ABSTRACT:Circulating tumor cells (CTC) represent a high-risk biomarker in newly diagnosed multiple myeloma (NDMM); however, their prognostic value among transplant-eligible (TE) patients receiving daratumumab/bortezomib/lenalidomide/dexamethasone (D-VRd) remains unknown. In this study, we analyzed CTC in the phase 3 PERSEUS/EMN017 trial. TE-NDMM patients were randomized (1:1) to D-VRd with daratumumab/lenalidomide maintenance (D-VRd group) or bortezomib/lenalidomide/dexamethasone (VRd) with lenalidomide maintenance (VRd group), both with transplant. A subset of 451 of 709 patients from PERSEUS (D-VRd, 231/355; VRd, 220/354) had screening blood samples collected for CTC analysis by flow cytometry. CTC were detected in 370 patients (82%; median limit of detection, 0.0004%). CTC were prognostic of progression-free survival (PFS), independent of other factors, as a continuous (hazard ratio [HR], 1.36 [95% confidence interval (CI), 1.15-1.60]; P< .001) and categorical variable (≥0.175% CTC-high, optimal threshold). D-VRd improved PFS vs VRd in CTC-low patients (4-year rates: 88% vs 74%; HR, 0.42 [95% CI, 0.25-0.70]; P = .0013). Regardless of study treatment, minimal residual disease (MRD)-negativity rates were lower in CTC-high vs CTC-low patients (10-5: 52.2% vs 66.2%; 10-6: 34.8% vs 52.4%). D-VRd significantly increased MRD-negativity rates vs VRd among CTC-high (10-5: 69.4% vs 33.3%; 10-6: 47.2% vs 21.2%; both P< .05) and CTC-low patients (10-5: 74.4% vs 57.8%; 10-6: 65.6% vs 38.5%; both P< .001), with similar observations for sustained MRD-negativity. CTC levels are an independent prognostic factor in TE-NDMM treated with standard-of-care frontline quadruplet. D-VRd improved and sustained MRD-negativity rates in CTC-high and CTC-low, and improved PFS for CTC-low with a positive trend in CTC-high patients. This trial was registered at www.clinicaltrials.gov as #NCT03710603.

BACKGROUND:Proteasome inhibitors (PIs) are indispensable for the treatment of multiple myeloma (MM), the second most common hematologic malignancy. Although primary resistance to PIs is rare, most patients eventually relapse and develop acquired resistance, with underlying mechanisms that remain incompletely understood and appear to be drug-specific. In the case of bortezomib, resistance is often associated with PSMB5 mutations. In contrast, resistance to carfilzomib (CFZ) is mediated by overexpression of the drug efflux transporter ABCB1. However, the regulatory mechanisms driving ABCB1 upregulation in CFZ-resistant MM remain unclear. METHODS:An integrative multi-omics analysis was conducted using paired samples from a CFZ-sensitive and -resistant MM patient, alongside resistant cell line models. Whole-genome sequencing (WGS), whole-genome bisulfite sequencing (WGBS), and RNA sequencing (RNA-seq) were used to assess the genotype (structural variants, single nucleotide variants, and copy number variations), methylation status, and the expression of the ABCB1 locus. ABCB1 promoter methylation levels and expression levels in an independent MM subcohort were analyzed to determine clinical relevance. Functional validation was performed using dual-luciferase reporter assays, DNMT1 knockdown, and treatment with DNA methyltransferase inhibitors (DNMTis) to evaluate methylation-dependent regulation of ABCB1 expression. RESULTS:Significant hypomethylation of the ABCB1 downstream promoter region was identified (GH07J087598) in a CFZ-resistant patient sample, which correlated with elevated ABCB1 expression. Consistent with the paired CFZ-resistant case, the independent MM subcohort showed a significant inverse association between ABCB1 promoter methylation and ABCB1 expression. These findings align with results obtained from CFZ-resistant MM cell line models, which demonstrated reduced promoter methylation and elevated ABCB1 expression compared to their wild-type counterparts. Furthermore, treatment with DNA methyltransferase inhibitors as well as DNMT1 knockdown enhanced ABCB1 expression while demethylating the promoter, thereby validating the functional significance of promoter hypomethylation in ABCB1 overexpression. CONCLUSIONS:Our findings highlight ABCB1 promoter hypomethylation as a potential epigenetic driver of CFZ resistance in MM. These results underscore the clinical relevance of epigenetic regulation in drug resistance and the potential of targeting DNA methylation as a therapeutic strategy to overcome resistance in MM.

Chimeric antigen receptor (CAR) T cells exhibit high response rates in B-cell malignancies, but most patients eventually relapse. A key mechanism of treatment failure is the loss or downregulation of tumor antigen expression, yet strategies to modulate cell surface levels of CAR T-cell targets remain largely unexplored. Here, we identify B-cell maturation antigen (BCMA), a central CAR T-cell target in multiple myeloma (MM), as a highly shortlived protein that undergoes K48-linked polyubiquitylation at the plasma membrane, leading to its p97-dependent degradation via the ubiquitin-proteasome system (UPS). This previously unprecedented mechanism of plasma membrane protein regulation enables significant enhancement of BCMA expression via proteasome inhibitors (PIs). The clinically approved PI carfilzomib (CFZ) significantly enhances the efficacy of BCMAdirected CAR T cells against both PI-sensitive and-refractory MM cells in vitro and in vivo. Notably, CFZ treatment of 10 patients with BMCA CAR T-cell therapy relapse, under the CFZ after BCMA CAR T-cell (CarCAR) protocol, resulted in increased BCMA expression in all patients. However, clinical responses were observed only in those with residual and/or expanding CAR T cells, suggesting restored CAR T-cell function. These findings provide a rationale for the use of CFZ treatment in relapsed or refractory MM after BCMA CAR T-cell therapy, advocate for future trials combining CFZ with BCMA CAR T cells, and provide a framework for exploring UPS-dependent degradation of other immunotherapy antigens.

Abstract Background: Overexpression of NAD(P)H:quinone oxidoreductase 1 (NQO1) has been linked to poor prognosis and therapy resistance in solid tumors, but its role in multiple myeloma (MM) remains unclear. Combination therapies including proteasome inhibitors (PIs), immunomodulatory drugs (IMiDs), and/or immunotherapies have improved outcomes in MM, yet most patients eventually develop resistant and relapse. We recently identified NQO1 overexpression across PI-resistant MM models and that high NQO1 expression is associated with inferior outcome in PI treated MM patients. Hence, we here extend these findings in an independent MM cohort (n = 93) from our institution and in NQO1 upregulated MM cell line models to examine how NQO1 affects MM immunotherapy targets and sensitivity to standard treatment regimens. Methods: RNA-seq was performed on bone marrow-derived CD138+ cells from 24 newly diagnosed MM (NDMM) and 69 relapsed/refractory MM (RRMM) patients, as well as MM cell lines. Surface density of immunotherapy targets was quantified via direct stochastic Optical Reconstruction Microscopy (dSTORM). Sensitivity to PIs and IMiDs was assessed using AlamarBlue assays. Results: In the MM RNA-seq cohort, NQO1 expression was significantly higher in RRMM compared with NDMM (p = 0.0002), supporting an association of NQO1 with treatment-exposed, advanced disease. Furthermore, overexpression of NQO1 in MM cell lines led to an increased IC50 for bortezomib and carfilzomib, whereas IMiDs sensitivity remained intact. No upregulation of PI resistance-related genes was identified by RNA-seq. To test whether NQO1-dependent mechanisms are causative for PI resistance, NQO1-high MM cells were treated with NQO1 inhibitor ES936, which restored the sensitivity to both PIs to the wild type levels. In subsequent dSTORM analysis, no differences were observed in BCMA, SLAMF7, or GPRC5D, whereas CD38 surface density was significantly reduced in both NQO1-High myeloma cell lines. Consistent with the expression data, NQO1-overexpressing cells showed reduced sensitivity to CD38-directed therapies (daratumumab (Dara) and isatuximab (Isa). Notably, NQO1 inhibition with ES936 restored CD38 surface expression and rescued Dara and Isa responses, supporting that NQO1 is directly involved in CD38 downregulation and resistance to anti-CD38 antibodies. Of note, RNA-seq comparison of NQO1-high vs. WT MM models did not reveal changes in CD38 mRNA levels, pointing toward a post-translational mechanism, such as altered protein stability or trafficking. Conclusions: NQO1 is upregulated in RRMM and functionally induces resistance to PIs and CD38-directed therapies which are widely used in current MM treatment. Inhibition of NQO1 with ES936 rescues CD38 expression, and restores PI sensitivity and anti-CD38 responses, supporting NQO1 as a promising target in treatment-exposed or relapsed MM. Citation Format: Seungbin Han, Christina Verbruggen, Lenka Besse, Shilpa Kurian, Silvia Nerreter, Umair Munawar, Marietta Truger, Elena Gerhard-Hartmann, Andrej Besse, Ann-Sophie Hainold, Cornelia Vogt, Emma Besant, Nina Rein, Max Köppel, Xiang Zhou, Claudia Haferlach, Andreas Rosenwald, Christoph Driessen, Ondrej Slaby, Hermann Einsele, Leo Rasche, Markus Sauer, Johannes Waldschmidt, K. Martin Kortüm. NQO1 as a target to overcome therapy resistance in multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3912.

The study investigated the effectiveness of dysgeusia-specific acupuncture combined with self-acupressure compared to supportive acupuncture and self-acupressure for chemotherapy-induced dysgeusia in cancer patients with cancer-related fatigue (CRF) and dysgeusia. This multicenter, randomized, controlled, two-armed parallel-group, single-blind trial included cancer patients with CRF and dysgeusia during chemotherapy. All participants received CRF treatment consisting of eight acupuncture sessions over 8 weeks and had access to an eLearning course for self-acupressure. Patients in the intervention group received additional dysgeusia-specific acupuncture treatment and an eLearning with instructions to acupressure points, whereas the control group received an intervention focused solely on addressing CRF. The main outcome was perceived dysgeusia over 8 weeks, measured weekly after each acupuncture session on a numeric rating scale from 1 to 10. Other outcomes included taste and smell tests, and assessments of perceived dysgeusia, fatigue, distress, and quality of life. Due to a small sample size, results can be only judged exploratory. The study was approved by the Cantonal Ethics Committee (CEC) (Kanton Zürich Kantonale Ethikkommission) (approval no. KEK-ZH-Nr. 2020—01900). Since we included only twenty-one patients (11 intervention, 10 control, 90

We performed a comprehensive review with meta- and network meta-analyzes of maintenance-therapy studies. Lenalidomide, proteasome inhibitors, and CD38 antibodies improved progression-free survival (PFS). However, overall survival (OS) benefit appeared only with lenalidomide in transplant-eligible (TE) patients, while CD38-directed therapy showed a trend toward improved OS not seen with proteasome inhibitors. The network meta-analysis ranked regimens against observation: daratumumab-lenalidomide (DR) yielded the greatest PFS benefit, followed by carfilzomib-lenalidomide-dexamethasone (KRd), KR in TE, and DRd in transplant-ineligible patients. Significant OS prolongation in TE patients occurred with KR, DR, and lenalidomide alone.

Proteasome inhibitors (PIs) are frontline therapies for multiple myeloma (MM). Although MM patients initially respond to PIs, resistance frequently emerges. While all PIs nominally target the same proteasomal catalytic subunit (PSMB5), the extent to which resistance mechanisms are the same or different among different PIs or between patients is poorly understood. To address this, we performed proteome and phosphoproteome profiling of 12 MM cell line models, comprising four parental lines (AMO-1, ARH77, L363, and RPMI8226) paired with lines that acquired resistance to bortezomib (BTZ) or carfilzomib (CFZ). Over 7,000 proteins and up to 10,000 phosphopeptides were identified per cell line, enabling a comprehensive comparative analysis of shared and cell line-specific resistance signatures at the protein level. Data analysis revealed surprisingly few changes in the phosphoproteome but substantial reprogramming of the proteome in most models. Beyond known adaptations such as the overexpression of the PI target PSMB5 and the drug efflux transporter ABCB1, we identified the oxidoreductases NQO1 and NQO2 as significantly upregulated proteins under chronic proteotoxic stress across several models. Pharmacological follow up in PI resistant AMO-1 cells showed that NQO2 inhibition by imatinib fully restored CFZ sensitivity, validating NQO2 as a contributor to resistance formation in this model system.

Background:Bone-marrow minimal residual disease (MRD) and positron emission tomography/computed tomography (PET-CT) are central to response assessment in multiple myeloma (MM); however, their agreement, discordant patterns, and how their joint results relate to progression-free survival have not been systematically quantified. Methods:We conducted a systematic review and random-effects meta-analysis, and searched PubMed and Cochrane CENTRAL from Jan 2015 to Sept 30, 2025, with an updated search to Jan 31, 2026. We included studies reporting paired MRD and PET-CT assessments, and, when available, progression-free survival (PFS), in patients with MM. Study-level 2 × 2 MRD/PET-CT tables (each test positive or negative) were abstracted or reconstructed from published counts or percentages. Primary outcomes were (1) cross-modality agreement (observed agreement and Cohen's κ) and directional discordance (log-odds of MRD-/PET-CT+ vs MRD+/PET-CT-), and (2) the prognostic effect of dual negativity (MRD-/PET-CT- vs all other combinations) on PFS. We used random-effects models (REML with Hartung-Knapp adjustment) to estimate pooled hazard ratios for PFS. Sensitivity analyses were informed by QUADAS-2 (Quality Assessment of Diagnostic Accuracy Studies 2) and QUIPS (Quality in Prognosis Studies) risk-of-bias assessments. This study was registered with Open Science Framework (OSF; DOI 10.17605/OSF.IO/3CH9E). Findings:Ten cohorts contributed 1138 paired MRD/PET-CT assessments. The joint 2 × 2 distribution was: (a) MRD-/PET-CT+ 145 (12.7%), (b) MRD-/PET-CT- 499 (43.8%), (c) MRD+/PET-CT- 310 (27.2%), and (d) MRD+/PET-CT+ 184 (16.2%). Observed agreement was 60.0%, and pooled κ was 0.14 (95% CI 0.03-0.25; I2 57%). Discordant results predominantly reflected MRD+/PET-CT- rather than MRD-/PET-CT+ (primary pooled log-odds -0.83, 95% CI -1.87 to 0.21; I2 93%; k = 10). In five studies reporting PFS by joint MRD/PET-CT status, dual negativity was associated with substantially longer PFS than all other categories (pooled HR 0.34 (95% CI 0.22-0.51); τ2 0.03; I2 27%; k = 5). Interpretation:MRD and PET-CT interrogate distinct but complementary disease compartments, yielding low statistical concordance yet strong joint prognostic value. Dual MRD/PET-CT negativity reproducibly identifies a low-risk subgroup, whereas discordant patterns capture biologically heterogeneous residual disease. Prospective myeloma trials should prespecify all four MRD/PET-CT categories, align assessment timing and thresholds, and routinely report concordance and discordance to enable response-adapted treatment. Funding:None.

Background: The AML60+ score has been proposed for risk stratification in intensively treated elderly patients with acute myeloid leukemia (AML) or high-risk myelodysplastic neoplasms (MDS). Its prognostic impact in patients treated with hypomethylating agents (HMA) is unknown. Methods: Patients ≥ 60 years of age diagnosed with AML or MDS/AML according to ICC2022 were eligible for this retrospective and multicenter chart review if they had received at least one cycle of HMA-based treatment. Results: A cohort of 142 patients was analyzed. During follow-up (median 8 months), 114 patients died. The molecular Prognostic Score (mPRS) was available for 121 patients, the European Leukemia Net (ELN) 2022 classification for 117 patients, and the AML60+ for 105 patients. According to AML60+, 33 patients (31.4%) were classified as very poor risk, 36 (34.3%) as poor risk, and 34 (32.4%) as intermediate risk. Two patients (1.9%) were classified as favorable. Median overall survival (OS) was 21.7 months (mo) for the combined intermediate/favorable group, 7 mo for the poor risk group and 3 mo for the very poor risk group (p < 0.0001). Cox regression analysis (reference category: very poor) showed a significantly lower risk of death for both intermediate/favorable risk patients (HR 0.17, 95% CI 0.10–0.31, p < 0.001) and poor risk patients (HR 0.47, 95% CI 0.28–0.78, p = 0.004). The concordance score was 0.67 for AML60+, 0.60 for mPRS, and 0.58 for ELN2022. Conclusions: The AML60+ may represent a useful prognostic tool for elderly AML patients treated with HMA-based therapies. In particular, it could help to identify a group with a relatively favorable prognosis that is not clearly identified by the ELN2022 or the mPRS risk classification. However, analyses of larger cohorts are necessary to confirm our findings.

Multiple myeloma is a malignant disease characterised by the clonal proliferation of plasma cells. Since the last update of the Swiss recommendations for the diagnosis and treatment of multiple myeloma in 2019, the therapeutic landscape has evolved significantly, with the development of new monoclonal antibodies, novel combination therapies, and the introduction of T-cell-redirecting treatments such as bispecific antibodies and CAR T-cell therapy. This article summarises the current diagnostic procedures and therapeutic recommendations in Switzerland.

Introduction: Proteasome inhibition (PI) remains a cornerstone of multiple myeloma (MM) therapy. Despite decades of clinical use, the molecular mechanisms underlying the development of PI resistance remain only partially understood. In patients, epigenetic repression of PSMD5 and overexpression of the drug efflux transporter ABCB1 were found to induce PI resistance. However, these mechanisms account only for a subset of patients and appear to be drug-specific. Using proteomic profiling,we recently identified upregulated NAD(P)H:quinone oxidoreductase 1 (NQO1) in bortezomib (BTZ)- as well as in carfilzomib (CFZ)-resistant MM cell line models. NQO1 is a cytoprotective oxidoreductase known to prevent redox cycling and the formation of reactive oxygen species (ROS) by catalyzing two-electron reductions of quinones. While NQO1 overexpression has been associated with chemoresistance in solid tumors, its role in hematologic malignancies, especially multiple myeloma, is largely unexplored. Here, we provide a comprehensive functional and clinical evaluation of NQO1 as a novel mediator of PI resistance, providing a potential therapeutic target in MM to overcome PI resistance. Methods: Patients were first stratified according to their NQO1 expression levels using the CoMMpass dataset (IA22): the top quartile (Q4) was defined as NQO1-high, and the lower three quartiles (Q1–Q3) as NQO1-low. Progression-free survival (PFS) was then compared between these groups, further stratified by whether first-line treatment regimens contained PIs or were based on non-PI therapies. NQO1-overexpressing MM cell lines (AMO1-NQO1 and RPMI8226-NQO1) were generated using the Sleeping Beauty System, and their drug sensitivity was assessed using the AlamarBlue assay following treatment with PIs (BTZ, CFZ) and immunomodulatory drugs (lenalidomide, pomalidomide). To evaluate whether PI resistance was NQO1-dependent and reversible, cells were co-treated with the irreversible NQO1 inhibitor ES936, followed by reassessment of PI sensitivity. Results: In the CoMMpass dataset (IA22), high NQO1 expression correlated significantly with inferior PFS in patients (n=544) receiving PI-containing first-line regimens (26.2 vs. 40.6 months, p = 0.0004; HR 1.49, 95% CI: 1.17–1.91). This association remained significant when the analysis was restricted to patients treated with PI-containing but IMiD-sparing regimens (n=157; 16.4 vs. 29.5 months, p = 0.0185). In contrast, no significant correlation was observed in patients receiving non-PI-based regimens (n = 39; 40.4 vs. 32.1 months; p = 0.8093), suggesting that elevated NQO1 levels may confer PI-specific resistance and adversely impact survival following first-line therapy. To functionally validate these clinical observations, we overexpressed NQO1 in AMO1 and RPMI8226 MM cell lines and assessed their drug sensitivity. Overexpression of NQO1 decreased sensitivity to both BTZ and CFZ while increasing IC₅₀ values by ~2.3× and ~1.3× in AMO1 and ~1.5× and ~1.2× in RPMI8226, respectively. Consistent with our clinical observations, IMiD sensitivity remained unaffected in our models. To validate whether acquired PI resistance was NQO1-dependent and reversible, cells were treated with the irreversible NQO1 inhibitor ES936. Co-treatment reduced the CFZ IC₅₀ by ~28% in AMO1 and by ~38% in RPMI8226, restoring PI sensitivity. As ABCB1 overexpression is a known driver of CFZ resistance, we next investigated whether combined inhibition of NQO1 and ABCB1 could further enhance drug sensitivity. In CFZ-resistant AMO1 cells, treatment with the ABCB1 inhibitor tariquidar decreased the CFZ IC₅₀ to 23.2 nM, and co-treatment with ES936 further reduced it to 14.3 nM. Conclusion: Our findings identify NQO1 as a novel mediator of PI resistance in MM. High NQO1 expression is associated with significantly inferior PFS in PI-treated patients, but not in those receiving non-PI-based regimens, indicating a PI-specific mechanism of resistance. Functional studies confirmed that NQO1 overexpression reduces sensitivity to BTZ and CFZ, while pharmacologic inhibition with ES936 restored drug responsiveness. Furthermore, combined inhibition of NQO1 and ABCB1 synergistically enhanced CFZ sensitivity in resistant cells. Together, these results support NQO1 as both a predictive biomarker and a promising therapeutic target, offering a rationale for the development of combinatorial strategies to overcome PI resistance and improve outcomes in MM.

AMG 420 is a first-in-class bispecific T-cell engager (BiTE®) molecule directing a cytotoxic T-cell response toward multiple myeloma cells. This phase 1b, open-label, dose-expansion study (NCT03836053) evaluated the safety, tolerability, and efficacy of AMG 420 monotherapy in patients with relapsed/refractory multiple myeloma. Twenty-three patients received continuous intravenous infusion of AMG 420 (200-600 µg/day) in a 6-week cycle. Two dose-limiting toxicities (grade 3 staphylococcal sepsis and recurrent grade 2 cytokine release syndrome [CRS]) were reported. Commonly reported treatment-related adverse events included CRS, headache, and pyrexia. Overall response rate was 34.8%; median progression-free survival was 2.83 months; and minimal residual disease-negative complete responses were reported in 8.7% of patients. Overall, the safety profile and efficacy from AMG 420 established the proof-of-concept of T-cell engager therapy as a promising therapeutic class in multiple myeloma and BCMA as an effective target for T-cell engager therapy.

Given the evolving understanding of genetic risk factors in multiple myeloma (MM), this paper assesses whether next-generation sequencing (NGS) could complement or even replace fluorescence in situ hybridization (FISH) at diagnosis. A structured consensus process within European Myeloma Network (EMN) clinical and laboratory groups was conducted to establish recommendations on routine clinical deployment of NGS in MM risk assessment. Four key questions were addressed: (1) should NGS be used in addition to, or alternatively to FISH in identifying prognostic genetic markers, (2) which prognostic markers are most relevant for analysis by NGS, (3) which patients should be offered NGS testing, and (4) what is the optimal timing for performing NGS. The panel reviewed current literature, evaluated available NGS technologies, and compared their performance with that of FISH-based methodologies. The paper reviews current standard NGS protocols, quality control measures, and provides practical points for the implementation of an NGS diagnosis in MM. While NGS shows promise in improving risk stratification, challenges such as cost, accessibility, and clinical workflow integration must be addressed. The consensus supports the initial incorporation of NGS as a complementary tool to FISH. Recommendations emphasize that: a broader list of genetic events should be incorporated into such a test than what currently requested by risk scores; the test should be offered at least to the fit patients who could be candidates for modern triplet or quadruplet treatments; the test should be repeated at the time relapse, especially in the future when targeted treatments may mandate the use of predictive markers of response. This consensus provides a foundation for future research and policy development, guiding the adoption of NGS in MM risk assessment.

Proteasome inhibitor (PI) resistance remains a major therapeutic obstacle in the treatment of multiple myeloma (MM). MM cells demonstrate pronounced dependence on insulin and insulin-like growth factor-1 signaling via their cognate receptors, IGF-1R and INSR. In this study, we identify ceritinib, a clinically approved inhibitor of anaplastic lymphoma kinase (ALK), as a drug, which can inhibit IGF-1R/INSR activity and downstream PI3K/AKT/mTORC1 signaling. Ceritinib can overcome PI-resistance in MM when used in combination with carfilzomib. This synergy was consistently observed across in vitro and in vivo models, and primary patient-derived MM cells. Mechanistically, MM cells exploit IGF- 1R/INSR signaling to sustain expression of key molecular chaperones, including HSP70 and BiP, which are critical for maintaining proteostasis under conditions of high protein synthesis and turnover. Pharmacological inhibition of IGF-1R/INSR signaling by ceritinib abrogates this adaptive stress response, thereby preventing the upregulation of cytoprotective heat shock proteins upon proteasome inhibition. This disruption results in enhanced accumulation of protein aggregates, increased protein polyubiquitination, endoplasmic reticulum stress, and activation of apoptotic pathways. Collectively, our findings support the repurposing of ceritinib in combination with carfilzomib as a translationally relevant and safe strategy to circumvent PI resistance in MM, warranting further clinical investigation in the relapsed/refractory disease setting.

Background The combination of proteasome inhibitors (PI) and bispecific antibodies (bsAb) has demonstrated both synergistic and potentially antagonistic effects in the treatment of multiple myeloma (MM). While PIs may enhance bsAb-mediated cytotoxicity by increasing surface expression of target antigens, emerging evidence suggests that PIs may also promote immune evasion through up-regulation of immune checkpoint molecules. In this study, we aimed to dissect the immunological consequences of this combination by investigating the impact of the most commonly used PI bortezomib (BTZ) on T-cell subset composition and function in the context of bsAb therapy in MM. Methods Peripheral blood samples and clinical data from 104 pre-treated MM patients were prospectively collected and T-cell subsets were analyzed by flow cytometry. In vitro functional assays were conducted for BTZ, teclistamab (Tec) and talquetamab (Tal) using the PI-resistant MM cell line L363 and healthy donor T-cells. Proteasome subunit activity (β1, β2, β5) in T-cells was visualized with activity-based probes. MM cell viability was assessed, and cytokine concentrations were measured via ELISA in culture supernatants. Results Peripheral blood samples of 104 patients with pre-treated MM were analyzed. Median age of our cohort was 68 years (range 43-89) and patients were pre-treated with a median of 2 lines of therapy (range 1-8). Overall, 89 (85.6%), 27 (25.9%) and 13 (12.5%) patients had received PI, bsAb and chimeric antigen receptor modified (CAR) T-cell therapy, respectively, and 63 (60.6%) patients had prior high-dose melphalan and autologous stem cell transplant (ASCT). At sampling, 76 (73.1%) patients received ongoing anti-MM therapy while the remaining 28 (26.9%) patients were in a therapy-free interval. In a multivariate generalized linear model, prior Tal therapy was related with an increased proportion (%) of PD-1+ T-cells (β=3.55, P=0.03) while Tec positively correlated with increased frequencies of LAG-3+ (β=1.07, P=0.04) and TIM-3+ (β=9.16, P<0.001) T-cells. Moreover, exposure to BTZ led to a decrease of CD8+ LAG-3+ T-cells (β=-6.63, P=0.03). Since time between last treatment and sampling did not correlate with frequencies of different T-cell subsets in our study, these results suggest that anti-MM therapies including bsAb and BTZ may lead to persistent changes in circulating T-cell subsets in MM patients, potentially influencing the efficacy of T-cell based immunotherapies. Next, we performed functional assays to investigate whether the addition of BTZ to a bsAb modulates T-cell subsets and alters immune checkpoint expression on T-cells. Healthy donor T-cells were pre-treated with BTZ for 72 hours prior to co-culture to exclude potential confounding effects from reduced T-cell viability. A significant toxicity to healthy T-cells was observed upon treatment with 100 nM BTZ, resulting in >30% reduction in T-cell viability and significant inhibition (>50%) of proteasome subunits (β1, β2, and β5) after 72 hours. Co-culture of PI-resistant L363 cells with donor-derived T-cells plus Tal or Tec led to a significant increase in MM cell viability in the presence of BTZ (Tal: 2-fold increase, P=0.006; Tec: 5-fold increase, P<0.001). This finding was consistent with a marked reduction in cytokine production by T-cells, including IFN-γ (0.3-fold, P<0.001) and IL-6 (0.6-fold, P=0.03) in Tec with vs. without BTZ co-cultures. Phenotypic analysis of T-cell subsets revealed that BTZ significantly increased the CD4/CD8 ratio (Tal: P=0.005; Tec: P<0.001) and the frequency of effector memory T cells re-expressing CD45RA (Temra) (Tal and Tec: P<0.001), while reducing the proportion of central memory T cells (Tcm) (Tal and Tec: P<0.001). Additionally, we observed a significant decrease in the expression of the immune checkpoint markers PD-1, LAG-3 and TIM-3 on T-cells following BTZ co-treatment (PD-1: Tal and Tec, P<0.001; LAG-3: Tal P=0.03; Tec P<0.001; TIM-3: Tal P=0.006; Tec P<0.001). Our results suggest that the addition of BTZ to bsAbs may alter the T-cell activation by bsAb, characterized by decreased cytokine production, shift in T-cell subsets and lower expression of activation markers on T-cell, e.g. immune checkpoints. Conclusion Taken together, our findings provide the first evidence that BTZ may impair T-cell activation by bsAb in MM in vitro, thereby potentially diminishing the anti-tumor efficacy of bsAb.

Since the publication in 2021 of the European Hematology Association (EHA) Clinical Practice Guidelines for the treatment of patients with smouldering multiple myeloma (SMM) and multiple myeloma (MM), developed in collaboration with the European Society for Medical Oncology, a novel international staging system (R2-ISS) has been developed, several prognostic factors are entering clinical practice (such as minimal residual disease, circulating plasma cells and monoclonal protein assessed by mass spectrometry) and, at the time of writing, 14 novel regimens have been approved by the EMA and/or the FDA for the treatment of patients with MM. A multidisciplinary group of experts from the EHA and European Myeloma Network, based in various institutions mostly located in Europe, have updated the previous guidelines and produced algorithms for everyday clinical practice that incorporate levels of evidence and grades of recommendation based on the aforementioned new data. In these Evidence-Based Guidelines, we provide key treatment recommendations for both patients with newly diagnosed MM and those with relapsed and/or refractory MM, including guidance for the use of established drugs as well as contemporary immunotherapies. Novel approaches for the management of patients with SMM focus on those who might require early intervention. Finally, we provide recommendations for myeloma-related complications and adverse events, such as bone disease, renal impairment and infections, as well as for those associated with T cell-mobilizing therapies, such as cytokine-release syndrome and immune effector cell-associated neurotoxicity syndrome. In these Evidence-Based Guidelines, a multidisciplinary panel of experts from the European Hematology Association and the European Myeloma Network provide key treatment recommendations for patients with smouldering multiple myeloma, and newly diagnosed or relapsed and/or refractory multiple myeloma, addressing the use of established drugs and novel immunotherapies as well as the management of disease-specific and treatment-related complications and adverse events.

Multiple myeloma (MM) cells secrete high levels of immunoglobulin and are therefore addicted to mechanisms that maintain proteome homeostasis (proteostasis). While proteasome inhibitors that target the degradative aspect of proteostasis have proven effective, only limited attempts have been made to target protein secretion. Here we show that the receptor tyrosine kinase LTK is a regulatory node in the proteostasis network that responds to secretory load and helps cells maintain a high secretory output. LTK is a highly similar paralog to ALK and by repurposing existing ALK inhibitors, we demonstrate that targeting LTK causes immunoglobulin retention, ER stress and subsequent apoptosis of primary MM cells, even in patients refractory to proteasome inhibitors. Thus, LTK is a novel therapeutic target in the biosynthetic pathway of proteostasis, with significant potential for MM treatment.

The treatment landscape of heavily pretreated relapsed/refractory MM has changed considerably in recent years with the introduction of novel BCMA- and GPRC5D-directed immunotherapies, including CAR T-cell therapy, bispecific antibodies (BsAbs), and antibody-drug conjugates (ADCs). Treatment selection and sequencing become increasingly complex with the broad range of therapeutic options. In this review, the European Myeloma Network provides recommendations on how to best incorporate these novel therapies into the present treatment landscape using current evidence. The optimal treatment sequence depends on various patient- and tumor-related features, but also reimbursement and availability issues. In addition, mechanisms underlying relapse (e.g., antigen loss, reduced T-cell fitness, or outgrowth of T-cell resistant clones) dictate the efficacy of sequential BCMA- or GPRC5D-directed immunotherapy. BCMA-targeting BsAbs and ADCs should preferably be avoided prior to CAR T-cell therapy, as some studies have shown that these agents negatively influence clinical outcomes after CAR T-cell therapy. Therefore, we recommend the selection of CAR T-cell therapy first, and BsAbs and/or belamaf later in the disease course, if patients are eligible for CAR T-cell therapy and in case CAR T-cell therapy is available within a short time frame. However, bridging therapy with GPRC5D-directed BsAbs (initiation after apheresis) can be considered to significantly reduce tumor burden, because this was shown to improve the efficacy of consecutive BCMA-directed CAR T-cell therapy. Sequential treatment with agents targeting the same antigen, but with different modes of action, is feasible, but several studies have demonstrated that target switch is a more effective strategy. In addition, there is increasing evidence indicating that the efficacy of sequential use of BsAbs can be improved by creating a BsAb-free interval.

Overcoming proteasome inhibitor (PI) resistance remains an unmet medical need in multiple myeloma (MM) therapy. Anaplastic Lymphoma Kinase (ALK) inhibitors co-target related tyrosine-kinases, such as InsR and IGF1R. Ceritinib, a second-generation ALK inhibitor, has anti-MM activity in monotherapy or in combination with PI carfilzomib (CFZ). MM cells do not express ALK, but are dependent on InsR/IGF1R to sustain their high metabolic demand. Here, we aimed to identify the mechanism of action of ceritinib and the mechanism of synergy between ceritinib and CFZ in MM cells in vitro, in vivo, and ex vivo. PI-naïve and PI-resistant MM cell lines and primary cells were used. Genome-wide CRISPR/Cas9-based screening was performed to identify ceritinib resistance candidate genes. A pam-kinase screening, RNA sequencing and untargeted LC-MS/MS were used to elucidate mechanism of action of treatment combination. RNA-seq data from patients included in the CoMMpass study and from patients treated at the Wurzburg University Hospital were analyzed for InsR and IGF1R expression. An orthotopic in vivo mouse model with PI-resistant cells was used to determine the effect of the drug combination in vivo. Among several approved ALK inhibitors tested, ceritinib shows the highest synergistic cytotoxicity in combination with CFZ in MM cell lines and primary cells. The combination of ceritinib and CFZ is active in PI-resistant MM in vivo. MM cell lines and primary cells are negative for ALK but positive for InsR and IGF1R expression, which is increased in the relapsed/refractory MM cohort. Kinase screening identified InsR and IGF1R as major kinases inhibited by ceritinib in MM. CRISPR/Cas9-screening identified transcription factor FOXO1, a downstream target of InsR/IGF1R/Akt, as the top resistance candidate to ceritinib. Subsequently, FOXO1 inhibition protected MM cells from ceritinib-induced cytotoxicity. Ceritinib in combination with CFZ caused rapid translocation of FOXO1 to the nucleus, suggesting its activation. Consistently, ceritinib induced the expression of genes related to cell cycle arrest, which are regulated by FOXO1. Ceritinib, by targeting the InsR/IGF1R, impaired glycolysis and amino acid homeostasis, observed as an accumulation of aromatic amino acids. Collectively, ceritinib decreases Akt activity via InsR/IGF1R inhibition, leading to metabolic and proliferative shut-down. In combination with proteasome inhibition, this has a detrimental effect, impairing protein homeostasis and inducing starvation in MM cells. Ceritinib in combination with carfilzomib represents a promising treatment option for MM to be tested in the clinic. Supported by the project National Institute for Cancer Research (Programme EXCELES, ID Project No. LX22NPO5102) - Funded by the European Union - Next Generation EU. Andrej Besse, Tiberiu Totu, Marianne Kraus, Max Mendez Lopez, Jana Veprkova, Ondrej Slaby, Marija Buljan, Leo Rasche, K. Martin Kortum, Christoph Driessen, Lenka Besse. Combination of IGF1R/InsR and proteasome inhibition shows strong antitumor activity in proteasome inhibitor resistant multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4430.