ABSTRACT Extracellular vesicles (EVs) are promising delivery vehicles capable of transporting therapeutic agents across biological barriers. However, native EVs primarily accumulate in liver, spleen and lungs, limiting targeted delivery to disease sites. To enhance their targeting efficiency for the plasma cell cancer multiple myeloma (MM), localized in the bone marrow (BM), we engineered HEK293‐derived EVs to display a nanobody (Nb) against the MM cell surface marker CS1. We confirmed enrichment of the Nb construct on engineered EVs and binding of α‐CS1 EVs to CS1. In vitro, we found enhanced α‐CS1 EV uptake by MM cell cultures. In vivo, we first compared the biodistribution of HEK293‐derived native EVs in healthy and MM‐bearing mice. Although native EVs reached the BM in both groups, MM‐bearing mice showed increased liver and lung accumulation together with reduced BM delivery. α‐mCS1 EV delivery to the BM of MM‐bearing mice was only slightly increased compared to native EVs, while off‐target accumulation also increased. At the cellular level, no changes in EV delivery to MM cells were detected. In conclusion, while CS1 targeting enhances in vitro EV uptake by MM cells, in vivo biodistribution remains suboptimal. Further optimization is needed to improve EV‐based drug delivery for MM.

Immunotherapy has emerged as a promising strategy for multiple myeloma (MM), yet relapse remains frequent due to the immunosuppressive bone marrow (BM) microenvironment, characterized by T cell dysfunction and accumulation of immunosuppressive myeloid cells. The co-stimulatory receptor 4-1BB (CD137, TNFRSF9) can enhance T and NK cell effector functions, but its therapeutic utility in MM is not well established. Tasquinimod (TQ), a clinical-stage S100A9 inhibitor, offers a complementary approach by limiting the recruitment and activity of suppressive myeloid cells. 4-1BB expression was assessed during disease progression in MM mice and in newly diagnosed MM patients using single-cell RNA sequencing and flow cytometry. Therapeutic potential was evaluated in 5TGM1 tumor-bearing mice treated with two 4-1BB agonists, LOB12.3 (IgG1κ) and 3H3 (IgG2a), using isotype controls. The lead agonist was subsequently combined with TQ to investigate dual targeting of the immunosuppressive tumor microenvironment. Tumor burden was quantified via BM and spleen plasmacytosis and serum M-protein levels. Immune modulation was analyzed using multi-parameter flow cytometry. Statistical significance was determined using the Mann–Whitney U test or one-way ANOVA (p < 0.05). 4-1BB expression progressively increased on T and NK cells during tumor development in mice. In primary MM patient BM samples, ex vivo 4-1BB stimulation with urelumab enhanced effector responses, increasing IFN-γ+ and Granzyme B+ CD3+ T cells, alongside trends toward increased CD56+ NK cells and elevated IFN-γ+ NK cell activity. In vivo, 4-1BB agonist treatment promoted expansion of T cell subsets, with clone-specific effects: the IgG2a clone 3H3 significantly reduced M-protein levels and BM plasmacytosis, whereas the IgG1 clone LOB12.3 induced NK cell depletion and demonstrated limited anti-tumor activity. Combining 3H3 with TQ provided superior anti-myeloma efficacy, reducing BM plasmacytosis from 62.5

INTRODUCTION:Multiple myeloma (MM) is a plasma cell malignancy characterized by frequent relapse and resistance to therapy. Caveolin-1 (Cav-1), a scaffolding protein that forms plasma membrane caveolae, has been demonstrated to regulate key processes including cell signaling, metabolism, autophagy, and interactions with the bone marrow microenvironment. AREAS COVERED:This review outlines Cav-1's role in MM progression and therapy resistance, including its effects on cell survival, adhesion, and communication with the bone marrow environment. Preclinical approaches to target Cav-1, such as small molecules, peptides, RNA-based methods, CRISPR, and tumor-specific delivery, are summarized, including combination with proteasome inhibitors. Challenges for clinical translation, such as the lack of selective inhibitors and possible toxicity, are also discussed. EXPERT OPINION:Cav-1 is a context-dependent therapeutic vulnerability in MM. Blocking Cav-1 can restore drug sensitivity, reduce protection from the bone marrow environment, and improve immune killing of tumor cells. Given the multifaceted nature of Cav-1 and its prevalence in normal tissues, the development of selective or tumor-targeted delivery mechanisms is imperative. New strategies, including inhibitors and nanoparticle delivery, combined with biomarker-guided patient selection, may offer safe and effective targeting of Cav-1 and support combination treatments for resistant MM.

T cell exhaustion and T cell senescence constitute distinct yet partially overlapping differentiation states that collectively constrain T cell functionality. T cell exhaustion arises under conditions of chronic antigen exposure and is characterised by a progressive, hierarchical loss of effector capacity, sustained expression of inhibitory receptors, and extensive transcriptional, epigenetic and metabolic reprogramming. By contrast, T cell senescence represents a more stable and terminal state, driven by replicative history, age-associated decline or stress-induced damage, and is defined by durable cell cycle arrest, altered differentiation, metabolic remodelling and acquisition of a pro-inflammatory secretory phenotype. In the context of cancer, dysfunctional T cells contribute to tumour progression, while also representing a major barrier to the success of T cell-based immune therapies, which strongly rely on the fitness, persistence and functional plasticity of T cells. Although substantial efforts have focused on overcoming exhaustion and optimising T cell manufacturing, senescence remains comparatively underexplored and presents unique therapeutic challenges due to its relative resistance to functional reprogramming. This review provides a comprehensive overview of T cell replenishment in homeostasis, followed by the molecular hallmarks and signalling pathways of T cell senescence and exhaustion. We discuss the current landscape of T cell-based immune therapies, including immune checkpoint blockade, T cell engagers and adoptive cell therapies, and explain how T cell dysfunction impacts their therapeutic outcomes. Finally, we highlight emerging strategies to prevent or overcome T cell dysfunction in adoptive cell therapy products.

Multiple myeloma (MM) is a hematological malignancy characterized by plasma cells residing in the bone marrow. Despite advancements in treatment, including proteasome inhibitors (PIs) such as bortezomib (Bz), drug resistance remains a major challenge. Metabolic reprogramming supports MM survival and drug resistance, with mitochondria emerging as promising therapeutic targets through their control of OXPHOS and mitochondrial reactive oxygen species (Mito-ROS). Using metabolic flux analyses, flow cytometry, and Western blot analysis, we identified pyruvate as a central metabolic intermediate, which not only enhances mitochondrial respiration and Mito-ROS production, but also the Integrated Stress Response (ISR) pathway. Conversely, metformin, an inhibitor of OXPHOS, was still able to activate the ISR pathway, but rather reduced Bz-induced cytotoxicity by decreasing both protein synthesis, and ROS production. Results were confirmed on primary murine and patient samples. Moreover, analysis of the CoMMpass study revealed that patients with prolonged progression-free survival under PI treatment showed enrichment in OXPHOS-related genes, highlighting the importance of mitochondrial metabolism in regulating MM responses to Bz. These data suggest that targeting pyruvate metabolism to increase ROS production could offer a strategy to enhance Bz activity in MM.

Adrenergic signaling regulates immune homeostasis through neuroendocrine pathways, but its role in hematologic malignancies remains poorly understood. Multiple myeloma (MM) originates in the bone marrow (BM), a highly innervated niche where neural, immune, and stromal signals are integrated. Whether adrenergic stimulation shapes BM immunity and influences myeloma progression is unclear. Using the 5T33MM mouse model, we investigated how adrenergic activation affects tumor growth and the BM immune landscape. Sustained adrenergic signaling was induced in vivo, while pharmacological β-adrenergic stimulation was used to assess effects on tumor-immune interactions. Immune composition and tumor burden were analyzed in BM and spleen, β2-adrenergic receptor expression was examined across murine and human immune populations, and ex vivo assays were performed using murine and patient-derived BM samples treated with adrenergic agonists alone or combined with relevant myeloma immunotherapies. Chronic restraint stress remodeled the BM immune compartment in MM-bearing mice, characterized by expansion of innate effector populations, including neutrophils and natural killer cells, and was associated with a significant reduction in BM tumor burden. These effects were compartment-restricted and not reflected in splenic tumor load or systemic disease markers. Pharmacological β-adrenergic stimulation reproduced these immune alterations and transiently delayed myeloma progression. In contrast, short-term β-adrenergic agonism ex vivo did not alter MM cell viability but consistently reduced the efficacy of MM immunotherapies. In summary, adrenergic signaling exerts context-dependent effects on myeloma progression, restraining tumor growth in vivo while impairing immunotherapy efficacy ex vivo, identifying a BM-specific neuroimmune axis relevant for cancer treatment.

Multiple myeloma (MM) develops in the hypoxic bone marrow (BM) microenvironment, which alters tumor behavior and immune responses. While hypoxia is known to directly suppress immune function, its effect on immunotherapy-relevant antigen expression and the MM secretome remains underexplored. Here, we investigated how hypoxia affects BCMA expression and BCMA-targeted CAR T cell responses. MM cells were cultured under normoxia (21

The role of the sympathetic nervous system (SNS) in cancer biology has gained increasing attention, and its ability to affect immunotherapy is starting to become clearer. Extensive evidence shows that neuro-onco-immune interactions significantly influence tumor progression and the effectiveness of cancer treatments. Blocking SNS signaling, primarily through β-adrenergic receptors, enhances immune cell functions, by increasing CD8+ T-cell activation and cytokine production, while reducing immunosuppressive cell populations. This review explores the relationship between SNS signaling and cancer immunotherapy, emphasizing how SNS activation affects the efficacy of various immunotherapies, including immune modulators, immune checkpoint inhibitors, oncolytic virus therapy, therapeutic vaccines, and CAR-T cell therapies. We summarize retrospective studies investigating the use of β-blockers during immunotherapy, suggesting potential benefits for treatment outcomes of blocking SNS signaling in the tumor microenvironment. We examine ongoing clinical trials that evaluate the use of beta-blockers with immune checkpoint inhibitors, which aim to improve patient outcomes. While translational and preclinical studies provide ample evidence for targeting SNS signaling in cancer immunotherapy, clinical studies are only beginning to emerge. Ultimately, this review underscores the need for further research to better understand how SNS signaling can be targeted to optimize immunotherapy, paving the way for more effective treatment strategies.

Abstract Background Accumulation of malignant plasma cells in the bone marrow causes lytic bone lesions in 80% of multiple myeloma patients. Frequently fracturing, they are challenging to treat surgically. Myeloma cells surviving treatment in the presumably protective environment of bone lesions impede their healing by continued impact on bone turnover and can explain regular progression of patients without detectable minimal residual disease (MRD). Locally applicable biomaterials could stabilize and foster healing of bone defects, simultaneously delivering anti-cancer compounds at systemically intolerable concentrations, overcoming drug resistance. Methods We developed silica-collagen xerogels (sicXer) and bortezomib-releasing silica-collagen xerogels (boXer) for local treatment of osteolytic bone disease and MRD. In vitro and in vivo (tissue sections) release of bortezomib was assessed by ultrahigh-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Material impact on bone formation was assessed in vitro regarding osteoclast/osteoblast numbers and activity. In vivo, drilling defects in a rat- and the 5T33-myeloma mouse model were treated by both materials and assessed by immunohistochemistry, UPLC-MS/MS, µCT, and ToF-SIMS. The material’s anti-myeloma activity was assessed using ten human myeloma cell lines (HMCLs) and eight primary myeloma cell samples including four patients refractory to systemic bortezomib treatment. Results sicXer and boXer show primary stability comparable to trabecular bone. Granule size and preparation method tailor degradation as indicated by release of the xerogel components (silica and collagen) and bortezomib into culture medium. In vitro, both materials reduce osteoclast activity and do not negatively interfere with osteoblast differentiation and function. The presumed resulting net bone formation with maintained basic remodeling properties was validated in vivo in a rat bone defect model, showing significantly enhanced bone formation for boXer compared to non-treated defects. Both materials induce myeloma cell apoptosis in all HMCLs and primary myeloma cell samples. In the 5T33-myeloma mouse model, both materials stabilized drilling defects and locally controlled malignant plasma cell growth. Conclusions The combination of stabilization of fracture-prone lesions, stimulation of bone healing, and anti-tumor effect suggest clinical testing of sicXer and boXer as part of a combined systemic/local treatment strategy in multiple myeloma and non-malignant diseases.

Abstract Introduction Immunotherapy has shown clinical promise in multiple myeloma (MM), but relapse remains common, largely due to the immunosuppressive bone marrow (BM) microenvironment, characterized by T cell exhaustion and the accumulation of myeloid-derived suppressor cells (MDSCs). Recent advances highlight the therapeutic promise of 4-1BB (CD137) agonist antibodies in enhancing T cell–mediated anti-tumor responses, though their efficacy in MM remains poorly defined. Investigating 4-1BB expression dynamics in a preclinical MM model informed our rationale for targeting this pathway. Tasquinimod (TasQ), a small-molecule immunomodulatory agent, currently evaluated in a phase Ib/IIa clinical trial in MM patients (NCT04405167), offers a complementary strategy. By inhibiting the S100A9 signaling pathway, TasQ interferes with the recruitment and function of MDSCs, resulting in a less suppressive tumor microenvironment (TME) which reestablishes the anti -tumor immunity. In this study, we assessed the therapeutic potential of two distinct 4-1BB agonist antibody clones in an immunocompetent preclinical MM model. Building on these findings, we further explored the combination of the most effective 4-1BB agonist with TasQ, aiming to overcome the immunosuppressive tumor microenvironment, and strengthen anti-myeloma immune activity. Methods 4-1BB expression was analyzed during disease progression in 5T33MM mice using single-cell RNA sequencing of spleen and BM, 2 myeloma-infiltrating organs. These findings were validated by flow cytometry in both 5T33MM and 5TGM1 models. Therapeutic potential was assessed by treating 5TGM1 tumor-bearing mice with two 4-1BB agonist clones—LOB12.3 (IgG1κ; n=7/group) and 3H3 (IgG2a; n=5/group), using clone-specific isotype controls. Beginning on day 3 post-tumor inoculation, mice received 100µg of antibody intraperitoneally twice weekly until end stage. In a follow-up experiment, the lead 4-1BB agonist was combined with TasQ, administered at 30mg/kg in drinking water, to evaluate the impact of dual targeting the immunosuppressive TME (n=11/group). Tumor burden was assessed by determining the percentage of plasmacytosis in BM and spleen through cytospin stainings, together with M-protein measurement by serum electrophoresis. Immunomodulating effects were investigated using multi-parameter flow cytometry. Statistical significance was determined using the Mann–Whitney U test or one-way ANOVA, with p<0.05 considered significant. Results 4-1BB was predominantly detected on T cells and natural killer (NK) cells, with its expression further increasing as the disease progressed. Treatment of 5TGM1 mice with 4-1BB agonists significantly increased the percentage of CD4+ and CD8+ T cells in the BM and spleen. Interestingly, treatment with clone LOB12.3 resulted in a significant reduction in NK cell percentages in both the BM and spleen, while clone 3H3 selectively reduced splenic NK cells. Therapeutically, clone 3H3 significantly decreased M-protein levels and BM plasmacytosis (p<0.01), while no significant effects were observed for clone LOB12.3. Treatment with the IgG2a-formatted 4-1BB agonist combined with TasQ led to a significant reduction in M-protein levels and BM plasmacytosis (p<0001). Plasmacytosis was 62.50% in the isotype control group, decreased to 36.18% with 4-1BB agonist, 37.64% with TasQ, and further reduced to 14.09% in the combination therapy group, highlighting the enhanced efficacy of dual treatment. These effects were mediated by increased granzyme B–mediated T and NK-cell activation and enhanced differentiation of effector T cells (CD44⁺CD62L⁻). While this therapy did not alter the frequency of dendritic cells (DCs) in the BM, it enhanced their maturation, as evidenced by increased CD86 expression—particularly on type 1 and type 2 conventional DC subsets. Conclusion In conclusion, our findings demonstrate that 4-1BB activation can enhance anti-tumor immunity in MM; however, the therapeutic efficacy is dependent on the specific agonist used. Notably, the IgG2a-formatted 4-1BB agonist showed superior activity, underscoring the importance of isotype selection in achieving optimal immunotherapeutic outcomes. Moreover, co-administration with TasQ enhanced therapeutic efficacy, supporting the potential benefit of a combinatorial approach. Further investigation is warranted to elucidate the mechanisms driving these responses and to refine 4-1BB-targeted strategies for effective clinical translation.

Objective Ovarian cancer (OC) is the deadliest gynecological malignancy, with patients experiencing late diagnosis and high recurrence. AXL is highly expressed in OC and correlates with worse prognosis. We aim to evaluate the use of AXL-targeting nanobodies as a novel therapeutic modality to inhibit the AXL-GAS6 axis in OC. Methods ID8-fLuc cells were in vivo passaged three times to obtain the fast-progressing P3 ID8 Thy1.1 cell line. The expression of AXL in ID8 lines was assessed by flow cytometry and immunofluorescence, while serum GAS6 levels in tumor-bearing and naïve mice were measured by ELISA. Using in silico analyses, we correlated AXL and GAS6 expression with OC patient outcomes and assessed AXL expression in different tissues and patient cohorts. Orthotopic ovarian tumors were processed to a single cell suspension and treated in vitro with the nanobody (AXL-Fc), a PARP inhibitor (Olaparib), or controls (PBS or DMSO) for 24 h or 72 h, followed by flow cytometry to assess stages of cell death and cancer cell proliferation. Results In vivo passaging of ID8-fLuc cells resulted in a faster-progressing P3 ID8 Thy1.1 cell line that recapitulates stages I-III observed in OC patients. In OC patients, AXL and GAS6 genes are highly expressed in primary and metastatic tumors, and enriched in platinum-resistant patients. AXL-Fc induces necrosis in OC cells from orthotopic ovarian tumors. AXL-Fc synergizes with Olaparib, resulting in decreased OC cell proliferation. Conclusions Leveraging the synergistic effects of AXL-Fc with Olaparib, we propose a new AXL-targeting treatment approach for OC that warrants further investigation.

Despite significant advancements, multiple myeloma (MM) remains incurable, largely due to drug resistance. Our previous research has demonstrated that proline metabolism plays a role in MM progression and that inhibiting PYCR1, the final enzyme in proline synthesis, enhances bortezomib sensitivity in MM cells. Given the high expression of PYCR1 in bone marrow stromal cells (BMSCs), we sought to investigate the effects of PYCR1 inhibition in BMSCs and its indirect influence on MM cell metabolism and viability. Culturing MM cells in conditioned medium (CM) of PYCR1-silenced BMSC significantly impaired oxidative phosphorylation and sensitised MM cells to bortezomib. Analysis of the CM secretome revealed a reduction in activin A release. Proline and activin A supplementation were able to counteract MM sensitivity to bortezomib. Combination therapy of the PYCR1 inhibitor pargyline and bortezomib reduced tumour load in a 3D model and reduced serum activin A levels in 5TGM1-bearing mice. This study demonstrates the contribution of stromal cell metabolism to MM progression. Inhibiting PYCR1 in BMSCs leads to less activin A release, limits oxidative phosphorylation in MM cells and enhances bortezomib efficacy.

BACKGROUND:The plasma cell malignancy multiple myeloma (MM) remains incurable due to the inevitable development of drug resistance (DR). Epigenetic modifiers are frequently mutated or deregulated in MM patients, contributing to MM progression and relapse. Overexpression of the de novo DNA methyltransferase 3B (DNMT3B) in MM has been reported, correlating with poor prognosis. However, its exact role in MM cell biology and relapse remains elusive. METHODS:To evaluate the basal expression and prognostic value of DNMT3B mRNA in terms of overall survival the publicly available gene expression profiling datasets GSE2658, GSE9782, GSE4581, E-MTAB-372, E-TABM-1088 and E-TABM-937 were used. Both the DNMT3B selective inhibitor Nanaomycin A and genetic knockdown using a doxycycline inducible shRNA against DNMT3B were used to target DNMT3B. Viability and apoptosis were assessed using respectively a CellTiter-Glo assay and AnnexinV/7AAD stainings. Cell proliferation was measured by BrdU incorporation and cell cycle analysis, while the clonogenic capacity was evaluated by a colony formation assay. Finally, RNA-seq was performed upon genetic knockdown. RESULTS:Here, we show that DNMT3B is significantly increased in the relapsed setting and high DNMT3B levels are strongly correlating with disease progression and high-risk disease, irrespective of the treatment. Targeting DNMT3B using either genetic inhibition or the selective inhibitor Nanaomycin A strongly impaired MM cell growth, survival and clonogenicity. Moreover, Nanaomycin A reduced viability of primary MM cells from newly diagnosed and relapsed patients. Mechanistic studies revealed that DNMT3B inhibition mainly affects cell cycle and stemness-related transcriptional programs. Notably, DNMT3B depletion affected the stability of the master cell cycle regulator MYC, thereby reducing c-MYC levels and cell viability both in parental and c-MYC overexpressing cells. Finally, Nanaomycin A (re)sensitized MM cells to bortezomib, melphalan and anti-CD38 monoclonal antibodies (daratumumab, isatuximab). CONCLUSION:Collectively, our findings uncover DNMT3B as a targetable vulnerability in high-risk patients with high DNMT3B/MYC levels.

Multiple myeloma (MM) remains incurable due to the development of drug resistance. We previously showed that communication between bone marrow stromal cells (BMSCs) and MM cells supports MM growth and triggers therapy resistance. This communication occurs through a plethora of mechanisms, including the release of cytokines and small extracellular vesicles (sEVs). The PDZ protein syntenin is a master regulator of intercellular communication, in particular via sEVs. In this study, we aimed to explore whether targeting syntenin, by genetic alteration or pharmacological inhibition, can disrupt BMSC-MM crosstalk, thereby rendering the MM cells more sensitive to therapy. We found that syntenin (SDCBP) is highly expressed in inflammatory BMSC of MM patients and that its expression in BM aspirates correlates with poor patient survival. Using in vitro models, we established that knockout of syntenin in BMSC alters their secretome and abolishes BMSC-induced bortezomib resistance of MM cells via regulation of STAT3, MAPK, and AKT-mTOR pathways. Pharmacological inhibition of syntenin decreases syntenin and IL-6 sorting into BMSC sEVs and enhances bortezomib-induced MM cell death. Finally, we validated the therapeutic added value of syntenin inhibition in combination with bortezomib in vivo, using the 5TGM1 MM mouse model. In conclusion, our findings show that syntenin supports the secretion of pro-tumoral factors by BMSCs and qualifies as a possible novel therapeutic target in MM.

BACKGROUND:Invariant natural killer T (iNKT) cells and CD8+ T cells are key in the immune response against multiple myeloma (MM), a largely incurable blood cancer. Immunization is a promising strategy to activate these T cell populations. To our knowledge, immunization with messenger RNA (mRNA) and the iNKT agonist, α-galactosylceramide (αGC), has not been studied in MM, as knowledge on clinically relevant antigens in preclinical MM models is lacking. METHODS:Microarray data and immunopeptidomics (imPep) were used to identify candidate antigens for immunization in 5TMM models. Galsomes, lipid nanoparticles containing antigen mRNA and αGC were used to immunize 5T33MM-bearing mice. This treatment was combined with a CD40 agonist. Tumor burden and activation of iNKT cells and CD8+ T cells were studied using M-protein electrophoresis, flow cytometry and ELISA. RESULTS:RNA transcripts revealed survivin as a candidate antigen. Prime-boost Galsomes therapy targeting survivin significantly reduced M-protein levels despite low survivin-specific T cell responses. Further analysis showed potential T cell fratricide. ImPep revealed HSP60, Idiotype, PICALM and EF1A1 as candidate antigens. Prime-boost therapy with Galsomes targeting these antigens reduced MM growth significantly when combined with a CD40 agonist, coinciding with significantly improved antigen presentation, costimulation and cytotoxicity of iNKT cells and CD8+ T cells. CONCLUSION:These findings highlight the potential of Galsomes, an mRNA vaccine designed to activate CD8+ T cells and iNKT cells, for MM therapy, and emphasize the importance of combinatorial approaches, addressing immune anergy for effective MM immunotherapies.

BACKGROUND:CAR T cell therapy targeting BCMA has shown remarkable efficacy in multiple myeloma (MM), but relapses occur due to T cell exhaustion and the emergence of BCMA-negative subpopulations. Novel targets are needed to overcome antigen escape. METHODS:B7-H3 (CD276) expression was assessed on primary MM patient samples. We engineered nanobody-based CAR T cells (nanoCARs) targeting B7-H3 and evaluated their cytotoxicity and cytokine production in vitro, including against patient-derived myeloma cells. Anti-tumor activity was tested in two different MM xenograft models. Dual CAR T cells (BCMA/B7-H3) and CARpooling (mix of BCMA and B7-H3 CAR T cells) were also tested for efficacy in antigen escape models. RESULTS:B7-H3 expression was detected on plasma cells in 60% of MM patients. B7-H3 nanoCAR T cells exhibited strong antigen-specific cytotoxicity and effector cytokine secretion, including against primary MM cells. In vivo, they reduced tumor burden and improved survival. Dual (BCMA/B7-H3) CAR T cells and CARpooling effectively eliminated heterogeneous tumor populations with mutually exclusive BCMA or B7-H3 expression. These findings show that BCMA/B7-H3 targeting may be a strategy to overcome antigen escape mechanisms. CONCLUSION:B7-H3 is a promising immunotherapy target in MM. B7-H3-specific and dual-targeting nanoCAR T cells could offer a strategy to prevent antigen escape and improve treatment durability.

AbstractUntil recently, treatment options for patients diagnosed with Acute Myeloid Leukemia (AML) were limited and predominantly relied on various combinations, dosages, or schedules of traditional chemotherapeutic agents. Patients with advanced age, relapsed/refractory disease or comorbidities were often left without effective treatment options. Novel advances in the understanding of leukemogenesis at the molecular and genetic levels, alongside recent progress in drug development, have resulted in the emergence of novel therapeutic agents and strategies for AML patients. Among these innovations, the receptor tyrosine kinase AXL has been established as a promising therapeutic target for AML. AXL is a key regulator of several cellular functions, including epithelial-to-mesenchymal transition in tumor cells, immune regulation, apoptosis, angiogenesis and the development of chemoresistance. Clinical studies of AXL inhibitors, as single agents and in combination therapy, have demonstrated promising efficacy in treating AML. Additionally, novel AXL-targeted therapies, such as AXL-specific antibodies or antibody fragments, present potential solutions to overcome the limitations associated with traditional small-molecule AXL inhibitors or multikinase inhibitors. This review provides a comprehensive overview of the structure and biological functions of AXL under normal physiological conditions, including its role in immune regulation. We also summarize AXL’s involvement in cancer, with a specific emphasis on its role in the pathogenesis of AML, its contribution to immune evasion and drug resistance. Moreover, we discuss the AXL inhibitors currently undergoing (pre)clinical evaluation for the treatment of AML.

CAR T cells are widely applied for relapsed hematological cancer patients. With six approved cell therapies, for Multiple Myeloma and other B-cell malignancies, new insights emerge. Profound evidence shows that patients who fail CAR T-cell therapy have, aside from antigen escape, a more glycolytic and weakened metabolism in their CAR T cells, accompanied by a short lifespan. Recent advances show that CAR T cells can be metabolically engineered towards oxidative phosphorylation, which increases their longevity via epigenetic and phenotypical changes. In this review we elucidate various strategies to rewire their metabolism, including the design of the CAR construct, co-stimulus choice, genetic modifications of metabolic genes, and pharmacological interventions. We discuss their potential to enhance CAR T-cell functioning and persistence through memory imprinting, thereby improving outcomes. Furthermore, we link the pharmacological treatments with their anti-cancer properties in hematological malignancies to ultimately suggest novel combination strategies.