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CAR T cell therapies in multiple myeloma
which was associated with rapid removal of IL-6 from the culture supernatant. Furthermore, T-cell proliferation increased while the signaling and function of IL-6 was neutralized. T cells co-expressing mbaIL6 and anti-CD19 CAR, neutralized IL-6 derived from macrophages while maintaining antitumor activity in vitro and in a xenograft model.72 Thus, CAR-T incorporating the capacity to remove IL-6 may provide new strategies for preventing CRS. Another strategy is the incorporation of “suicide” switches, such as constructs containing a CAR and inducible caspase 9. Administration of a small molecule causing dimerization of inducible caspase 9 resulted in apoptosis and CAR-T-specific depletion.73
Reducing off-tumor on-target toxicity
In order to avoid adverse off-tumor effects, spatial and temporal activity of CAR-T need to be limited. Under this hypothesis, GPRC5D has been proposed as a novel target antigen, expressed on almost all CD138+ cells.74 Like BCMA, its expression is restricted to plasma cells, except for hair follicles. Preliminary results of anti-GPRC5D CAR-T showed potent anti-MM efficacy in vitro and in a mouse model, with the encouraging finding that these cells also effectively eradicated MM after treatment with anti-BCMA CAR-T. Most recently, it was demonstrated that simultaneous targeting of GPRC5D and BCMA could prevent relapse mediated by BCMA escape. Several multi- target constructs were compared and in BCMA-negative disease, dual-target (bicistronic) and pooled approaches exhibited the highest efficacy, whereas for GPRC5D/BCMA-expressing disease, the dual-target appeared to be more efficacious. Mechanistically, express- ing two CAR on one cell enhanced the strength of CAR- T/target interactions.
Reducing immunogenicity and simplifying structures
In order to reduce the immunogenicity of the CAR bind- ing domain, human or humanized scFv have been used more frequently in recent studies, instead of murine sequences.68,75 Furthermore, a reduction of immunogenici-
ty might be achieved by the incorporation of heavy-chain- only binding domains, which subsequently simplify the structure of the CAR antigen-binding domain without having a light-chain domain.76 In general, simplified struc- tures may facilitate better gene expression by transduced T cells.76,77 Moreover, limiting the size of expressed genes is important for the potential expression of >1 protein.78,79 A recent study demonstrated that CAR with antigen- recognition domains consisting of only a fully human heavy-chain variable domain (FHVH33) in addition to 4- 1BB and CD3ζ domains mediated comparable cytokine release, reduction in tumor burden, and degranulation in mice when compared to an identical CAR with a conven- tional scFv.76,80 Further investigations identified a crucial contribution of 4-1BB in reducing activation-induced cell death, enabling survival of T cells expressing FHVH33- CAR.76
Efficacy
Understanding antigen loss
Some relapses are either antigen-negative or antigen- low.80 One study in leukemia mouse models could dissect evidence for CAR promoting reversible antigen loss through a mechanism called trogocytosis.81 This mecha- nism defines an active process of rapid intercellular trans- fer of membrane fragments and related molecules. The specific target antigen is transferred to T cells resulting in decreased density on tumor cells, leading to declined T- cell activity by boosting fratricide T-cell killing and exhaustion.81 These cascades affected CAR constructs that included different costimulatory domains (CD28 or 4- 1BB), and the effect was dependent on antigen density. Thus, it was hypothesized that multi-target CAR-T could overcome these limitations.81
Multi-targeting
T-cells expressing single-chain bispecific CAR are able to prevent antigen escape.68,82 Moreover, CAR pools com- bining two single-input CAR-T products have been pro- posed (Figure 2). Pooling a humanized anti-CD19 and a
Table 4. Limitations and ways to improve CAR-T therapy in multiple myeloma. What may limit CAR-T therapy?
How to improve CAR-T therapy?
Anti-IL6 treatment and prevention
Safeguard designs incorporating drugs such as rituximab/cetuximab Tackling immunogenicity
Simplified CAR structures (e.g., heavy-chain-only binding domains)
Multi-targting therapy (dual-target, OR-target, CARpool) More accurate measurement of expansion/persistence “Suicide switches”
Combination of immunomodulatory modulation and CAR-T Senolytic CAR-T (?)
Address trogocytosis
Increase antigen density (e.g. γ-secretase-inhibition for anti-BCMA therapy)
Increase comparability and knowledge sharing of intensive care unit management and other care settings
Outcome prediction
Allogeneic CAR-T
Optimize supply chain models (e.g., intermediate players for cryopreservation)
Toxicity
Resistance
Management
Availability
On-target, on-tumor On-target, off-tumor
Impaired CAR-T expansion/persistence
Immunosuppression induced by BM microenvironment Antigen loss or downregulation
Suboptimal recognition and treatment of severe events
Lack of scale-up High costs
No stockpiling Time
BCMA: B-cell maturation antigen; IL6: interleukin-6; BM: bone marrow.
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