ]. Consistent with this, the impact of B cell depletion on pathogenic, tissue-localized T-bet + CD11c + age-associated B cells (ABCs) remains poorly defined [ 85 ]. Effective control of autoimmune diseases, and potentially even an “immunological reset,” may require deep and sustained B cell depletion [ 23 , 86 ].

Another therapeutic challenge stems from the phenotypic plasticity of B cell surface markers—particularly since CD20 expression is absent on terminally differentiated B-lineage cells, including plasmablasts, plasma cells, and ABCs [ 87 ]. Furthermore, while conventional anti-inflammatory agents can effectively suppress the inflammatory response, they often require continuous administration over extended periods—potentially for years or even a lifetime. Such prolonged therapeutic regimens are associated with an increased risk of secondary infections and malignancies, significantly compromising patients’ quality of life. Additionally, existing B-cell depletion therapies, including monoclonal antibodies targeting CD19 or CD20, demonstrate limited efficacy against plasma cells residing within the central nervous system [ 88 ]. In light of these limitations, current investigations are focusing on developing immune-reprogramming platforms capable of resetting autoreactive immunity, potentially enabling sustained disease remission and complete withdrawal of maintenance immunosuppression.

CAR-T cell therapy as an emerging approach

CAR-T cell therapy, initially pioneered for hematologic malignancies, is rapidly evolving as a transformative strategy in autoimmune diseases management. By leveraging the precision of synthetic immunology, CAR-T cells are engineered to recognize specific antigens, reprogram immune responses, and restore immune homeostasis—representing a paradigm shift from broad immunosuppression to targeted immune modulation. The application of CAR-T therapy to autoimmune diseases began in 2021 with the first reported SLE case, and has since expanded rapidly across rheumatology (20 + cases), neurology (15 + cases), and NMOSD (12 patients) by 2025 [ 89 – 98 ]. This therapy involves genetically engineering a patient’s T cells to express synthetic receptors that enable targeted recognition and destruction of pathogenic immune cells [ 99 ].

The basic structure and function of CAR-T cells

CAR-T cell therapy relies on genetically engineered receptors that enable T cells to recognize and eliminate target cells in an MHC-independent manner. Typically, a CAR comprises four key components [ 100 – 102 ]: the antigen-binding domain, hinge (spacer) domain, transmembrane domain, and intracellular co-stimulatory and signaling domain. Each component can be fine-tuned to optimize specificity, sensitivity, and activation efficiency. Figure 2 presents the functions, major segments, and current engineering strategies for each structure.

Antigen-binding domain (ABD)

The ABD is the molecular backbone of CAR-T cell specificity, directing antigen recognition through MHC-independent mechanisms. It can incorporate a range of binding motifs, including monoclonal antibody-derived single-chain variable fragments (scFvs [ 103 ]), single-domain antibodies (such as nanobodies [ 104 , 105 ] and variable new antigen receptors [ 106 ]), natural ligand-receptor pairs (APRIL [ 107 ], BAFF [ 108 , 109 ]), and engineered small-molecule binders [ 110 , 111 ]. A key challenge is balancing sensitivity and specificity: high-affinity ABDs increase detection but risk off-target toxicity and tonic signaling, while low-affinity variants improve specificity but may reduce potency. Recent refinements include dual/multi-specific ABDs to mitigate antigen escape [ 112 – 114 ], linker engineering to prevent scFv aggregation [ 115 ], and computational affinity maturation to refine binding kinetics [ 116 ].

Hinge (Spacer) domain

Traditionally derived from IgG1 hinge regions, the hinge domain modulates antigen engagement and signaling by adjusting the spatial separation between the scFv and transmembrane domain. Optimal positioning of epitopes—whether membrane-proximal or membrane-distal—requires careful consideration of ligand density, hinge rigidity, and steric accessibility [ 101 ]. Non-IgG-based spacers, such as those from CD8 and CD28, have emerged to reduce Fcγ receptor-mediated off-target effects while maintaining robust efficacy [ 117 ].

Transmembrane domain

This domain anchors the CAR and helps stabilize signal transmission, often using motifs from membrane proteins like CD28 or CD3ζ. Proper selection is essential for synergistic interactions with extracellular and intracellular modules. A notable innovation is the split-CAR architecture (Fig. 3 A), partitioning CARs into two non-functional polypeptides anchored within the transmembrane domain [ 118 – 120 ]. Upon administration of soluble switch molecules, these chains are brought into proximity, enabling AND-gate antigen recognition, titratable activity, and on-demand reprogramming without re-engineering the T cell [ 121 ].

Intracellular domain .

The intracellular portion consists of co-stimulatory elements (e.g., CD28, 4-1BB) that regulate T-cell survival, and the TCR-derived CD3ζ signaling unit that initiates activation. Third-generation CARs incorporate multiple co-stimulatory domains to enhance persistence. These domains exert differential effects on basal signaling—CD28 amplifies, while 4-1BB attenuates spontaneous CD3ζ activation, which may otherwise cause tonic signaling and T-cell exhaustion [ 122 , 123 ]. Recent studies focus on genetic modification of motifs (e.g., 4-1BB/CD28 hybrids) and the development of novel variants (e.g., ICOSζ, OX40ζ, CD27ζ) 122 . The sequence and combination of co-stimulatory domains influence effector functions [ 124 , 125 ], making their selection context-dependent.

The CD3ζ chain contains a basic amino acid-rich sequence (BRS) sequestering three ITAMs in its resting conformation. Antigen engagement induces conformational changes, exposing ITAMs for phosphorylation by Src-family kinases (e.g., Lck) and recruitment of downstream effectors. Both ITAM multiplicity and proximity to the transmembrane domain modulate signal intensity and CAR-T persistence [ 126 – 128 ]. While increased ITAM density or reduced distance enhances activation, excessive signaling can trigger activation-induced cell death (AICD), compromising efficacy. For example, adjusting the number of functional ITAMs in CD3ζ can extend CAR-T persistence and potency in preclinical models [ 128 ].

Refinements in CAR design focus on amplifying activation signals, prolonging in vivo persistence, improving tissue infiltration, and enabling precise antigen discrimination to spare healthy tissues [ 129 ]. Restricted antigen recognition domains (such as those targeting tumor-specific neoantigens) help reduce off-target effects [ 130 , 131 ]. Functional modules can be “armored” to further optimize activation, persistence, and specificity, leading to the development of third- to fifth-generation CARs.

Iterative structural optimization of CAR-T cells

The evolution of CAR-T technology reflects iterative optimization to enhance efficacy and safety: First-generation CARs, developed by Zelig Eshhar in 1993, combined an scFv with the CD3ζ signaling domain [ 132 ]. While they enabled antigen recognition and cytotoxicity, their reliance solely on CD3ζ signaling led to limited T-cell proliferation, transient persistence, and insufficient cytokine secretion, ultimately compromising potency [ 133 ]. Second-generation CARs incorporated a single co-stimulatory domain (such as CD28, 4-1BB, or OX-40) alongside CD3ζ [ 134 ]. These domains synergistically amplify activation: CD28 promotes immediate cytokine release and rapid expansion, while 4-1BB supports mitochondrial biogenesis and long-term persistence [ 135 – 137 ]. Clinical studies have demonstrated that CD28-based CAR-T cells yield robust initial responses, while 4-1BB-based cells provide sustained survival, as reflected in their different cytokine profiles and pharmacokinetics [ 135 , 138 ]. Third-generation CARs are defined by the incorporation of two distinct co-stimulatory domains (e.g., CD28 and 4-1BB) in tandem with CD3ζ, further improving activity in preclinical models, though concerns about T-cell exhaustion due to overstimulation emerged [ 139 – 141 ]. Fourth-generation CARs (also known as TRUCKs: T cells Redirected for Universal Cytokine Killing) are characterized by the addition of inducible transgenes—typically cytokines (e.g., IL-12, IL-15) or chemokines—that are expressed upon CAR activation) [ 142 ]. These “armored” CAR-T cells enhance the microenvironment by recruiting endogenous immune cells and countering immunosuppressive signals. Early trials (such as BCMA-targeted CAR-T cells secreting IL-7 and CCL19) have shown prolonged remission in multiple myeloma with manageable toxicity [ 121 , 143 ]. A representative fourth-generation CAR example involves inclusion of pre-activation domains, like CD137, priming T cells into a metabolically active state before antigen engagement, thereby enhancing cytotoxicity [ 144 – 146 ]. Notably, the defining feature of fourth-generation CARs is antigen-dependent transgene expression, not pre-activation domains. Fifth-generation CARs integrate intracellular domains from cytokine receptors (e.g., IL-2Rβ) that activate JAK/STAT signaling in addition to CD3ζ and co-stimulatory domains, or employ advanced synthetic biology modules such as logic gates or inducible safety switches [ 147 ]. These advances aim to improve safety (e.g., mitigating on-target/off-tumor toxicity) and adaptability for complex diseases, including autoimmune disorders. Logic-gated transcription circuits (e.g., SynNotch CAR cascades) and modular CAR systems enable post-infusion dose titration or emergency shutdown in cases of severe toxicity [ 148 , 149 ](Fig. 3 A).

These iterative CAR innovations underscore the interplay between synthetic biology and clinical needs, providing a blueprint for tailoring therapies to diverse contexts [ 150 , 151 ]. Among these features, the extracellular antigen-binding domain determines target specificity and is essential for extending CAR-T application to autoimmunity. Conventional CARs for hematologic malignancies typically employ an scFv targeting CD19-positive B cells [ 152 – 154 ] or both CD19-positive B cells and BCMA-positive plasma cells [ 114 , 155 , 156 ]. Accordingly, CD19 and BCMA are the most extensively studied antigens in autoimmune diseases contexts [ 157 , 158 ]. Beyond these, novel structural optimizations have been developed to further specify targeting and reduce treatment-related adverse effects in ADs [ 159 , 160 ].

The mechanism of action of CAR-T cells: targeted recognition and immune regulation

Targeted recognition and CAR-T cell activation

Upon recognizing their target antigen, CAR-T cells form a non-classical immune synapse with the target cell. Unlike the conventional TCR-mediated synapse, which depends on peptide–MHC interactions, this synapse is established through direct antigen binding, initiating cytotoxic activity [ 161 ]. The three ITAMs within CD3ζ undergo conformational changes, exposing tyrosine residues for phosphorylation by Src-family kinases (e.g., Lck). Phosphorylated ITAMs act as docking sites for ZAP-70, which then phosphorylates adaptor proteins LAT, SLP-76, and Link CAR triggering downstream signaling cascades including the MAPK, NF-κB, and Ca²⁺-NFAT pathways [ 162 , 163 ]. Ultimately, CAR-T cells mediate target cell elimination primarily through direct cytolytic activity, while also secreting cytokines that amplify and sustain the immune response.

Direct cellular killing

CAR-T-cell-mediated cytotoxicity operates via two principal pathways: (1) Perforin and Granzyme Pathway: Upon activation, CAR-T cells release cytotoxic granules containing perforin and granzymes. Perforin forms transmembrane pores in the target cell, allowing granzymes to enter the cytoplasm and cleave intracellular substrates, leading to cytolysis [ 164 ]. This is considered the primary mechanism of redirected cytotoxicity by CAR-T cells. (2) Fas/FasL Pathway: FasL expressed on CAR-T cells binds to Fas receptors on target cells, triggering the caspase cascade and programmed cell death [ 164 ]. Though slower than the perforin/granzyme pathway, it is effective when granule-mediated killing is compromised [ 165 , 166 ].

Generally, CAR-T-cell cytotoxicity in hematologic cell lines is more sensitive to granzyme inhibition, whereas killing in solid tumor cells relies more on Fas-mediated apoptosis [ 165 ]. This functional redundancy enables both rapid and sustained cytolytic responses against diverse targets.

Cytokine release and immune modulation

Activated CAR-T cells exert potent immunomodulatory effects by releasing a range of cytokines, such as IL-2, IL-6, and IFN-γ, that reprogram the immune microenvironment [ 167 ]. These effects can be classified into four main mechanisms: (1) Amplified Cytotoxicity and Immune Cell Recruitment: Pro-inflammatory cytokines enhance CAR-T-cell cytotoxicity and attract additional immune cells, including NK cells and endogenous T cells [ 167 , 168 ]. Notably, an NFAT-responsive cassette that enhances cytokine expression (e.g., IL-12) has been incorporated into fourth-gen CARs to promote recruitment and activation of NK cells and other effectors [ 142 ]. (2) Broad Host Immune Activation: CAR-T cells can initiate systemic immune activation, recruiting antigen-presenting cells such as dendritic cells and macrophages. They also promote polarization of tumor-associated macrophages toward the pro-inflammatory M1 phenotype, enhancing antitumor activity [ 169 , 170 ]. Critically, in autoimmune contexts, this inflammatory remodeling—combined with deep depletion of autoreactive B-cell clones—may disrupt self-sustaining autoimmune circuits and create a permissive window for immune reconstitution from hematopoietic precursors, a process central to the “immune reset” phenomenon. (see Box 1). (3) Bystander Effect: Activation of non-engineered endogenous T cells (bystander cells) is driven by the inflammatory milieu created by CAR-T cells [ 166 , 171 , 172 ]. Lysis of antigen-expressing cells releases antigens, which can be processed and presented by APCs to endogenous T cells, extending the immune response to targets not initially recognized by the CAR [ 172 ]. (4) Modulation of Tregs: The pro-inflammatory tumor microenvironment can suppress Treg infiltration and activity, thereby enhancing CAR-T-cell persistence and efficacy. For example, CAR-T-cell secretion of IL-12 decreases Treg levels [ 173 , 174 ], and reducing Treg abundance can further promote immune responses [ 175 , 176 ].

Clonal expansion and persistence

After cytolysis, CAR-T cells undergo clonal expansion, maintaining their cytotoxic potential [ 177 ]. In some instances, they persist in circulation for months [ 178 ], contributing to durable remissions, particularly in hematologic malignancies.

Box 1. Immunological reset following CAR T-cell infusion

“Immune reset,” as opposed to mere immunomodulation, involves the elimination of autoreactive lymphocytes followed by the regeneration of a balanced, self-tolerant immune repertoire in ADs. Unlike traditional depletion strategies such as rituximab, CAR T-cell therapy can achieve comprehensive clearance of effector and memory immune cells—even within central and peripheral lymphoid tissues—thus providing a deeper reset of immune architecture. In addition, the preparative lymphodepletion regimen used prior to CAR infusion may further facilitate immune reconstruction by enabling the regeneration of naïve, non-pathogenic immune cells [ 179 , 180 ].

This immunological reset can be characterized by a reduction in the levels of effector and memory B cells [ 181 ], an increase in naïve and non-pathogenic immune cells [ 182 , 183 ], diversification of B-cell and T-cell repertoires [ 183 , 184 ], and alterations in lymphocyte subsets—including elevated regulatory T cells [ 185 ], faster recovery of CD8 + T cells relative to CD4 + T cells [ 183 ], increased peripheral CD8 + CD28 − CD57 + T cells [ 183 ], and an rise in IL-10-producing B cells—and is reflected clinically by sustained drug-free clinical remission and clearance of serum autoantibodies.

In non-human primate models treated with CAR T-cells, researchers observed an immune reset, evidenced by the re-emergence of naïve B cells, occurring on average 21 days post-infusion [ 183 , 186 ]. In clinical observations of ADs, flow cytometric analyses of peripheral blood post-infusion indicate that pathogenic autoreactive B cells are completely eliminated before CAR-T cells contract to low levels and B-cell reconstitution begins spontaneously [ 89 , 90 , 94 ]. This limited persistence of CAR-T cells in ADs contrasts with findings in B-cell malignancies, where CAR-T cells can persist for years and B-cell aplasia often lasts much longer [ 90 , 94 , 178 ]. Interestingly, even though B-cell reconstitution occurred relatively early (with a reported average of 110 days in one follow-up cohort), no patients experienced clinical relapse within up to 12 months of observation [ 90 , 94 ]. These findings suggest that immune reset is not merely B-cell depletion, but involves the ablation of autoreactive memory compartments and the regeneration of a naïve, diverse B-cell pool from bone marrow precursors—potentially facilitated by lymphodepletion-induced niche opening. Further analyses of immunoglobulin light and heavy chain usage in reconstituted B cells show patterns closely resembling those of healthy individuals [ 90 , 94 ]. Phenotypic profiling also supports this, with characteristics similar to re-emerging B cells observed after myeloablative conditioning and autologous HSCT [ 187 , 188 ]. This durable disease remission, along with the restore of a healthy immune system, is central to the therapeutic promise of CAR-T in autoimmunity. Besides, it is noteworthy that in some ADs, such as SLE, autoantibody accumulation can precede clinical manifestations by several years [ 189 ]. Therefore, the absence of disease recurrence during early follow-up may reflect insufficient time for a pathogenic antibody buildup to reach a clinically relevant threshold.

CAR T-cell-based therapy: from cancer to autoimmunity

The on-target off-tumor rebalance in autoimmune diseases

In hematologic malignancies, CAR-T cells are engineered for high-affinity on-target lethality to eradicate malignant clones effectively. However, ADs arise from dysregulated immune homeostasis rather than uncontrolled proliferation, necessitating a paradigm shift in CAR-T design to reconcile on-target specificity (targeting pathogenic immune cells) with off-tumor safety (preserving physiological immune tolerance). This requires multidimensional optimization, including antigen selection refinement, tissue-specific activation control, and tolerogenic immune modulation, to mitigate autoimmune pathology while avoiding disruption of healthy immunity.

The extracellular scFv domain enables CAR-T cells to recognize disease-associated markers, such as CD19 on autoreactive B cells in SLE or BCMA on plasma cells in RA [ 190 ]. Affinity tuning of scFvs ensures selective targeting of cells with high antigen density, minimizing off-tumor toxicity to healthy tissues expressing low levels of the same antigen [ 103 ]. Novel antigen-binding domains, such as structurally stabilized “binders,” further enhance CAR-T persistence and functionality by reducing immunogenicity and improving pharmacokinetics [ 110 ].

Beyond direct cytotoxicity, CAR-T cells modulate the immune microenvironment via cytokine secretion and synaptic signaling. TRUCKs integrate inducible cytokine modules (e.g., IL-10 or TGF-β) to locally promote Treg expansion or suppress pro-inflammatory Th17 responses, thereby fostering immune tolerance [ 191 ]. CAR-Tregs, engineered to recognize tissue-specific antigens (e.g., insulin for type 1 diabetes), migrate to inflamed sites and exert suppressive effects through contact-dependent mechanisms (such as CTLA-4 engagement) or paracrine signaling (such as IL-35 secretion), offering a targeted approach to re-establish immune equilibrium [ 192 , 193 ].

Distinct immune dynamics following CAR T cell therapy in hematologic malignancies versus autoimmune diseases

Given the hyperactive immune state in ADs, immune responses following CAR T-cell therapy may differ from those observed in hematologic malignancies. First, long-term immune recovery after CAR T-cell therapy in hematologic cancers typically restores a functional immune system capable of combating infections and maintaining immune homeostasis [ 194 , 195 ]. In contrast, immune reset in ADs leads to the establishment of immune tolerance, effectively preventing autoreactive responses without compromising overall immune surveillance (Box 1). Clinical trials in SLE and myasthenia gravis have observed sustained remission post-CD19 CAR-T therapy, correlating with B-cell regeneration and reduced autoantibody titers [ 196 ]. The underlying alterations in cellular components—spanning from a reduction in autoreactive immune cells, diversification of T-cell and B-cell repertoires, to variations in T-cell and B-cell subsets [ 183 ]—also signify the ablation of an overactivated immune system and the subsequent reconstruction of a homeostatic one(Box1). This process may contribute to the prolonged therapeutic effects observed in ADs. Second, the incidence and severity of cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) appear to be generally lower in ADs. For instance, CRS has been reported in 42% to 95% of patients receiving CD19-targeted or BCMA-targeted CAR T-cell therapy for hematologic malignancies [ 197 , 198 ]. A recent systematic review further confirmed that most patients treated with CAR T-cell therapy for ADs experienced only low-grade CRS, partly owing to a lower antigen burden and relatively reduced immune activation [ 199 ].

Challenges in clinical application

The ultimate goal of CAR T-cell therapy in autoimmune diseases is to achieve long-term, drug-free remission. Encouraging results have emerged from previous studies. For example, in a cohort of patients with refractory SLE, all individuals achieved sustained drug-free remission and resolution of nephritis following CAR T-cell injection [ 90 ]. Moreover, significant improvements in quality of life have been reported in patients with antisynthetase syndrome, as assessed by Physician Global Assessment (PGA), functional muscle testing, pulmonary evaluations, and other clinical measures [ 89 , 91 , 92 ]. Since the first report in 2021 of a patient with refractory SLE successfully treated with anti-CD19 CAR T cells [ 1 ], this therapeutic approach has shown immense promise—but with that have come several areas of concern.

Safety concerns (e.g., CRS, long-term immunosuppression)

The most common adverse effect associated with CAR T-cell therapy is CRS, which can be life-threatening in severe cases. CRS is believed to be triggered by the excessive release of pro-inflammatory cytokines, such as IL-6, and is further amplified by intense immune activation [ 200 – 202 ]. Another frequent acute reaction is ICANS, typically occurring within the first 10 days post-infusion [ 200 , 203 ], with incidence rates ranging from 15% to 64% among the six FDA-approved CAR T-cell products [ 83 ]. As previously noted, both CRS and ICANS tend to present more mildly in autoimmune diseases compared to malignancies [ 90 , 197 , 198 , 200 , 203 , 204 ]. This difference can be explained by the absence of malignant tumor burden and relatively lower levels of immune activation in patients with autoimmune disorders [ 204 ].

Once activated, CD19-targeted CAR T cells eliminate B cells expressing the target antigen, resulting in a temporary state of complete B-cell depletion, known as B-cell aplasia. Although long-term B-cell aplasia is rare, patients may experience hypogammaglobulinemia and an increased risk of infection during this period [ 196 ]. For instance, in a follow-up study of 15 patients with ADs who received CAR T-cell therapy, 2 individuals developed secondary hypogammaglobulinemia during the observation period spanning from the 3rd to the 12th month (the study endpoint) [ 94 ]. However, in contrast to cancer settings, in which the median duration of B-cell aplasia exceeds four months [ 200 ], B-cell recovery tends to occur more rapidly in autoimmune diseases. For example, follow-up data in rheumatology indicate that the median duration of B-cell aplasia post-treatment is approximately 112 days [ 90 , 205 ]. In addition, vaccine-induced antibody titres remain stable at pre-treatment levels, and total IgG concentrations do not decline substantially in patients with ADs receiving anti-CD19 CAR T-cell therapy [ 94 , 191 ]. Collectively, these findings suggest that the shorter duration of B-cell aplasia, preservation of antibody levels, and milder cytotoxic exposure may contribute to a lower risk of infection in ADs patients. Although treatment-related adverse effects tend to be milder in autoimmune diseases compared to cancer, both acute and long-term side effects should be carefully monitored and managed.

Durability and relapse risk

Following infusion, CAR T cells undergo rapid expansion, reaching a peak level before persisting at much lower concentrations for months or even years after treatment [ 176 , 206 ]. Concurrently, B cells—the primary target in most cases—undergo complete depletion in response to the swift proliferation of CAR T cells. Circulating B cells typically begin to recover once CAR T-cell levels decline. The duration of this process varies among individuals. As noted above, in contrast to patients with hematologic malignancies, CAR T cells in ADs generally exhibit shorter persistence and lower peak expansion [ 90 , 94 ], potentially due to the lack of malignant masses and reduced cytotoxic stimulation.

In the context of malignancies, early CAR T-cell exhaustion followed by reconstitution of target cells often leads to disease relapse [ 207 ]. However, emerging evidence in ADs, particularly SLE, suggests a distinct trajectory. Remarkably, patients can sustain durable remission and symptom resolution even after B-cell reconstitution. Long-term follow-up studies support this observation [ 90 , 94 , 96 ]. In a cohort of patients with refractory SLE treated with CD19-targeted CAR T cells, no signs of disease recurrence were observed for up to 12 months post-infusion, despite the average duration of B-cell aplasia being only 110 ± 32 days [ 90 ]. Further immunoglobulin analyses revealed that the reconstituted B-cell populations displayed immunoglobulin light and heavy chain repertoires comparable to those found in healthy individuals [ 90 , 94 ]. This supports the notion of an immune reset, whereby autoreactive clones are eliminated, allowing for the regeneration of a more tolerant and functional B-cell repertoire (described in detail in Box 1).

In addition, the clearance of CD19-positive B cells may not fully eliminate all autoreactive components involved in the pathogenesis of ADs, as CD19-negative plasma cells can continue to produce pathogenic autoantibodies that damage healthy tissues. Dual targeting of CD19 and BCMA, or BCMA-targeted CAR T-cell therapy alone, may address this limitation in subgroups of ADs primarily driven by long-lived plasma cells [ 91 , 155 , 208 ]. Furthermore, even in autoimmune diseases primarily mediated by B-lineage cells, autoreactive T cells play a significant role, particularly those activated or sustained independently of B-cell interactions [ 209 , 210 ]. As these T cells are not targeted by anti-CD19 or anti-BCMA CAR T-cell therapy, their persistence may contribute to disease relapse or primary resistance [ 85 , 188 ].

Current clinical evidence suggests that relapses following CAR T-cell therapy in patients with ADs are rare, though not entirely absent [ 96 , 211 ]. This favorable outcome may be attributed to the immune system reconstitution facilitated by the infusion of living cellular products. However, it is important to note that long-term follow-up data remain limited, as the clinical application of CAR T-cell therapy in ADs is still relatively recent. Accordingly, extended monitoring and more longitudinal studies are required to better assess relapse risk and inform clinical practice.

Limitations to widespread CAR T cell use

Thus far, CAR T-cell therapy has primarily been employed as a salvage strategy in patients with ADs who have failed two or more lines of immunosuppressive treatment and are at high risk for permanent organ damage or mortality. However, by this advanced stage, the alleviation of disease manifestations may be limited, quality-of-life improvements may be modest, and tissue damage may be irreversible [ 188 , 212 ]. Moreover, as discussed earlier, although the risk is extremely low, CAR T-cell therapy carries the potential for severe adverse effects, including life-threatening complications [ 20 , 90 , 198 – 200 , 204 ]. Consequently, adequate functional organ reserves are essential to mitigate treatment-associated risks [ 188 ]. In this context, patients with mild-to-moderate ADs, who typically respond well to conventional therapies, may not be ideal candidates for CAR T-cell intervention, further complicating the process of patient selection.

In parallel, the quality and quantity of autologous T cells are critical for the successful manufacture and infusion of CAR T cells. Current statistics indicate that at least 7% of patients are deemed ineligible for CAR T-cell therapy due to insufficient T-cell yield or viability following leukapheresis [ 213 ]. This limitation is particularly relevant for individuals who have undergone repeated rounds of immunosuppression, which can impair T-cell integrity [ 214 , 215 ]. While glucocorticoids are generally considered the least detrimental immunosuppressive agents for T-cell preservation, their use remains controversial in this setting, underscoring the need for careful pre-treatment evaluation and optimization [ 214 – 216 ].

Standardization of CAR T manufacturing for autoimmune diseases

At present, no CAR T-cell therapies have been formally approved for the treatment of ADs. However, several investigational therapies are undergoing clinical evaluation, including trials targeting CD19 and BCMA antigens in conditions such as SLE, myositis, and systemic sclerosis. The manufacturing processes for CAR T-cell products vary among manufacturers, which may lead to variability in both product quality and therapeutic outcomes.

In addition, the extremely high cost and patient-specific variability inherent to autologous T-cell sources pose significant challenges to broader clinical adoption. Allogeneic, or “off-the-shelf,” therapies seek to overcome these limitations by using T cells from healthy universal donors, engineered for use across multiple recipients [ 15 , 217 , 218 ]. Several allogeneic CAR T-cell products are currently in clinical trials for ADs (e.g., NCT06106893 , NCT06212154 , NCT06294236 ). Even so, key obstacles remain, particularly the risks of immune rejection in both host-versus-graft and graft-versus-host directions [ 219 ]. Gene-editing technologies offer potential solutions to mitigate immune rejection and GVHD, but they also introduce new concerns, including off-target effects and genotoxicity. Moreover, due to host immune resistance and intrinsic defects introduced by gene editing, allogeneic CAR T cells are theoretically expected to exhibit shorter in vivo persistence than autologous counterparts [ 220 ]. Interestingly, clinical trials to date have reported unexpectedly prolonged survival of these cells, with detectable levels persisting beyond 2 months—and in some cases, up to 6 months post-infusion [ 221 , 222 ].

Other challenges and considerations

In clinical practice, the FDA recommends lifelong monitoring for patients who receive cell-based therapies, including CAR T-cell treatments, due to potential long-term risks. While the ultimate therapeutic goal is to achieve sustained, drug-free remission, long-term regular follow-up assessments and monitoring for potential relapses and adverse effects are necessary. In addition, the application of CAR T-cell therapy in ADs presents unique challenges. Unlike cancer patients, who often face urgent, life-threatening conditions, individuals with ADs typically experience chronic but insidious disease courses. The high cost of CAR T-cell therapy, typically ranging from $373,000 to over $500,000 per infusion [ 146 , 223 , 224 ], raises important questions about cost-effectiveness, especially when alternative, less expensive treatment options are available. Moreover, the risk-benefit balance must be carefully considered, as CAR T-cell therapy may cause serious adverse effects such as CRS and neurotoxicity. In cases where the autoimmune condition is not immediately life-threatening, these risks may outweigh the potential benefits.

Considering the adaptation of CAR T-cell therapy for ADs, standardizing clinical indications remains paramount. Currently, most physicians reserve CAR T as a salvage treatment for refractory cases [ 89 , 90 ]; however, this strategy often coincides with end-stage organ damage, limiting potential improvements in quality of life despite CAR T administration. Given that residual organ functionality is critical for tolerating therapy-related toxicities (e.g., acute kidney injury [ 225 ]), experienced clinicians advocate for preemptive intervention during the disease course—particularly in patients with high-risk prognostic markers—to maximize clinical benefit [ 188 ].

Unlike oncology, where global consensus guidelines (e.g., NCCN or EBMT criteria) define CAR T eligibility for hematologic malignancies [ 226 , 227 ], AD-specific indications lack standardized protocols [ 228 ]. Nevertheless, a framework for patient selection has emerged through expert consensus, underscoring core criteria (e.g., disease scope, organ-function threshold, and treatment center) to guide clinical trials and pragmatic use [ 229 ]. Building on this, we propose selective inclusion criteria for CAR T candidates in ADs: (1) permissible pre-treatment vaccination status to mitigate infection risks during lymphodepletion; (2) severe, refractory disease unresponsive to conventional immunosuppressants or biologic agents; and (3) preserved organ function to ensure resilience against potential adverse events. These criteria aim to balance therapeutic efficacy with safety, especially given the permanent sequelae risks in late-stage ADs and the prohibitive cost of CAR T re-treatment. Looking ahead, the establishment of quantifiable biomarkers, such as baseline cytokine profiles or organ-specific injury scores, and time-to-intervention thresholds will be critical for standardizing patient selection and optimizing long-term outcomes.

CAR-T cell therapy in autoimmune diseases

CAR T-cell therapy holds significant promise for treating ADs. Emerging evidence suggests this approach induces more profound biological alterations in autoimmune conditions compared to conventional therapies. Current clinical trial activity underscores this potential: as of this review, more than 200 studies registered on ClinicalTrials.gov are investigating allogeneic (universal) CAR-T therapies. This substantial research effort highlights the growing interest in developing effective off-the-shelf treatments for ADs, including SLE, RA, T1D, and multiple sclerosis (Table 1 ), as well as certain hematologic autoimmune disorders including ITP and AIHA (Table 2 ). Although CAR-T therapies remain associated with risks such as CRS and ICANS, numerous approaches are being pursued to refine administration protocols and manufacturing processes; these efforts aim to mitigate toxicities and enhance therapeutic efficacy (Fig. 4 ).

SLE

SLE is characterized by overactivation of polyclonal B cells and autoantibody production [ 230 ]. CD19 is broadly expressed in B cell lineages (except long-lived plasma cells) and is an ideal target for CAR-T cell therapy [ 231 ]. Injection of anti-mouse CD19 CAR-T cells in MRL-lpr mice led to persistent depletion of CD19 + B cells, improvement of lupus symptoms, delayed disease progression, and prolonged survival. Mice injected with CAR-T cells showed prolonged lifespan and regression of splenomegaly compared with mice injected with PBS (Phosphate-Buffered Saline, PBS), and the injection of 4-1BB CAR-T cells had a better improvement effect on lupus symptoms than CD28 CAR-T cells [ 232 ]. However, low-dose total-body irradiation (TBI) or lymphodepleting chemotherapy (e.g., cyclophosphamide and fludarabine) is typically required before CAR-T therapy [ 233 ].

Andreas Mackensen et al. first demonstrated that CD19 CAR-T cell therapy yielded positive long-term outcomes in patients with refractory SLE, with loss of anti-dsDNA antibodies, B-cell reconstitution, and no SLE recurrence during extended follow-up 90 . Multiple clinical studies have since confirmed that CAR-T cell therapy can provide effective and durable symptom relief in SLE [ 234 – 237 ]. Clinical follow-up has shown a marked reduction of CD19 + CD27 + memory B cells post-therapy, with no moderate or severe CRS or ICANS and no long-term myelotoxic effects [ 94 ]. In refractory SLE patients with thrombocytopenia unresponsive to steroids, cyclophosphamide, and rituximab, a single autologous CD19 CAR-T cell infusion led to increased platelet counts and decreased SLEDAI, suggesting sustained response [ 238 ].

Combined BCMA-CD19 CAR-T cell (“cCAR”) therapy has induced medication-free remission and depletion of pathogenic autoantibodies in patients with lupus nephritis [ 157 ]. Following CAR-T treatment, anti-dsDNA and anti-ssDNA antibodies were significantly reduced, along with various other autoantibodies, while the native humoral immune response was preserved [ 239 ]. Inflammatory cytokine levels also declined. Although some patients achieve long-term drug-free remission (e.g., no recurrence after 2 years), side effects such as CRS and ICANS can still increase infection risk. CRS may be alleviated with tocilizumab and glucocorticoids [ 240 ]. The persistence of CAR-T cells in vivo and the stability of immune reconstitution require further study and data [ 241 ].

RA

RA is a chronic autoimmune disease marked by persistent inflammation and joint damage, with self-reactive B cells playing a major role in pathogenesis. CD19 CAR-T cell therapy induces complete depletion of bone marrow B cells, concurrent with PR3-ANCA seroconversion, supporting the effectiveness of B-cell targeting in organ-specific autoimmunity [ 242 ]. CD19 CAR-T appears more effective than CD20 monoclonal antibodies, likely due to targeting a broader B-cell population, including plasma cell precursors [ 243 , 244 ]. Universal CAR-T therapies can specifically eliminate B cells secreting autoantibodies (such as ACPA + B cells) via a FITC-antigenic peptide linker, with efficacy correlated to linker dose. In a collagen-induced arthritis mouse model, autoantibody-secreting cells in joint synovium were significantly reduced, relieving inflammation [ 245 ].

Clinical studies show that 70% of RA patients unresponsive to biologics or targeted agents achieved ACR50 after CAR-T therapy, with a 60% reduction in ultrasound synovitis scores, decreased inflammation, and a 40% drop in ACPA levels, though RF antibodies were unchanged [ 246 , 247 ]. This suggests B-cell targeting may be more effective against specific antibody subtypes. In clinical trials, 80% of patients experienced grade 1–2 CRS (fever, fatigue), with no serious ICANS events. Long-term follow-up showed infection rates similar to healthy controls and no opportunistic infections [ 247 ]. Neutropenia (47.8%) and anemia (20.3%) were manageable post-therapy [ 248 ]. High CAR-T doses (e.g., 11 × 10⁶ cells/kg) may increase CRS risk, highlighting the need for dose optimization or use of reduced co-stimulatory domains [ 246 ]. Fourth-generation CAR-T therapies, such as those co-expressing IL-6 antibodies, have been reported to lower CRS risk by locally remodeling the inflammatory microenvironment [ 249 ].

MS

MS is a chronic autoimmune disease involving abnormal B and T cell activation in the CNS [ 250 ]. Current therapies, such as CD20-targeted monoclonal antibodies, are limited in advanced cases. CAR-T therapy depletes CD19 + B cells [ 251 ]. Animal studies show anti-CD19 CAR-T cells deeply deplete CNS B cells in EAE models [ 252 ]. Clinically, CAR-T cell expansion in cerebrospinal fluid, reduced OCBs, and lower EDSS scores have been observed in progressive MS patients post-treatment. This suggests that CAR-T enrichment in the CNS is linked to reduced intrathecal antibodies and enables targeting of central B cells via the blood-brain barrier [ 97 , 253 ].

Bispecific CAR-T approaches, such as CAAR-T (targeting autoantigens) and CATCR-T (targeting T cell receptors), can more precisely eliminate pathogenic cells and reduce off-target risk [ 244 ]. CD19 targeting may excessively deplete normal B cells, raising infection risk; BCMA targeting focuses more on plasma cells but may impact long-lived plasma cells in bone marrow [ 254 ]. Importantly, CAR-T-associated neurotoxicity (such as ICANS) remains a risk, especially in MS patients with active CNS inflammation [ 255 ].

T1D

T1D is a metabolic disease characterized by absolute insulin deficiency caused by autoimmune destruction of pancreatic beta cells. Clinical trials have shown that treatment with Fc receptor-nonbinding anti-CD3 monoclonal antibodies can delay T1D progression by modulating CD8 + T lymphocytes, the effectors that destroy beta-like cells [ 256 ] ( NCT01030861 ). Ma et al. used the human ESC genome project to develop a CD19-CAR-T autoimmune diabetes mouse model, showing that CAR-T cells could kill beta-like cells expressing CD19 both in vitro and in vivo, and that PDL1 overexpression in beta-like cells conferred resistance against reactive T cells [ 257 ]. A single infusion of 287-CAR-T cells temporarily suppressed islet autoimmunity and delayed T1D onset in NOD mice, but this protection was not sustained. Studies indicate that CAR-T therapy does not significantly alter immune cell profiles [ 258 ]. Current research focuses on developing allogeneic universal CAR-Tregs to overcome limitations of individualized approaches. The observed improvement in C-peptide levels among patients receiving autologous polyclonal regulatory T cell therapy, along with the in vivo detection of labeled therapeutic cells, supports the potential efficacy of this immunomodulatory approach in preserving β-cell function in type 1 diabetes [ 259 ]. However, most of the clinical cases of CAR-T therapy for T1D are still under investigation. Specific enrichment serves as the crucial mechanism through which A2-CAR-Tregs exert a potent protective effect. This mechanism may contribute to promoting infection tolerance by modulating the local immune microenvironment. For instance, it can induce the polarization of M2 macrophages and inhibit effector T cells, among other regulatory functions. Genetic engineering is expected to overcome the bottleneck of current Treg therapy and achieve more precise, effective and durable immune tolerance induction [ 260 – 262 ].

Other autoimmune diseases

Clinical studies have shown that BCMA CAR-T targeting plasma cells, such as Descartes-08, leads to clinically meaningful improvement in MG by reducing anti-AChR antibody levels, with no CRS or ICANS observed [ 93 , 95 , 263 ]. Rituximab offers limited benefit in skin sclerosis, whereas CD19 CAR-T has been associated with >50% reductions in mRSS and stable lung function in multiple studies. In diffuse systemic sclerosis, mRSS decreased significantly post-therapy, DLCO improved to 55%, and cardiac MRI showed less fibrosis [ 15 , 247 ]. In immune-mediated necrotic myopathy (IMNM), autologous CD19 CAR-T cell therapy significantly improved myasthenia, with CK levels dropping by 80% and anti-HMGCR antibody titers declining to the lower limit of detection [ 264 ].

In summary, CAR T-cell therapy has demonstrated substantial progress in treating autoimmune diseases, but further clinical trials are needed to validate efficacy, and improved clinical management strategies remain essential.

Innovative strategies in CAR-T cell therapy for autoimmunity

Recent advances in CAR-T cell engineering aim to improve specificity, safety, and long-term efficacy, responding to the unique challenges posed by ADs. Beyond CD19 and BCMA, emerging targets such as CD7 and B-cell receptor–derived epitopes allow more selective elimination of pathogenic lymphocyte subsets. Concurrently, innovations in CAR architecture—including bispecific and synthetic TCR-based designs—improve signaling precision, while allogeneic or iPSC-derived platforms offer scalable, off-the-shelf alternatives. These converging strategies underpin the next generation of CAR-T therapies tailored for chronic autoimmune conditions. (Fig. 5 ).

CAR-T structure modification

The architecture of the chimeric antigen receptor (CAR) is critical in determining specificity, activation kinetics, and safety of CAR-T cells. Structural innovations include adjustments to the antigen-binding domain (scFv), hinge, transmembrane, and intracellular signaling domains, all intended to enhance precision and reduce off-target effects (Fig. 3 A).

The scFv domain is pivotal for determining CAR-T cell function [ 265 ]. For instance, Kang et al. used the KHYG-1 NK/T-cell line to screen eight different CD19 scFvs and found that 4G7 performed as well as FMC63, the most widely used scFv, offering an alternative for patients resistant to FMC63-based CAR-T therapy [ 266 ]. Drent et al. compared CD38 CARs with different antigen affinities and found that low-affinity CAR-T cells (A4) showed anti-myeloma efficacy similar to high-affinity CAR-T (028) in vitro and in vivo, but with less fratricide and reduced toxicity to normal CD38 + cells [ 267 ]. Grada et al. introduced TanCAR, a bispecific CAR with two scFv domains that can each recognize a different target, increasing the chance of effective activation and reducing antigen escape; however, the dual-scFv architecture carries a theoretical risk of cross-reactivity due to potential mispairing between light and heavy chain variable domains of distinct scFvs, which could generate neo-specificities—though this has not been observed in preclinical models to date [ 268 – 270 ].

Co-stimulatory signals, such as CD28 and 4-1BB, play a crucial role in CAR-T cell function by influencing persistence, apoptosis resistance, and cytotoxic ability. CD28 promotes rapid expansion but shorter cell persistence, while 4-1BB prolongs cell survival and reduces exhaustion [ 271 ]. Roseli et al. designed a CD19 CAR-T with both 4-1BB and a mutant CD28 domain, enhancing persistence and reducing exhaustion, but as Wijewarnasuriya et al. reported, excessive co-stimulation can cause T-cell dysfunction [ 272 ]. Despite these advances, the full complexity of CAR-T signaling remains unraveled, and further research into the interplay of co-stimulatory pathways is needed to optimize efficacy and safety.

Another prominent innovation is CAAR-T cell therapy. Unlike conventional CAR-T, CAAR-T cells have an extracellular domain derived from autoantigen, enabling them to specifically recognize and eliminate B lymphocytes with self-reactive BCRs [ 160 ]. For example, studies of CAAR-T cells expressing the Dsg3 domain have demonstrated superior and more durable effects in treating pemphigus vulgaris compared to CD19 CAR-T cells, providing long-lasting relief and safety [ 273 ].

Target selection beyond CD19 and BCMA

CD19 and BCMA remain the primary targets for CAR-T therapy in autoimmune diseases such as SLE, lupus nephritis (LN), and systemic sclerosis (SSc). As of this review, 95 clinical trials involving CD19, BCMA (Table 1 ) CAR-T for ADs are registered on clinicaltrials.gov. However, concerns about broad B-cell depletion—including prolonged immunosuppression and infection risk—have prompted intense interest in expanding target diversity [ 274 , 275 ](Table 3 ). For instance, NCT06279923 explores CD19-BAFF CAR-T cells targeting multiple antigens in various autoimmune diseases, demonstrating enhanced specificity and efficacy. CD7 is emerging as a new target, particularly in SSc and other severe T-cell–mediated autoimmune conditions. Studies combining single-cell transcriptomics, flow cytometry, and multiplex immunofluorescence of samples from SSc patients and healthy controls show that CD7 regulates cytolytic activity of cytotoxic T cells and NK cells. CD7 is highly expressed on pathogenic cytotoxic CD4⁺ and CD8⁺ T cells as well as NK cells that infiltrate affected tissues and promote fibrosis [ 276 ]. Selective depletion of CD7 + cells inhibits fibroblast contraction and stabilizes disease in severe SSc, positioning CD7 as a promising target for selective removal of pathogenic cells [ 276 ]. Additionally, NCT05239702 evaluates the safety and efficacy of CD7 CAR-T cells in refractory Crohn’s disease and ulcerative colitis, highlighting the versatility of this approach across different autoimmune conditions.

CD20 has been widely used in antibody therapies (e.g., rituximab) and is now being explored for CAR-T therapy in diseases such as MS and T1D [ 277 ]. In MS, CD20⁺ B cells act as antigen-presenting cells and cytokine producers within the CNS, sustaining neuroinflammation and demyelination; in T1D, they support autoreactive T-cell responses in pancreatic islets [ 278 , 279 ]. Preclinical MS models demonstrate that anti-CD20 CAR-Ts can cross the blood-brain barrier, eliminate infiltrating B cells, and attenuate demyelination [ 280 , 281 ]. Nevertheless, safety concerns persist—specifically regarding long-term immunosuppression—prompting the development of engineered CARs with transient or regulated activity [ 282 ]. Improved anti-CD20 antibodies remain under investigation in ongoing clinical trials [ 283 ].

Beyond single-product CAR-T regimens, co-infusion of distinct CAR-T populations targeting complementary antigens offers a pragmatic strategy to broaden efficacy while preserving manufacturing flexibility. For example, NCT06056921 reports on the safety and efficacy of sequential infusion of anti-CD19 and anti-BCMA CAR-T cells in refractory systemic lupus erythematosus, achieving significant remission rates without dose-limiting toxicities. In a recent phase 1 trial in refractory systemic lupus erythematosus, co-administration of autologous anti-CD19 and anti-BCMA CAR-T cells achieved 80% remission rates at 12 weeks with no dose-limiting toxicities, effectively eliminating both circulating B cells and long-lived plasma cells—the dual sources of pathogenic autoantibodies [ 284 ]. This aligns with other emerging reports: a patient with treatment-resistant ulcerative colitis attained drug-free remission and mucosal healing after CD19 CAR-T therapy [ 285 ], and five patients with severe lupus nephritis achieved sustained clinical and histological remission following infusion of YTS109, a CRISPR-engineered allogeneic anti-CD19 STAR-T cell product lacking TCR and HLA expression [ 286 ]. However, repeated CAR-T infusions, particularly in non-lymphodepleted settings, may provoke anti-CAR immune responses that limit re-dosing. A recent case series in pediatric CNS tumors highlighted the development of neutralizing anti-CAR antibodies after multiple intracerebroventricular administrations, leading to loss of CAR-T persistence and therapeutic resistance [ 287 ]. These findings underscore the need for strategies to mitigate immunogenicity—such as humanized scFvs or transient expression platforms—when designing multi-dose or multi-target regimens for chronic autoimmune conditions.

Method of CAR gene transfer

Gene delivery for CAR expression is central to CAR-T production. Current methods include viral vectors (lentivirus, retrovirus) and non-viral systems (transposons, CRISPR-Cas9) [ 288 ]. Viral vectors ensure stable genomic integration and high transduction efficiency but raise concerns about insertional mutagenesis and manufacturing complexity [ 289 ]. Non-viral approaches, such as mRNA electroporation, provide transient CAR expression and may reduce long-term immunosuppression risks. Newer platforms like nanoparticle-mediated delivery and Sleeping Beauty transposons are being optimized for higher efficiency and safety in AD applications [ 290 , 291 ](Fig. 3 B).

Gene-editing synergies

Gene-Editing and Synthetic Biology approaches, particularly CRISPR-Cas9, enable precise deletion of immune checkpoint genes (e.g., PD-1, CTLA-4) in CAR-T cells, enhancing their resistance to inhibitory signals and sustaining effector function in inflammatory microenvironments [ 292 ]. Synthetic biology is driving innovations such as AND/NOT logic gates, which require dual-antigen recognition or exclude normal tissue antigens, increasing specificity for heterogeneous AD targets and minimizing off-tissue toxicity [ 293 , 294 ].

Another major safety innovation is the integration of “suicide switches” into CAR-T cells, allowing for irreversible cell elimination upon administration of an external agent in case of severe toxicity. However, this approach eliminates all CAR-T cells at once [ 295 ]. Reversible “off switches,” which temporarily inhibit CAR-T activity, have also been developed; these use drugs or soluble factors to modulate CAR-T function without cell death [ 296 , 297 ]. NOT-gated CAR-Ts seek to integrate safety directly into antigen recognition logic, preventing activation against healthy tissues.

Optimization of CAR-T cell production

Optimizing CAR-T manufacturing is essential for broader accessibility, reduced costs, and improved therapeutic outcomes in ADs. Two key innovations are in vivo CAR-T generation and allogeneic “off-the-shelf” CAR-T therapies [ 298 ](Fig. 3 C). (1) In Vivo CAR-T Generation: This approach bypasses traditional ex vivo manufacturing by directly transducing patient T cells in vivo using viral (e.g., AAV) or non-viral (e.g., LNP) vectors. Lentiviral vectors are common but limited by scalability and cost. Alternatives like AAV enable stable CAR integration, while antibody-conjugated LNPs allow targeted mRNA delivery to T cells [ 299 – 301 ]. For example, CD3-targeted LNPs delivering IL-6 shRNA and CD19 CAR mRNA have generated functional CAR-T cells in vivo and mitigated CRS [ 302 ]. These methods eliminate ex vivo manipulation, shorten production time to days, and lower costs [ 303 ]. (2) Allogeneic CAR-T Therapies: Healthy donor- or iPSC-derived CAR-T cells are advancing as scalable “off-the-shelf” options [ 304 , 305 ]. iPSCs reprogrammed from mature T cells retain TCR rearrangement and can be differentiated into potent CD8αβ + CAR-T cells [ 306 ]. Gene editing (e.g., CRISPR/Cas9) addresses risks such as graft-versus-host disease (GVHD) by knocking out TCR or HLA genes [ 307 ]. Triple-edited CAR-T cells (TCR-/HLA-I-/HLA-II-) have shown improved persistence and NK cell evasion [ 308 ]. Donor-derived CAR-T, in clinical trials, have demonstrated low GVHD and promising efficacy [ 309 ]. Preconditioning regimens and pharmacokinetic modeling are being refined to optimize engraftment and persistence [ 310 , 311 ]. Notably, NCT06106893 evaluates the safety and efficacy of universal CAR-γδT cells in active systemic lupus erythematosus, representing a step forward in allogeneic CAR-T therapies for autoimmune diseases.

Notably, both in vivo and allogeneic approaches offer the potential to eliminate the need for lymphodepleting preconditioning—a major advantage in autoimmune settings, where chemotherapy-induced cytopenia and infection risk are particularly concerning [ 312 ]. The first-in-human application of in vivo CD19 CAR-T therapy using CD8-targeted lipid nanoparticles (HN2301) in five patients with refractory systemic lupus erythematosus demonstrated rapid B-cell depletion, reduction in disease activity, and only low-grade cytokine release syndrome without lymphodepletion or severe toxicity, supporting its feasibility in ADs [ 313 ]. Importantly, emerging data suggest that durable clinical responses in autoimmunity do not require permanent B-cell ablation but instead reflect an “immune reset” driven by transient, profound depletion of pathogenic B-cell subsets. This reset is mechanistically characterized by the near-complete elimination of autoreactive memory B cells and plasmablast precursors, followed by reconstitution from bone marrow progenitors that repopulate the periphery with a naïve, non-autoreactive B-cell repertoire [ 186 ]. Concurrent restoration of T-cell homeostasis and reduction in inflammatory mediators further support long-term tolerance. Preclinical studies confirm that lymphodepletion-free CAR-T regimens can achieve this reset: a single dose of MDF-modified VivoVec particles induced sustained B-cell aplasia for over 10 weeks in non-human primates without preconditioning, markedly outperforming conventional ex vivo CAR-T products that required lymphodepletion yet exhibited B-cell rebound within 5 weeks [ 314 ]. Similarly, CD8-targeted LNPs delivering transient CAR mRNA achieved rapid B-cell clearance and features of immune reset without genotoxic integration or chemotherapy [ 186 ]. These findings support a paradigm shift toward safer, preconditioning-free CAR-T strategies tailored to the distinct therapeutic goals of autoimmunity.

For vector and Process Innovations: Non-viral platforms like transposons and LNP-mRNA co-delivery avoid genome integration risks and improve safety [ 315 ]. Combinatorial strategies—such as delivering CAR mRNA alongside siRNA to knock down PD-1—enhance antitumor activity without impairing T cell activation [ 316 ]. AI-driven process optimization is being applied to standardize CAR-T production and assure batch consistency [ 317 , 318 ].

Expanding the scope: targeting other immune cells

Growing recognition of the heterogeneous cellular landscape underlying autoimmune pathology has stimulated exploration of diverse immune cell sources for CAR engineering. The choice of source cell population—such as Tregs, NK cells, or macrophages—fundamentally shapes the therapeutic potential of CAR-based immunotherapy, influencing cytotoxic efficiency, immune tolerance, and safety profiles [ 319 ]. Enhancing efficacy in autoimmune diseases may thus depend on strategically selecting or genetically modifying optimal immune cell subsets during the CAR manufacturing process (see Fig. 6 ).

CAR-modified regulatory T cells

Tregs, defined by their CD25 and FOXP3 expression, are essential enforcers of immune homeostasis, maintaining self-tolerance and suppressing pathogenic autoreactivity in autoimmune diseases [ 320 , 321 ]. Their potential has been increasingly recognized in both transplantation tolerance and autoimmunity.

CAR-Tregs are generated by equipping Tregs with CARs tailored to recognize specific antigens, thereby redirecting their potent suppressive function to disease-relevant tissues. Numerous innovations in CAR design—such as choice of co-stimulatory domain and antigen specificity—have optimized Treg stability, persistence, and suppressive efficacy. For example, CD28-containing CAR-Tregs displayed superior immunosuppressive function in vivo, potently restraining effector T cell responses and allograft rejection [ 322 ]. Overexpression of FOXP3 in CAR-Tregs further enhances their stability and therapeutic function, as demonstrated by FoxP3-overexpressing anti-CD19 CAR-Tregs (Fox19 CAR-Tregs) that disrupted autoimmunity and tissue injury in SLE models [ 323 ]. Similarly, FoxP3-CAR-Tregs have shown robust efficacy in preventing allograft rejection and efficiently filling the Treg niche, supporting their clinical translation [ 324 ].

CAR-Tregs have been explored for a broad range of indications. In multiple sclerosis, MOG-specific CAR-Tregs exhibited minimal tonic signaling with strong disease suppression in EAE models [ 325 ]. In T1D, MHC class II peptide-specific CAR-Tregs prevented autoimmune diabetes in mice [ 193 ]. Studies have also targeted key immune modulators, the researchers used XCR1-targeting CAR-T cells for selective depletion and immunomodulation of pathogenic cDC1s, modestly suppressing Th1-driven neuroinflammation in EAE [ 326 ]. Additionally, Engagement of OX40L-expressing cells by OX40L-specific CAR-Tregs triggers suppression via trogocytosis, resulting in the removal of surface OX40L. This process attenuates dendritic cell stimulation and potentiates the intrinsic function of the Tregs, without altering their anti-inflammatory cytokine secretion profile [ 327 ]. CAR-Tregs have demonstrated promise in clinical and preclinical models for GVHD, asthma, hemophilia, transplantation, and diverse autoimmune diseases [ 328 ]. Their therapeutic efficacy hinges on maintaining FOXP3 expression, antigen specificity, and persistence in hostile inflammatory microenvironments. Advanced engineering—such as constitutive FOXP3 expression or CRISPR-mediated knockout of destabilizing genes (e.g., IFN-γ or IL-17)—is under investigation to further stabilize the regulatory phenotype [ 324 , 329 , 330 ]. Collectively, these advances underscore the capacity of CAR-Tregs to restore immune tolerance and resolve inflammation with high antigen specificity.

CAR-NK cells in autoimmune diseases

CAR-NK cells have emerged as an attractive alternative to CAR-T cells, offering distinct advantages such as reduced risk of cytokine release syndrome, lower graft-versus-host potential, and broader applicability [ 331 – 333 ]. Aberrant Tfh cell responses drive many autoimmune conditions. Reighard et al. developed PD-L1-expressing CAR-NK cells to selectively deplete PD-1 hi Tfh cells, showing efficacy both in vitro and in humanized mouse models [ 334 ].

Autoantibodies against La/SSB protein are involved in early-stage SLE, Sjögren’s syndrome (SS), and neonatal lupus [ 335 ]. By engineering the La/SSB epitope into a CAAR expressed in NK92MI cells, researchers demonstrated specific elimination of La/SSB-autoreactive B-cell clones [ 336 ]. These findings underscore the potential for CAR-NK cells to target auto-reactive lymphocytes and modulate disease activity with a favorable safety profile.

Macrophage-targeting CAR approaches

CAR-macrophages (CAR-Ms) leverage the innate phagocytic capacity and tissue-penetrating abilities of macrophages to eliminate pathogenic and abnormal cells, representing a new frontier in CAR-based cell therapy [ 337 , 338 ]. Recent preclinical models demonstrate that CAR-Ms targeting fibroblast activation protein (FAP) effectively infiltrate myocardial tissue and enhance cardiac function in murine models of fibrosis [ 339 – 341 ]. Given the contribution of organ fibrosis to advanced autoimmune diseases, CAR-Ms—with their superior tissue access—offer novel strategies for treating fibrotic manifestations such as cutaneous and pulmonary fibrosis.

To address resistance to efferocytosis, researchers developed CD47-targeted CAR-M platforms capable of engulfing apoptosis-resistant cells. Combined with lipid nanoparticles designed to enhance liver X receptor (LXR) signaling, these CAR-Ms exhibited improved phagocytic function [ 342 ]. Macrophage polarization plasticity also presents therapeutic opportunities: by engineering a CAR-M system incorporating α1β1 integrin-mediated FcγRI signaling, scientists promoted pro-inflammatory M1 polarization, thereby enhancing tumor control and survival via M1-associated pathways [ 343 ]. Moreover, Lei A et al. developed induced pluripotent stem cell–derived macrophages (iMACs) expressing TIR domain-containing CARs, which induced NF-κB-dependent M1 polarization, suppressed M2 phenotypes, and modified the tissue microenvironment—suggesting potential applications in modulating autoimmune tissue pathology [ 344 ].

In summary, CAR-engineered immune cells—including CAR-Tregs, CAR-NK, and CAR-M—represent a new generation of precision therapies for autoimmune diseases (Table 4 ). CAR-Tregs restore immune tolerance through antigen-specific suppression; CAR-NK cells selectively eliminate pathogenic cells while preserving protective immunity; and CAR-M therapies uniquely combine targeted cell clearance with inflammation resolution and tissue repair. Although each approach demonstrates transformative potential, their clinical success will depend on further engineering to optimize targeting specificity, persistence, and functional control within diseased tissues.

Challenges and future perspectives

Application of novel technologies

Current advances in CAR-T cell engineering are enriched by the integration of diverse, cutting-edge technologies designed to enhance therapeutic efficacy, predict and reduce toxicities such as CRS, and individualize cell therapy for autoimmune diseases [ 345 ]. Notably, Artificial Intelligence (AI), organoid technology, and microphysiological systems (MPS) are at the forefront of these innovations, each providing unique capabilities for discovery, optimization, and translation in the CAR-T field (Fig. 5 d).

Artificial intelligence

AI, encompassing both machine learning (ML) and deep learning (DL), is transforming the landscape of CAR-T cell therapy. By enabling the rapid processing and analysis of multidimensional datasets, AI is being harnessed to optimize all critical steps in CAR-T cell therapy development. This includes target antigen selection, vector design, manufacturing process optimization, and data-driven clinical decision-making [ 346 ].

Recent investigations have focused on the application of ML algorithms to refine CAR architecture and preemptively model CRS, with the goal of enhancing efficacy and safety while making therapies more accessible and affordable. For example, Lupeng Ye et al. established a dead-guide RNA (dgRNA)-mediated CRISPR activation screening platform in primary CD8 + T cells, which enabled the identification of gain-of-function (GOF) targets for CAR-T engineering. Among these, PRODH2 was found to be a promising target, enhancing CAR-T cell efficacy in multiple cancer models [ 347 ].

The clinical translation of CAR-T therapies, however, faces significant economic and logistical hurdles, particularly in manufacturing and scaling production cost-effectively [ 348 ]. To address these, AI-driven platforms are being designed in advanced smart hospitals to provide real-time process characterization and quality control. Integration of ML algorithms can utilize real-time data to facilitate adaptive automation, dynamically adjusting production parameters to enhance cell growth and viability while minimizing manual intervention [ 349 ]. This could significantly reduce both costs and human error in CAR-T cell production.

Moreover, DL has shown promise in predicting patient-specific therapeutic responses using comprehensive clinical and biological data, thereby facilitating personalized treatment optimization. For example, Wei et al. demonstrated that AI could accurately predict and evaluate adverse events, including CRS, prior to therapy [ 350 ]. Similarly, AI has been used to predict outcomes of bridging radiotherapy (BRT) in relapsed/refractory large B-cell lymphoma patients awaiting CAR-T therapy, where all treated patients achieved symptomatic improvement and subsequently received successful CAR-T infusion [ 351 ].

Despite these advances, the successful implementation of AI/ML in CAR-T cell immunotherapies requires rigorous quality controls, robust and standardized datasets, and careful handling of biological heterogeneity and variable study designs. The ongoing evolution of regulatory and ethical frameworks will also be crucial for the safe and effective integration of AI in clinical practice.

Organoids

Organoid technology has emerged as a transformative tool in cell therapy research, enabling the generation of three-dimensional, patient-specific tissue models that closely recapitulate the in vivo microenvironment. These organoids serve as valuable platforms for optimizing CAR-T cell production, assessing antitumor activity, and understanding cellular interactions prior to clinical translation.

Multiple studies have utilized patient-derived glioblastoma organoids (GBOs) to evaluate the real-time bioactivity of CAR-T cells, providing insights into immunotherapy efficacy, tumor cell killing, and resistance mechanisms [ 352 , 353 ]. These models allow researchers to simulate complex tumor-immune interactions, predict therapeutic outcomes, and identify potential obstacles to CAR-T cell infiltration and persistence.

Organoid modeling has also revealed strategies to enhance CAR-T cell efficacy against solid tumors. For example, co-expression of CCR2b in CAR-T cells enhances chemotaxis, thereby improving their migration toward and infiltration into solid tumor brain metastases [ 354 ]. Additionally, Wang et al. utilized artificial thymic organoids (ATOs) to derive functional CD19-targeting CAR-T cells from human induced pluripotent stem cells (iPSCs), showing that these cells exhibit comparable phenotypic characteristics and anti-leukemic efficacy as conventional CAR-T products in human CD19 + leukemia xenograft models [ 306 ]. This approach holds promise for producing standardized, scalable, and patient-specific CAR-T products, potentially overcoming limitations of donor-derived cells.

However, challenges remain in ensuring the reproducibility, scalability, and standardization of organoid cultures. Continued integration with high-content imaging, single-cell sequencing, and automated analysis will be essential to fully realize the potential of organoid platforms in CAR-T research and therapy development.

Microphysiological systems

Advancements in microphysiological systems (MPS), often referred to as organ-on-a-chip (OoC) platforms, are providing new in vitro models that recapitulate the dynamic biochemical, biophysical, and mechanical environments of human tissues [ 355 ]. These microfluidic-based systems emulate key features of the human microenvironment, making them invaluable for preclinical CAR-T research, especially in modeling TME. For example, breast cancer-on-chip platforms incorporating integrated endothelial barriers have been developed to study real-time immune cell transmigration, tumor infiltration, and cytokine release during perfused cultures [ 356 ]. This allows for a more accurate assessment of CAR-T cell trafficking, activation, and functional responses in a complex, physiologically relevant context. Benteng Ma et al. employed a digital nanoplasmonic biosensing platform to quantitatively monitor cytokine concentrations—including TNF-α, IFN-γ, MCP-1, GM-CSF, IL-1β, and IL-6—during the first five days of CAR-T cell therapy in a microfluidic leukemia-on-a-chip model [ 357 ]. These high-sensitivity measurements provide critical insights into the kinetics of immune activation and the pathogenesis of CRS, supporting the development of safer and more effective therapies.

The modular design of tumor-on-chip platforms enables systematic investigation of diverse cell types within the TME, facilitating bench-to-bedside translation and accelerating preclinical development of CAR-T cell therapies. While technical complexity and cost remain barriers to widespread adoption, ongoing innovations in microfabrication, imaging, and data analysis are rapidly enhancing the utility of MPS in CAR-T research.

Combinational strategies

Despite remarkable clinical advances, maintaining the long-term efficacy of CAR-T cell therapy—especially in the context of immune evasion, exhaustion, and the suppressive microenvironment—remains a significant challenge [ 358 ]. Recent research emphasizes the importance of combinational strategies, including metabolic interventions and the use of molecular targeted drugs, to enhance therapeutic outcomes, reduce resistance, and improve safety [ 359 ] (Fig. 7 ).

Metabolic interventions

Metabolic regulation critically influences CAR-T cell persistence, capacity to overcome immunosuppression, and maintenance of functional phenotypes. Both glycolytic and mitochondrial oxidative phosphorylation (OXPHOS) pathways are essential for maintaining cellular homeostasis [ 360 ]. Physiologically, naïve T cells rely on OXPHOS for survival, but after antigen stimulation, they differentiate into effector T cells and shift their metabolic strategy toward aerobic glycolysis to support proliferation and effector functions [ 361 ].

Recent investigations have shown that overexpression of glucose transporters (GLUTs) augments metabolic adaptability and tumoricidal capacity of CAR-T cells [ 362 – 364 ]. Specifically, GLUT1 overexpression enhances glycolytic flux, OXPHOS, and metabolic reserve, correlating with reduced exhaustion markers and promoted Th17 polarization [ 362 , 363 ]. Similarly, GLUT3 overexpression activates the PI3K/Akt/mTOR signaling axis, improving mitochondrial bioenergetics and enhancing antitumor potency [ 363 ].

Mitochondrial loss and dysfunction are major contributors to T cell exhaustion and represent significant barriers to CAR-T efficacy [ 365 ]. CD38 has emerged as a critical regulator; its inhibition suppresses the CD38-cADPR-Ca²⁺ signaling pathway while activating the CD38-NAD⁺-SIRT1 axis, leading to glycolytic suppression and improved T cell function [ 366 ]. In lupus, BCMA-expressing germinal center B cells depend on glucose oxidation; combining APRIL-based CAR-T cells with glycolysis inhibition has been shown to synergistically deplete pathogenic B cells and extend the lifespan of lupus-prone mice [ 367 ].

Abnormal lipid metabolism impairs immune cell function by altering metabolism and generating byproducts such as ROS and lactate, which support the formation of an immunosuppressive microenvironment by recruiting Tregs, M2 macrophages, and MDSCs [ 368 ]. Lipid peroxidation and endoplasmic reticulum (ER) stress further drive immune suppression, while activation of HIF-1α promotes glycolysis and amino acid metabolism in Teff cells, inhibiting fatty acid oxidation and leading to Teff dysfunction [ 369 ]. Tregs, in contrast, exhibit a metabolic profile marked by elevated lipid metabolism and reduced glycolysis/OXPHOS [ 370 ]. Targeting lipid metabolism thus represents a novel approach to remodel the TME.

Preclinical studies demonstrate that CAR-T cells engineered to co-express FOXP3 undergo distinct metabolic reprogramming, characterized by downregulated glycolysis/OXPHOS and upregulated lipid metabolism, which enhances their therapeutic efficacy [ 371 ]. In addition, targeted reduction of lipid peroxidation via GPX4 overexpression, combined with limiting fatty acid transport through CD36 inhibition, has been shown to rescue the function of CD8 + TILs [ 372 ].

Amino acids are also essential for T cell immune responses, regulating activation and proliferation. Tryptophan metabolism by IDO produces kynurenine, which impairs T cell responses and promotes Treg differentiation. Leucine regulates T cell proliferation via the mTOR pathway [ 218 ]. These findings highlight the intricate crosstalk between metabolic pathways and immune function, and suggest that metabolic modification of both CAR-T cells and the immune microenvironment can potentiate immunotherapy efficacy.

Despite early clinical progress, further studies are needed to clarify the precise metabolic requirements of immune cells in different disease contexts and to optimize metabolic interventions for enhanced safety and efficacy (Fig. 8 ).

Molecular targeted drugs

Maintaining durable CAR-T cell activity and safety in autoimmune diseases is a central therapeutic challenge. Unlike cancer patients, autoimmune diseases patients often have a lower B-cell burden, and antigen escape following therapy may contribute to relapse. Molecular targeted drugs have emerged as important tools for enhancing antigen expression, overcoming resistance, and improving the cytotoxic efficacy of CAR-T cells [ 373 ].

In autoimmune diseases, immune suppression mediated by co-inhibitory molecules like PD-1, TIM-3, CTLA-4, LAG-3, and TIGIT drives T cell exhaustion and establishes an immunosuppressive microenvironment, constituting a key mechanism of resistance [ 374 , 375 ]. A variety of molecular targeted agents—including hypomethylating agents, TKIs, BTK inhibitors, PI3K inhibitors, and immune checkpoint inhibitors—have shown the ability to reverse exhausted phenotypes of CAR-T cells and enhance their proliferation and activity [ 373 ]. CRISPR/Cas9-mediated knockout of PD-1 further enhances cytokine secretion and in vivo efficacy by downregulating exhaustion-associated gene signatures [ 376 ].

Macrophages, Tregs, MDSCs, and DCs are major contributors to the immunosuppressive microenvironment, presenting significant barriers to CAR-T therapy success [ 377 , 378 ]. Targeting immunosuppressive molecular pathways, such as OX40/OX40L signaling, has been shown to reduce T cell exhaustion by activating NF-κB, MAPK, and PI3K-AKT pathways [ 379 ]. Additionally, innovative strategies such as engineering CAR-T cells to activate small-molecule drugs in situ are under investigation [ 380 ].

CAR-T cell therapy is also associated with unique toxicities, including CRS, ICANS, and cytopenias [ 239 ]. Key pathways such as JAK-STAT, TLR signaling, NET formation, and NLRP3 inflammasome activation are implicated in CRS pathogenesis [ 381 ]. Targeted agents—including JAK inhibitors, TNF blockers, dasatinib, tocilizumab, GM-CSF inhibitors, mTOR inhibitors, and lenzilumab—can effectively manage these toxicities without adding significant safety risks [ 382 ]. However, most molecular targeted drug combinations with CAR-T therapy in ADs remain in the preclinical stage; comprehensive studies are required to fully elucidate their toxicity and safety implications.

Conclusion

CAR-T cell therapy has rapidly emerged as a transformative approach for the treatment of autoimmune diseases, building on its established success in hematologic malignancies. Unlike conventional immunosuppressive therapies that require prolonged, sometimes lifelong administration and carry risks of infection or malignancy, CAR-T cell therapy is distinguished by its innovative design: a single, personalized infusion of genetically engineered autologous T cells, programmed to selectively target and eliminate autoreactive immune cells. This strategy is rooted in the concept of immune system “reset”—an intervention with the potential to induce deep, treatment-free remissions by eradicating the cellular drivers of autoimmunity.

The promise of CAR-T therapy in autoimmune diseases lies not only in its profound efficacy—demonstrated by early clinical trials in conditions such as systemic lupus erythematosus, myasthenia gravis, and autoimmune cytopenia—but also in its capacity to fundamentally alter the therapeutic paradigm. Rather than chronic suppression of immune activity, CAR-T therapy aims for durable immunological rebalancing, restoring tolerance, and allowing patients to achieve sustained remission without the burden of ongoing medication. This represents a significant advance over current standards of care and offers hope for patients with refractory or severe disease.

However, the translation of CAR-T therapy from oncology to autoimmunity brings unique challenges. The complex, individualized manufacturing process, the risk of acute toxicities such as cytokine release syndrome, and questions about the durability and mechanism of remission all require continued investigation. Furthermore, the heterogeneity of autoimmune diseases, the diversity of pathogenic immune cell subsets, and the dynamic nature of immune regulation necessitate ongoing research to optimize CAR design, target selection, and patient selection.

Large-scale, controlled studies are currently underway to better define the safety, efficacy, and long-term outcomes of CAR-T therapy across a spectrum of autoimmune diseases. The integration of novel technologies—ranging from artificial intelligence and organoid models to metabolic and combinatorial pharmacologic strategies—continues to expand the boundaries of what is possible in immune cell engineering. As our scientific understanding deepens and manufacturing platforms mature, CAR-T therapy is poised to move beyond proof-of-concept, offering the potential for lasting, treatment-free remission and fundamentally shifting the landscape of autoimmune diseases management.

Importantly, successful clinical translation will also depend on navigating evolving regulatory frameworks and addressing economic barriers. Regulatory agencies such as the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) are beginning to adapt oncology-derived CAR-T approval pathways to the distinct risk–benefit profiles of autoimmune indications—where long-term safety and the potential for cure must be weighed against acute toxicities in often younger, non-malignant populations. Parallel efforts to reduce costs are equally critical: innovations in manufacturing (e.g., point-of-care or automated platforms), the development of allogeneic “off-the-shelf” CAR-T products, and novel reimbursement models tied to durable remission could significantly enhance accessibility and scalability.

In summary, CAR-T cell therapy represents an ambitious and promising leap forward in the quest for curative treatments for autoimmune diseases. The coming years will be crucial in fully elucidating its potential, optimizing its safety and accessibility, and determining its ultimate place in clinical practice. If these challenges are met, CAR-T therapy may well inaugurate a new era in which the goal is not merely disease control, but true immunological cure.

Supplementary Information

Supplementary Material 1.

Acknowledgements

Not applicable.

Abbreviations

Anti-cyclic citrullinated peptide antibody

Antibody-dependent cellular cytotoxicity

Autoimmune diseases

Artificial intelligence

B-cell-activating factor

B-cell maturation antigen

Chimeric autoantibody receptors

Chimeric antigen receptor -T cell

Complement activation

Central nervous system

Cytokine release syndrome

Dendritic cell

Germinal centers

Glucose Transporters

Graft-versus-host disease

Immune effector cell-associated neurotoxicity syndrome

Interleukins

Induced pluripotent stem cells

Lipid nanoparticles

Macrophages

Major histocompatibility complex

Myeloid-derived suppressor cells

Multiple sclerosis

Nicotinamide adenine dinucleotide

Naïve B cells

Neutrophil extracellular traps

Natural killer cell

Rheumatoid arthritis

Single-chain variable fragment

Systemic lupus erythematosus

Systemic Sclerosis

Type 1 diabetes mellitus

T follicular helper

Toll-like receptor

Tumor microenvironment

Tumor necrosis factor receptor

T peripheral helper

Regulatory T cells

Author contributions

Xiaomei Chen, Kaifan Liu, and Bowen Liu analyzed the data and writed the manuscript, Shiyin Li, and Yulian Wang supervised the writing of the article, Xin Du and Jianyu Weng collected the clinic data; Peilong Lai, Kongming Wu, and Bing Song designed the research and provided critical reviews.

Funding

The author(s) declared that financial support was received for the research, authorship, and/or publication of this article. This research was supported by the National Natural Science Foundation of China (No. 82100228, 82270161, 82202978 and 82272794), Guangdong Basic and Applied Basic Research Foundation (No. 2024B1515020054), Guangzhou Basic and Applied Basic Research Foundation (No.2024A04J10011), Science and Technology Planning Project of Guangdong Province, China (No. 2023B1111050004), NSFC Incubation Project of Guangdong Provincial People’s Hospital (KY0120220024 and KY012021160), and High-level Hospital Construction Project of Guangdong Provincial People’s Hospital (DFJHBF202107).

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Xiaomei Chen, Kaifan Liu and Bowen Liu contributed equally to this work.

Contributor Information

Kongming Wu, Email: wukm_lab@163.com.

Peilong Lai, Email: lai_peilong@163.com.

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Associated Data

Supplementary Materials

Supplementary Material 1.

Data Availability Statement

No datasets were generated or analysed during the current study.