High-risk Multiple Myeloma Patients Research Articles

Post-ASCT consolidation with elotuzumab, lenalidomide, and dexamethasone or elotuzumab, pomalidomide, and dexamethasone in high-risk and ultra-high-risk multiple myeloma: a retrospective single-center study

Patients with high-risk multiple-myeloma (HRMM) and ultra-high-risk multiple-myeloma (UHRMM) show rapid disease progression and shorter survival compared to those with standard-risk multiple-myeloma (SRMM). Lenalidomide maintenance after autologous stem cell transplant (ASCT) has shown inferior outcomes in this subgroup compared to SRMM, and there is an unmet need for improved post-ASCT therapy. This retrospective study, from September 2016 to March 2023, assesses elotuzumab combined with lenalidomide or pomalidomide and dexamethasone (ERd or EPd) as consolidation therapy post-ASCT for HRMM and UHRMM patients. HRMM (1 cytogenetic abnormality) and UHRMM (≥2 cytogenetic abnormalities) were defined using IMWG and mSMART criteria. Among 75 patients (median age: 64 years), 59 received ERd and 16 EPd. Median progression-free survival was 29.3 months for all patients, 32.7 months for HRMM, and 21.9 months for UHRMM. Elotuzumab plus an IMiD consolidation therapy post-ASCT demonstrated promising efficacy compared to other studies, with a fixed duration and reduced lenalidomide-related toxicity.

Identifying high-risk multiple myeloma patients: A novel approach using a clonal gene signature.

Multiple myeloma (MM) is a heterogeneous disease with a small subset of high-risk patients having poor prognoses. Identifying these patients is crucial for treatment management and strategic decisions. In this study, we developed a novel computational framework to define prognostic gene signatures by selecting genes with expression driven by clonal copy number alterations. We applied this framework to MM and developed a clonal gene signature (CGS) consisting of 22 genes and evaluated in five independent datasets. The CGS provided significant prognostic values after adjusting for well-established factors including cytogenetic abnormalities, International Staging System (ISS), and Revised ISS (R-ISS). Importantly, CGS demonstrated higher performance in identifying high-risk patients compared to the GEP70 and SKY92 signatures recommended for prognostic stratification of MM. CGS can further stratify patients into subgroups with significantly differential prognoses when applied to the high- and low-risk groups identified by GEP70 and SKY92. Additionally, CGS scores are significantly associated with patient response to dexamethasone, a commonly used treatment for MM. In summary, we proposed a computational framework that requires only gene expression data to identify CGSs for prognosis prediction. CGS provides a useful biomarker for improving prognostic stratification in MM, especially for identifying the highest-risk patients.

ACUTE MYOCARDIAL INFARCTION DURING VRD CHEMOTHERAPY IN A PATIENT WITH MULTIPLE MYELOMA: A CLINICAL CASE

Acute myocardial infarction is a critical condition associated with significant morbidity and mortality rates. Myocardial infarction-related acute myocardial injury is characterized by a rapid elevation and subsequent decline in cardiac troponin concentration. According to the relevant data patients with multiple myeloma are in a high-risk category for venous and arterial thrombosis. Therefore, the incidence of cardiovascular complications, which include myocardial infarction, in these patients is higher than in the general population. The development of metaplastic anemia further compounds this risk by diminishing myocardial oxygen supply. Moreover, chemotherapy for oncohematological diseases carries the potential for cardiotoxic cardiovascular complications. Immunomodulator drugs like Thalidomide and Lenalidomide, frequently utilized in multiple myeloma treatment, have been associated with Lenalidomide-induced myocardial infarction—a prevalent adverse effect. The use of proteasome inhibitors such as Bortezomib and Carfilzomib poses an increased risk for myocardial infarction development. This clinical case presents an instance of acute myocardial infarction in a multiple myeloma patient during low cumulative chemotherapy dosage, comprising Lenalidomide and Bortezomib. It underscores the necessity for enhanced clinical, instrumental, and laboratory monitoring before each specific chemotherapy course in high-risk multiple myeloma patients. Such monitoring facilitates the early detection of chemotherapy-induced cardiotoxic effects, allowing for timely intervention and management.

Chinese expert consensus on diagnosis and treatment of high risk multiple myeloma (2024)

High-risk multiple myeloma (HRMM) refers to patients with multiple myeloma whose overall survival time is less than 2-3 years under current standardized diagnosis and treatment. By combining various static and dynamic prognostic factors, risk stratification is performed to identify HRMM patients early and treat patients with personalized strategies, with the aim of significantly improving adverse survival outcomes in HRMM patients. Although the clinical value of HRMM has reached a consensus domestically in recent years, there still exist confusions and ambiguities in the definition, high-risk factors, risk stratification, and treatment of HRMM, necessitating standardization. In order to enhance the diagnostic and treatment capabilities of Chinese physicians in HRMM, the Professional Committee of Hematologic Malignancies of the Chinese Anti-Cancer Association (CACA) and the Multiple Myeloma Expert Committee of the Chinese Society of Clinical Oncology (CSCO) have organized relevant experts to develop this consensus. This consensus aims to clarify the definition of HRMM, high-risk factors, and risk stratification system, and provide treatment recommendations for HRMM, thereby improving the quality of life and prognosis of Chinese HRMM patients.

Loss of GABARAP mediates resistance to immunogenic chemotherapy in multiple myeloma

AbstractImmunogenic cell death (ICD) is a form of cell death by which cancer treatments can induce a clinically relevant antitumor immune response in a broad range of cancers. In multiple myeloma (MM), the proteasome inhibitor bortezomib is an ICD inducer and creates durable therapeutic responses in patients. However, eventual relapse and resistance to bortezomib appear inevitable. Here, by integrating patient transcriptomic data with an analysis of calreticulin (CRT) protein interactors, we found that GABA type A receptor–associated protein (GABARAP) is a key player whose loss prevented tumor cell death from being perceived as immunogenic after bortezomib treatment. GABARAP is located on chromosome 17p, which is commonly deleted in patients with high risk MM. GABARAP deletion impaired the exposure of the eat-me signal CRT on the surface of dying MM cells in vitro and in vivo, thus reducing tumor cell phagocytosis by dendritic cells and the subsequent antitumor T-cell response. Low GABARAP was independently associated with shorter survival in patients with MM and reduced tumor immune infiltration. Mechanistically, we found that GABARAP deletion blocked ICD signaling by decreasing autophagy and altering Golgi apparatus morphology, with consequent defects in the downstream vesicular transport of CRT. Conversely, upregulating autophagy using rapamycin restored Golgi morphology, CRT exposure, and ICD signaling in GABARAPKO cells undergoing bortezomib treatment. Therefore, coupling an ICD inducer, such as bortezomib, with an autophagy inducer, such as rapamycin, may improve patient outcomes in MM, in which low GABARAP in the form of del(17p) is common and leads to worse outcomes.

Serum indicators in functional high-risk multiple myeloma patients undertaking proteasome inhibitors therapy: a retrospective study

ABSTRACT Objectives Multiple myeloma (MM) is a class of malignant plasma cell diseases. An increasing application of autologous stem cell transplantation (ASCT) and anti-myeloma agents represented by proteasome inhibitors (PIs) has improved the response rates and survival of MM patients. Patients progressing within 12 months were recently categorized with functional high-risk (FHR), which could not be clarified by existing genetic risk factors, with poor outcomes. Our study aimed to investigate clinical indices related to FHR and seek prognostic roles in transplant-eligible MM patients. Methods Demographic and individual baseline clinical characteristics were compared by using the Pearson's chi-square and Mann-Whitney U test. Progression-free survival (PFS) and overall survival (OS) were described by Kaplan-Meier estimates and compared using the log-rank test. Logistic regression analysis was used to assess the association of baseline characteristics at MM diagnosis with FHR status. Results From 18th January 2010 to 1st December 2022, 216 patients were included and divided into two groups according to the FHR status. There was no difference in baseline data between the two groups. Renal impairment (RI, Scr > 2 mg/dL) was common in MM patients and made sense in FHR status. AST levels were validated as independent predictors for FHR status (p = 0.019). Discussion Patients with RI or higher AST levels (AST > 40 U/L) tended to have worse outcomes. However, transplants had apparently improved prognoses. Conclusion Therefore, in the PIs era, transplantations are still effective therapies for transplant-eligible MM patients.

Efficacy of Allogeneic Hematopoietic Stem Cell Transplantation in Young and High-Risk Multiple Myeloma Patients

Objective: With the new therapeutic agents developed over last decades, the median survival of patients with multiple myeloma has approached 10 years. However, 15-20% of patients who are classified as High-risk multiple myeloma (HRMM) patients still have a median survival of less than 3 years, which leaves room for improvements. Simultaneously, transplantation related mortality (TRM) has been decreasing since progress of supportive care during these years. Thus, this study aims to evaluate the efficacy of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in young high-risk multiple myeloma (HRMM) patients and to conduct statistical analysis of various prognostic factors in the new era. Method: This retrospective study analyzed clinical data from 14 young multiple myeloma patients with high-risk cytogenetic abnormalities or high-risk disease biological factors who underwent allo-HSCT at the Hematopoietic Stem Cell Transplantation Center of the Institute of Hematology & Blood Diseases Hospital between November 2016 and November 2022. Results: There were 7 males and 7 females involved in this study, with a median age of 39.5 (31- 50) years at the time of transplantation. The median number of treatment lines before transplantation was 2 (1-6). Before transplantation, 42.9% (6/14) of the patients did not achieve complete remission (CR), with 35.7% (5/14) of the patients performing MRD (+). After transplantation, all patients could be evaluated for treatment response, in which the overall response rate was 100% (14/14). All 14 patients successfully underwent transplantation, with median engraftment times for neutrophils and platelets of 11 (10-14) days and 13 (9-103) days, respectively. Acute graft-versus-host disease (GVHD) of grades II-IV occurred in 5 patients (35.7%), and 2 patients (14.3%) developed moderate to severe chronic GVHD. The median follow-up time after transplantation was 18.93 (4.1-72.53) months, with the 2-year TRM of 7.1% (95%CI 0-19.7%) and the expected 2-year overall survival (OS) rate of 92.9% (95% CI 80.3-100%). Meanwhile, the expected 1-year and 2-year progression-free survival rates were 85.7% (95% CI 69.2-100%) and 45.7% (95% CI 20.9-99.9%), respectively, and the presence of chronic GVHD might influence the outcome positively. Conclusion: Allogeneic hematopoietic stem cell transplantation is an effective treatment to improve the prognosis of young and high-risk multiple myeloma patients.

Dysregulation of Cellular Immunity in High-Risk Multiple Myeloma

Introduction: Multiple myeloma (MM) is the second most prevalent hematological malignancy, characterized by the abnormal proliferation of plasma cells in the bone marrow. Approximately 20% of MM patients present with high-risk (HR) clinical and genetic features that are associated with aggressive disease course and poor prognosis. Novel immune therapies are changing the paradigm of MM treatment. A comprehensive understanding of immune dysregulation within the tumor microenvironment (TME) of HR vs standard risk (SR) MM patients could inform the development of targeted immunotherapeutic strategies for HR MM. Methods: 55 HR and 25 SR patients were selected from the Mt. Sinai Hematologic Malignancies Tissue Bank (IRB# STUDY-11-01978). HR included one or more cytogenic abnormalities (HRCA) such as translocations t(4;14), deletion of chromosome 17 (del17p), and gain/amplification of 1q (1q+/++) based on the Second Revised International Staging System (R2-ISS). We also included nine patients with t(14;16) translocation alone or combined with one or two of the other HRCA. We performed comprehensive immune phenotyping of the bone marrow micro-environment on specimens from HR and SR patients using mass cytometry (CyTOF) and single cell RNA sequencing (scRNASeq) of bone marrow mononuclear cells (BMMC) depleted of CD138 positive MM cells. Results: ScRNAseq of BMMC from HR MM patients revealed a robust cytotoxic and cytolytic signature on CD4+ T cells relative to SR, characterized by the upregulation of genes such as GNLY, NKG7, PRF1, and IFNG, and a higher frequency of Granzyme K-positive (GZMK+) CD8+ T cells, previously described as progenitors of exhausted cells. Both CD4 and CD8 T cells, along with monocytes, exhibited high expression of a group of Interferon-stimulated genes (ISGs) including IFI27, IFIT3, IFI44L, IFITM1/3, and OASL. Notably, high expression of this group of ISGs was associated with T cell exhaustion and poor prognosis in the setting of immune checkpoint inhibitor therapy for solid tumors. Mass cytometry analysis confirmed the activated state of T cells in HR MM patients, with increased expression of immune activation markers HLA-DR, CD38, and the senescence marker CD57. Furthermore, HR patients exhibited higher frequencies of mast cells, CD14+ Monocytes and plasmacytoid dendritic cells compared to SR, suggesting an inflammatory and immunosuppressive environment that may dampen effective antitumor immune responses. Conclusions: These results indicate distinct bone marrow microenvironments of HR and SR MM. CD4 and CD8 T cells appear to be more activated in HR patients compared to SR. However, the higher frequencies of mast cells and plasmacytoid dendritic cells, and the prominent expression of ISGs associated with inhibition of anti-tumor immunity suggest dysregulation and exhaustion of T cell activity in HR patients. Collectively, these findings highlight the complex immune dynamics in HR MM and provide insights into the underlying immune dysfunction which may contribute to disease progression and therapy resistance in HR MM. Such insights have the potential to inform the design of targeted therapeutic interventions aimed at modulating the immune response and improving treatment outcomes for HR MM patients.

Safety and Efficacy Outcomes for Patients with High-Risk Multiple Myeloma Receiving Idecabtagene Vicleucel: The MD Anderson Experience

Background: There is a paucity of data regarding the impact of high-risk features on the efficacy and safety of B-cell maturation antigen (BCMA)-directed autologous chimeric antigen receptor (CAR)-T therapy in patients with multiple myeloma (MM). Methods: In this retrospective single-center study we included all MM patients who received idecabtagene vicleucel (ide-cel) between 12/2019-10/2022, either in the setting of a clinical trial (i.e. KarMMa-2 and KarMMa-3) or as a standard of care (SOC). We evaluated the impact of high-risk disease features on the efficacy and safety outcomes. Patients with high-risk MM (HRMM) were defined as having at least one of the following: high-risk cytogenetic abnormalities prior to CAR-T infusion (deletion 17p, translocation (t)(4:14), t(14:16) or 1q gain/amplification (1q+)), extramedullary disease (EMD), plasma cell leukemia and Revised International Staging System (R-ISS) stage 3. Minimal residual disease (MRD) was evaluated using 8-color next-generation flow cytometry (NGF), with a sensitivity of 1/10-5 cells (0.001%) based on the acquisition and analysis of at least 2 million events. Efficacy outcomes included response rates, progression free survival (PFS) and overall survival (OS). Safety outcomes included incidence/grade of cytokine release syndrome (CRS), Immune effector cell-associated neurotoxicity syndrome (ICANS), hematological toxicity and infections. Results: A total of 74 patients were included in our analysis, 49 received SOC ide-cel and 25 were treated on clinical trial. The median age was 65.3 (range 37.9-90.3) years, and 49 patients (66%) were male. Median number of previous lines of therapy was 6 (range 1-18). The HRMM group included 52 patients (70%), and the most prevalent high-risk features were high-risk cytogenetic abnormalities (43%, most often 1q+ (n=26, 81%), followed by del17p (n=17, 33%), t(4:14) (n=9, 17%) and t(14:16) (n=4, 8%)) and EMD (19%). Seventy five percent of patients who received SOC ide-cel had HRMM compared to 25% of those who received ide-cel on clinical trial (p=0.014). Compared to patients with standard-risk MM (SRMM), those with HRMM more often had disease refractory to proteasome inhibitors (p=0.042) or anti-CD38 antibodies (p=0.01), and more often received bridging therapy prior to CAR-T infusion (p=0.021). After bridging therapy, 43% of HRMM patients had progressive disease, compared to 20% of SRMM patients (p=0.21). Sixty seven percent of the entire cohort received prior BCMA-directed therapy before ide-cel, without a significant difference between the two risk groups (p=0.80) ( Table 1). For the entire cohort, overall response rate was 85%, and 51% achieved ≥CR at best response. There was no significant difference between the HRMM and SRMM groups in response rates or in rates of MRD negativity at day 30 and at 3/6/9 months after CAR-T infusion, yet fewer HRMM had MRD negativity at 12 months after CAR-T (50% vs. 100% of evaluable patients, p=0.019). The 6-months PFS for the HRMM group and the SRMM group was 64% and 81%, respectively (p=0.11) ( Figure 1A), and the 6-months OS was 80% and 90%, respectively (p=0.067) ( Figure 1B); Thirty-nine patients in the HRMM group developed CRS (83% of evaluable patients), including one grade 3 and one grade 5 CRS, compared to 15 (75%, p=0.65) in the SRMM group (all grade <3). Eight patients in the HRMM group developed ICANS (17%), including two grade 4 ICANS, compared to 3 (15%, p=0.93) in the SRMM group (all grade <3). We did not observe a difference in rates of neutropenia (p=0.63), anemia (p=0.75) or thrombocytopenia (p=0.53) by day 30, and also not in the rates of infection (p=0.16) between the two groups. In multivariate analysis on the entire cohort, receiving <4 prior lines of therapy (HR 0.06, 95% CI 0.004-0.71, p=0.026) and ECOG≤2 (HR 0.07, 95% CI 0.08-0.53, p=0.01) were associated with superior OS. Conclusions: Patients with HRMM receiving ide-cel had mostly comparable response rates compared to those with SRMM. HRMM patients had numerically worse PFS and OS compared to patients with SRMM, though not reaching statistical significance. Overall rates of CRS/ICANS were comparable between the two groups, yet only patients with HRMM had grade ≥3 of these complications. Patients who received ide-cel after less than four previous lines of therapy had better overall survival.

Promising Safety and Efficacy of Trovocabtagene Autoleucel (C-CAR088) Followed ASCT in Ultra High-Risk Multiple Myeloma (UHR-MM) Patients Who Failed or Had Suboptimal Response to Standard First Line Triplet Based Therapy

Background: The prognosis of MM has improved significantly nowadays. However, patients with ultra-high-risk features (UHR-MM) had dismal outcomes even treated with standard front-line therapy. This study (NCT05632380) explores the potential of new CAR-T therapy in addressing the unmet need of these patients. Trovocabtagene autoleucel (trovo-cel), an anti-BCMA CAR T-cell manufactured by Cellular Biomedicine Group, Inc., showed deep and lasting responses with a good safety profile in relapsed or refractory multiple myeloma (R/R MM). Hereby, we are reporting a phase I, open-label, single-arm study, assessing the feasibility of trovo-cel followed ASCT in UHR-MM patients who failed or had suboptimal responses to the first-line standard therapy. Methods: Eligible patients are 18-70 years old, transplant-eligible and confirmed as UHR-MM defined as follows: 1) Genetic ultra-high risk: del(17p)≥60%; or ≥2 high-risk cytogenetic abnormalities including TP53 mutation, del(17p)/P53 deletion, t(4;14), t(14;16), t(14;20), 1q21 gain/ amplification, and did not reach CR or better response after 4 cycles standard first-line therapy; 2) Primary refractory: <MR after 2 cycles or <PR after 4 cycles standard first-line therapy; 3) Early relapse: lost best response within 6 months from the standard first-line therapy; 4) primary PCL history and < CR after 4 cycles standard first-line therapy. Standard first-line therapy should be a potent standard triplet-based regimen including one PI, one IMIDs, and glucocorticoid. Quadruplet or quintuplet regimens might be used. Prior ASCT, BCMA-targeted therapy, or CAR-T treatment are excluded. Patients undergo conditioning (melphalan alone or plus fludarabine) followed by ASCT on Day 0 and trovo-cel 3.0 or 6.0×10 6 cells/kg single intravenous infusion on Day 3. Maintenance is allowed after 3 months post-transplantation per investigator's decision. Results: As of July 24, 2023, 8 UHR-MM patients completed ASCT and trovo-cel infusion. Five patients received melphalan and 3 received fludarabine combining melphalan pretreatment. Trovo-cel was administrated at 3 × 10 6 cells/kg for all 8 patients. The median age of patients was 54.5 years (range: 39-66). The median number of prior lines of therapy was 2 (range: 1-2), with VRD exposure rate of 100%. Seven patients (87.5%) had received daratumumab, with six having been treated with dara-based quadruplet or quintuplet therapy as a front setting. Two patients (25%) had genetic ultra-high risk features, four patients (50%) had primary refractory myeloma, two (25%) had early relapse myeloma, and three patients (37.5%) had primary PCL history, two (25%) had extramedullary disease. Five patients (62.5%) received bridging and 6 received maintenance therapy. The combination of ASCT and Trovo-cel was well tolerated. With a median follow-up of 184 days (range: 13-332 days), no unexpected adverse events was observed. Among evaluable 7 patients, six (85.7%) had experienced grade 1 CRS and fully recovered. Two patients (28.6%) received tocilizumab and glucocorticoid for CRS. No ICANS event was reported. As expected, hematological toxicities are the most common AEs. All 7 patients achieved hematopoietic reconstitution successfully, with a median time of 14 days for ANC (≥0.5×10 9/L) and 13 days for PLT (≥20×10 9/L) post ASCT. The ORR was 100%, with 6 CR with MRD negative (NGF, 10 -5) (85.7%), 1 VGPR (14.3%). Robust CAR T-cell expansion was observed in blood of all patients. The median time of peak of CAR-T expansion (T max) was 7 days (range 4~7), the median peak CAR-T copy number (C max) was 891,963 copies /μg gDNA and the median AUC 0~28day was 6,009,475.5 copies /μg gDNA/day. The PK profile is similar to trovo-cel single agent treatment 1] but with earlier peak of CAR-T expansion. Conclusion: The preliminary results of this clinical trial show a promising safety and efficacy profile of trovo-cel followed ASCT in UHR-MM patients. Although the follow-up time is relatively limited, we are looking forward to a consistently good safety and efficacy outcome in long-term follow-up and in more patients recruited later. Research Sponsor: Institute of Hematology & Blood Diseases Hospital Collaborator: Cellular Biomedicine Group, Inc.

Discovery and Development of CD70 As a Cellular Therapy Target in High Risk Multiple Myeloma

Background: Even in the BCMA CAR-T era, multiple myeloma (MM) patients with high-risk genotypes have the poorest outcomes. We thus leveraged MMRF CoMMpass data to identify potential novel surface immunotherapeutic targets in high-risk MM. As described below, we focused on CD70, an immunotherapeutic target known to be upregulated on many hematologic and solid tumors but minimally expressed on normal tissue. While CD70 was previously explored in MM (McEarchern et al, Clin Cancer Res (2008)), it was not further investigated due to heterogeneous expression in newly diagnosed samples. Here we revisit this target to suggest cellular therapies against CD70 could be highly beneficial in high-risk MM patients, currently the greatest unmet need in the field. Methods: Primary patient samples were obtained under an IRB-approved protocol. CAR-T lentiviral transduction was performed on paired CD4/CD8 human T cells from at least 3 healthy donors. Effector function was assessed by cytotoxicity against MM cell lines and CAR-T profiling for memory and exhaustion markers. In vivo efficiency was assessed using bioluminescence. ATAC-seq peaks were called using Promo and HOMER (data from Jin et al. Blood (2018)). Machine learning was performed using XGBoost approach applied to CoMMpass RNA-seq data with 80% of samples for training and 20% as test set. Results: Analyzing CoMMpass mRNA data (release IA19) we first evaluated surface protein encoding-genes upregulated in tumors at relapse compared to diagnosis. Among >100 significantly upregulated genes, we focused on CD70 given known promise as an immunotherapy target. Further analysis across genomic subtypes revealed a specific, marked upregulation of CD70 in t(4;14) and gain 1q ( Fig. 1), as well as R-ISS 3 at diagnosis. Flow cytometry profiling of 8 cell line models, including t(4;14) models KHM11, KMS26, and LP1, revealed consistent CD70 protein expression with antigen density similar to BCMA. Profiling of relapsed primary MM samples confirmed that 19 of 28 samples had CD138+ plasma cells positive for CD70 expression. Of 5 patients in whom both BCMA and CD70 were measured, antigen densities were also similar. To target CD70, we then designed 10 unique anti-CD70 CAR-T constructs. 9 of these were scFv-based binders derived from therapeutic antibody fragments currently in preclinical or clinical development. By a structure-guided design based on the CD70:CD27 co-crystal structure (PDB: 7KX0), we also derived a unique truncated fragment of CD27 as an alternative CD70 binding element. All CD70 CARs were efficacious in in vitro cytotoxicity assays targeting MM cell lines AMO1, MM1.S, LP1, and RPMI8226. We then selected the top 3 performing scFv based-CARs as well as our natural ligand (CD27) CAR-T for in vivo studies. All 4 CD70 CAR-Ts prolonged survival of MM.1S-bearing mice compared to empty CAR ( n=5 mice/arm, p<0.005 by log-rank), similar to BCMA CAR-T controls. Remarkably, the CD27-based CAR showed superior expansion (50-85x, p< 0.0001 by ANOVA) and persistence compared to scFv-based ( Fig. 2). These results underscore that targeting CD70 using a natural ligand-based CAR-T represents a promising preclinical candidate for treatment of relapsed and high risk MM. We further investigated why CD70 may be upregulated in high risk MM. Analysis of ChIP-seq in ENCODE and primary MM tumor ATAC-seq was used to nominate a set of transcription factors that bind the CD70 locus. Extending from CoMMpass data, we built a predictive machine learning model for CD70 expression as a function of transcription factor gene expression, nominating key transcription factors TFAP2A, LEF1, MYB, and PAX5 as CD70 regulators; validation is now underway. Additionally, we showed that knockout of NSD2 in t(4;14) models KMS34 and KMS11 led to significantly decreased CD70 surface expression, implicating this central epigenetic driver in CD70 overexpression in this high-risk MM subtype. Finally, we also demonstrated that treatment with the hypomethylating agent azacytidine can drive increased CD70 expression, similar to that shown in AML models. Conclusion: Taken together, targeting CD70 appears to be a highly promising cellular therapeutic strategy for MM, with particularly high utility in high-risk patients after BCMA CAR-T relapse. We are continuing preclinical and future clinical development of this CAR-T strategy, using our natural ligand CD27 design with extended in vivo persistence.

Ultra High-Risk Multiple Myeloma Patients with Multi-Hit Tumours and SKY92 High Risk Signature Are at Increased Risk of Early Relapse Even When Treated with Extended Intensified Induction and Consolidation - Results from the Optimum/Muknine Trial

Background: We recently reported improved outcomes for patients with ultra-high risk (UHiR) newly diagnosed multiple myeloma (NDMM) by double hit/≥2 high-risk cytogenetic abnormalities (HRCA) and/or SKY92 risk signature or primary plasma cell leukemia (pPCL) in the OPTIMUM/MUKnine (NCT03188172; Kaiser et al, JCO 2023) trial. However, some participants still relapsed within the first 18 months of treatment despite stratified intensive therapy with Dara-CVRd, V-ASCT and extended Dara-VR(d) consolidation. In this analysis we characterise factors associated with early progression. Methods: In OPTIMUM, patients with UHiR NDMM (≥2 HRCA: t(4;14), t(14;16), t(14;20), gain(1q), del(1p), del(17p) or SKY92 high risk signature) or with pPCL received up to 6 cycles of Dara-CVRd induction, V-ASCT, 6 cycles Dara-VRd (Cons1) and 12 cycles Dara-VR (Cons2), before moving to monthly Dara-R maintenance until progression. OPTIMUM recruited 107 UHiR patients, including 9 pPCL. In this exploratory analysis, baseline clinical, laboratory and tumour molecular characteristics were compared between patients who experienced an Early Relapse, defined by MM relapse or MM-related death within 18 months from registration and those who did not, using descriptive statistics and hypothesis testing using two-sample t-tests or chi squared tests. Baseline characteristics were also compared by minimal residual disease (MRD; 10 -5 sensitivity by flow cytometry) status at the end of induction to elucidate whether baseline factors associated with early MRD negativity were overlapping with those associated with Early Relapse. Median follow-up was 51.5 months, and all patients had been on study for over 44 months. Results: Twenty-three (21%) of 107 UHiR patients experienced Early Relapse, versus 84 (79%) who did not. Patient age and gender were not different between groups. Platelet levels as a continuous variable were lower in the Early Relapse group vs those without (mean 179/nL, standard deviation (SD) 92/nL vs. mean 223/nL, SD 77/nL; P=0.0247), whereas haemoglobin, serum creatinine, calcium, presence of soft tissue plasmacytomas (PCM), bone lytic lesions or plasma cell bone marrow infiltration on diagnostic trephine were similar. ISS stage distribution differed between groups (P=0.0203): most Early Relapse participants were ISS stage 2 (13/23 (56.5%) vs 30/84 (35.7%) without Early Relapse), whereas ISS stage 3 was nominally similar (34.8% vs 31.0%). Most notably, only one of the 29 ISS stage 1 patients in OPTIMUM experienced Early Relapse. 57 of 107 (53.3%) patients entered OPTIMUM with ≥2 HRCA at diagnosis, and, of these, seven carried 3 HRCA. Five of seven participants (71.4%) with 3 HRCA experienced Early Relapse, opposed to only seven out of 50 (14%) with only 2 HRCA. 82 of 107 (76.6%) were SKY92 high risk at baseline. Only two of the 24 participants (8.3%) with SKY92 standard risk experienced Early Relapse, as opposed to 20 out of 82 (24.4%) with SKY92 high risk. Of individual HRCAs, only presence of del(17p) was associated with early relapse (P=0.002): 8 of 16 (50%) participants with del(17p) tumours experienced Early Relapse. However, seven of these 8 Early Relapse del(17p) tumours (88%) were also positive for SKY92 high risk signature. In contrast, the majority (62%) of del(17p) that did not relapse early were SKY92 standard risk. Of the 107 UHiR patients, 6 did not reach end of induction and 14 had no MRD result. Of 87 participants with interpretable MRD result at end of induction (EOI), 51% tested MRD-negative (44/87). Of clinical factors, only presence of PCM was associated with insufficient response (P=0.017, 5/5 MRD-positive), whereas ISS stage was not associated with MRD at EOI. Only one of four participants (25%) with 3 HRCA had MRD-negative status at EOI vs 66% of those with 2 HRCA. Also, the majority of patients (62%) identified by SKY92 high risk only were still MRD-positive. Conclusions: Our results suggest that presence of 3 or more HRCA, SKY92 high risk signature or del(17p) combined with SKY92 high risk are all associated with increased risk of Early Relapse despite intensified OPTIMUM treatment, and could help identify patients early who are candidates for novel, innovative first-line therapeutic approaches. Results also suggest utility of integrated molecular diagnostics, including gene expression risk profiling, to support clinical decision making in the era of advanced front-line combination therapies.

EDNRA-Expressing Mesenchymal Cells Are Expanded in Myeloma Interstitial Bone Marrow and Associated with Disease Progression.

Multiple myeloma (MM) induces dysfunctional bone marrow (BM) mesenchymal cells and neoangiogenesis. Pericytes and smooth muscle cells (SMCs) could detach from vessels and become cancer-associated fibroblasts. We found that the pericyte and SMC marker endothelin receptor type A (EDNRA) is overexpressed in whole MM bone biopsies; we sought to characterize its expression. EDNRA expression gradually increased with disease progression. High-risk MM patients had higher EDNRA expression than low-risk MM patients and EDNRA expression was highest in focal lesions. High EDNRA expression was associated with high expression of pericyte markers (e.g., RGS5, POSTN, and CD146) and the angiogenic marker FLT1. A single-cell analysis of unexpanded BM mesenchymal cells detected EDNRA expression in a subset of cells that coexpressed mesenchymal cell markers and had higher expression of proliferation genes. Immunohistochemistry revealed that the number of EDNRA+ cells in the interstitial BM increased as MM progressed; EDNRA+ cells were prevalent in areas near the MM focal growth. EDNRA+ cells were detached from CD34+ angiogenic cells and coexpressed RGS5 and periostin. Therefore, they likely originated from pericytes or SMCs. These findings identify a novel microenvironmental biomarker in MM and suggest that the presence of detached EDNRA+ cells indicates disrupted vasculature and increased angiogenesis.

CRIP1 INDUCES PIS RESISTANCE BY FORMING A COMPLEX WITH USP7 /PA200 AND ENHANCING PROTEASOME ACTIVITY AND AUTOPHAGY IN MULTIPLE MYELOMA

Introduction: Multiple myeloma (MM) is a plasma malignancy characterized by an accumulation of misfolded immunoglobulins in the cell. Although proteasome inhibitor (PI) can treat MM by inhibiting ubiquitin-proteasome system (UPS), autophagy as a compensatory protein clearance mechanism, it still leads to drug resistance. The discovery of new targets that can simultaneously inhibit UPS and autophagy will provide great potential for the treatment of MM. Methods: The proliferation, invasion, migration, proteasome activity and autophagy were detected in CRIP1- knockdown /overexpression and control MM cells. Co-IP (Co-Immunoprecipitation) with TAP/MS (Tandem affinity purification/Mass spectrum) was performed to detect the binding of USP7, CRIP1 and PA200. Myeloma xenograft model was used to determine the role of CRIP1 to promote proliferation of MM cells and induce BTZ resistance in vivo. Results: The levels of CRIP1 were significantly increased in plasma cells from NDMM (newly diagnosed multiple myeloma) compared to healthy donors, and further increased in patients with RRMM (relapsed/refractory multiple myeloma). Moreover, the high expression of CRIP1 is significantly related to the poor prognosis of MM patients, especially those treated with BTZ. Our results in vitro and in vivo showed that CRIP1 knockdown noteworthy inhibited the cell growth, invasion, migration. MM cells were more sensitive to PIs-induced cell growth arrest when CRIP1 knockdown. The proteasome activity and the protein level of LC3B-Ⅱ of the MM cells were significantly reduced with CRIP1 knockdown. Co-IP analysis showed that CRIP1 formed a complex with USP7/PA200. The protein level of CRIP1 decreased and the ubiquitination CRIP1 increased with USP7 inhibition. These data suggested that CRIP1 is the substrate of USP7.CRIP1 promotes the de-ubiquitination and stabilization of PA200 mediated by USP7 by interacting with USP7 and PA200, which in turn promotes MM cell survival and drug resistant. Functional inhibition of CRIP1 by knocking down of USP7 or PSME4 in CRIP1 OE MM cell line led to a decrease of the activity of proteasome and autophagy, thus enhancing the sensitivity to PIs in intro and in vivo. Conclusion: CRIP1 plays a critical role in MM progression and PIs resistance by formation of CRIP1/USP7/PA200 complex. High level of CRIP1 is a biomarker for high-risk MM patients. CRIP1 is a potential target for MM therapy, can simultaneously inhibit proteasome activity and autophagy. Encore Abstract - previously submitted to EHA 2023 The research was funded by the following foundations:(a) CAMS Innovation Fund for Medical Sciences (CIFMS) (2021-I2M-1-040 and 2022-I2M-1-022), (b) National NaturalScience Foundation of China (82170194, 81920108006, 82270175) and Natural Science Foundation of Fujian Province of China (2021J02040). Keywords: Diagnostic and Prognostic Biomarkers, Multiple Myeloma, Tumor Biology and Heterogeneity No conflicts of interests pertinent to the abstract.

P52 RISK STRATIFICATION COMBINING SKY92 GENE EXPRESSION PROFILING AND TRADITIONAL FISH IN MULTIPLE MYELOMA: THE FIRST PROSPECTIVE EVIDENCE IN THE R2-ISS ERA

High-risk (HR) multiple myeloma (MM) patients still have a very poor prognosis. However, the definition of HR MM remains controversial. Currently, FISH is the most commonly used method for risk stratification in MM. According to the Second Revised International Staging System (R2-ISS), HR cytogenetics is defined as presence of at least one of the following: del(17p), t(4;14), and 1q CNA. In recent years, gene expression profiling (GEP) approaches, e.g. SKY92, have been developed for detection of HR patients. In the R2-ISS era, however, data on MM risk stratification applying SKY92 in combination with traditional FISH is still lacking. Therefore, we performed the first prospective study evaluating the prognostic value of the combined SKY92 and FISH HR detection in newly diagnosed (ND) and relapsed/refractory (RR) MM. We prospectively collected bone marrow (BM) samples and clinical data of 147 MM patients. Cytogenetics were analyzed on purified CD138 positive MM cells by FISH, and HR cytogenetics was defined according to the R2-ISS classification. SKY92 risk status was determined with MMprofiler gene expression assay. Whole genome sequencing (WGS) was performed to compare the both risk stratification systems SKY92 and FISH. We included 147 patients in our study (NDMM: n=51, RRMM: n=96). SKY92 classification was available for 121 (82.3%) patients. HR SKY92 was significantly enriched in RRMM (40/76) compared with NDMM (6/45) (P<0.0001). RRMM patients with HR SKY92 showed significantly shorter progression free survival (PFS) (P<0.0001) and overall survival (OS) (P=0.0004) than standard-risk (SR). In NDMM, HR SKY92 also indicated a significantly inferior PFS (P=0.001) in comparison with SR. Of note, samples of 26 (17.7%) patients, who showed significantly lower bone marrow infiltration than the remaining patients (median: 25% vs 55%, P=0.038), did not meet the SKY92 quality control criteria. We then combined the SKY92 classification with cytogenetic analyses by FISH, which were available in 99 patients (NDMM: n=33; RRMM: n=66). We noticed a discrepancy between both risk stratification systems, with 44 (44.4%) and 58 (58.6%) patients being classified as HR by SKY92 and FISH, respectively. In total, 28 (42.4%) RRMM patients and 6 (18.2%) NDMM patients displayed HR in both SKY92 and FISH (double-HR). Regarding survival outcome in RRMM, double-HR patients showed the worst PFS (P<0.0001) and OS (P=0.0007). In NDMM, double-HR presented a negative prognostic factor for PFS (P=0.01). To compare the FISH and SKY92 classifications, we performed WGS in 16 patients who exhibited either only HR SKY92 (n=7) or only HR FISH (n=9). Interestingly, 1 patient with bi-allelic TP53 inactivation (deletion + mutation) and 6 patients harbouring 1q CNA were determined as SR by SKY92 but as HR by FISH. The median PFS was not reached after a median follow up of 371 days in these 9 patients. Moreover, 4 out of 7 patients with only HR SKY92 but SR FISH displayed 1q CNA, which was detected only by WGS, and del1p32 was found in 1 patients. Interestingly, we found CRBN mutation in 3 out of 7 patients with only HR SKY92 but SR FISH. The remaining 2 patients did not show any known HR genomic alterations, suggesting that HR MM may be associated with other factors, e.g. epigenetic modifications. Here, we provide the first prospective evidence in the R2-ISS era that advanced risk stratification combining SKY92 and FISH may help to identify the highest-risk MM.

Impact of clonal plasma cells in autografts on outcomes in high-risk multiple myeloma patients

Most patients with multiple myeloma (MM) undergoing autologous hematopoietic stem cell transplantation (autoHCT) eventually relapse, perhaps due to the presence of clonal plasma cells (CPC) in the autograft. We conducted a retrospective analysis to evaluate the impact of CPC in the autograft on the outcomes of high-risk chromosomal abnormalities (HRMM) patients undergoing autoHCT between 2008 and 2018. Patients were divided into CPC+ or CPC− in the autograft by next-generation flow cytometry (NGF). There were 75 CPC + autografts (18%) and 341 CPC− (82%). The CPC + group was less likely to achieve MRD-negative complete remission post-transplant (11% vs. 42%; p < 0.001). Median progression free survival (PFS) and overall survival (OS) were (12.8 vs. 32.1 months) and (36.4 vs. 81.2 months) in the CPC + and CPC− groups, respectively (both p < 0.001). Also in the subset of patients with MRD-negative ≥VGPR prior to autoHCT, those with CPC + autografts had inferior PFS (HR 4.21, p = 0.006) and OS (HR 7.04, p = 0.002) compared to CPC-. In multivariable analysis, the degree of CPC positivity in the autograft was independently predictive of worse PFS (HR 1.50, p = 0.001) and OS (HR 1.37, p = 0.001). In conclusion, both the presence and degree of CPC in the autograft were highly predictive of inferior PFS and OS.

Early Efficacy Observation of Pomalidomide-based Regimen in the Treatment of High-risk Multiple Myeloma

To investigate the efficacy and safety of pomalidomide based regimen in the treatment of high-risk multiple myeloma (MM). Clinical data of 27 high-risk MM patients treated in Shanxi Bethune Hospital from January 2021 to December 2022 were retrospectively analyzed. All patients were treated with pomadomide based regimen for at least 2 consecutive cycles, and the early therapeutic effect and safety were observed. Overall remission rate (ORR) was 63.0%(17/27 cases) and deep remission rate was 22.2%(6/27 cases) after 2 cycles of treatment; ORR was 90.5%(19/21 cases) and deep remission rate was 66.7%(14/21 cases) after 4 cycles of treatment. Both ORR and deep remission rate were significantly higher after 4 cycles of treatment than 2 cycles (P=0.044, P=0.003). Beyond that, in the newly diagnosed and relapsed refractory MM groups, ORR after 2 cycles of treatment were 75%(9/12 cases) and 60%(9/15 cases), and deep remission rates were 25%(3/12 cases) and 20%(3/15 cases), respectively; ORR after 4 cycles of treatment were 100%(9/9 cases) and 83.3%(10/12 cases), and deep remission rates were 77.8%(7/9 cases) and 58.3%(7/12 cases), respectively, while there was no significant difference in remission rates between the two groups (P>0.05). In the group of creatinine ≥177 μmol/L, the serum creatinine level was significantly decreased after 2 cycles of treatment compared with that pre-treatment (P=0.001). The 1q21 amplified subgroup accounted for the largest proportion (21/27 cases), ORR was 66.7%(14/21 cases) and deep remission rate was 23.8%(5/21 cases) after 2 cycles of treatment, ORR was 88.9%(16/18 cases) and deep remission rate was 66.7%(12/18 cases) after 4 cycles of treatment. In all the symptoms, the most common adverse reactions were pulmonary infection in 9 cases and hematological adverse reactions of grade 1-2 in 8 cases. The pomalidomide-based treatment regimen has good early curative effect on the newly diagnosed and relapsed refractory high-risk MM, and also benefits to the high-risk cytogenetic MM, or MM with renal impairment. Therefore, this treatment regimen showed a good safety, and the long-term curative effect needs to be further assessed by more clinical data.

Combined inhibition of Wee1 and Chk1 as a therapeutic strategy in multiple myeloma.

Multiple myeloma (MM) is a hematological malignancy characterized by an abnormal clonal proliferation of malignant plasma cells. Despite the introduction of novel agents that have significantly improved clinical outcome, most patients relapse and develop drug resistance. MM is characterized by genomic instability and a high level of replicative stress. In response to replicative and DNA damage stress, MM cells activate various DNA damage signaling pathways. In this study, we reported that high CHK1 and WEE1 expression is associated with poor outcome in independent cohorts of MM patients treated with high dose melphalan chemotherapy or anti-CD38 immunotherapy. Combined targeting of Chk1 and Wee1 demonstrates synergistic toxicities on MM cells and was associated with higher DNA double-strand break induction, as evidenced by an increased percentage of γH2AX positive cells subsequently leading to apoptosis. The therapeutic interest of Chk1/Wee1 inhibitors' combination was validated on primary MM cells of patients. The toxicity was specific of MM cells since normal bone marrow cells were not significantly affected. Using deconvolution approach, MM patients with high CHK1 expression exhibited a significant lower percentage of NK cells whereas patients with high WEE1 expression displayed a significant higher percentage of regulatory T cells in the bone marrow. These data emphasize that MM cell adaptation to replicative stress through Wee1 and Chk1 upregulation may decrease the activation of the cell-intrinsic innate immune response. Our study suggests that association of Chk1 and Wee1 inhibitors may represent a promising therapeutic approach in high-risk MM patients characterized by high CHK1 and WEE1 expression.

New agents-based induction chemotherapy followed by autologous stem cell transplantation and maintenance treatment strategy for multiple myeloma: a single center retrospective study of 300 cases

Objective: To examine the survival and influential factors of an integrated approach of novel agents, autologous hematopoietic stem cell (auto-HSCT) , and maintenance therapy in patients with multiple myeloma (MM) patients from a single center over the past 15 years. Methods: In our center, 300 MM patients who received an integrated strategy of new agents, auto-HSCT, and maintenance therapy over 15 years were retrospectively and prospectively analyzed. Results: The complete remission rates (CR) and ≥very good partial remission rates (VGPR) following induction therapy, transplantation, and maintenance therapy were respectively 35.3% and 55.2% , 72.4% and 80.0% , 89.2% , and 93.4% . When compared to patients receiving double-drug induction, the ≥VGPR and ORR of patients receiving triple-drug induction were improved. No difference existed in CR, ≥VGPR, and ORR between the PAD (bortezomib + liposome doxorubicin+ dexamethasone) and RAD (lenalidomide + liposome doxorubicin + dexamethasone) regimens, but the benefits speed differed. The negative rate of flow minimal residual disease following induction, transplantation, and maintenance was 18.8% (54 cases) , 41.4% (109 cases) , and 58.7% (142 cases) , respectively. The median time to progress (TTP) was 78.7 months and the median overall survival (OS) was 109 months. The median TTP for RISS-Ⅰ-Ⅲ patients were 111.8 months, 77.4 months, and 30.6 months, and the median OS was 118.8 months, 91.4 months, and 48.5 months, respectively. At various points during treatment, the TTP and OS of patients obtaining CR and MRD negative were longer than those of patients who did not obtain CR and MRD negative. TTP was noticeably shorter in high-risk cytogenetic patients compared to standard-risk patients even when CR was acquired during induction. There was no difference in TTP between patients with high-risk cytogenetics and those with standard-risk cytogenetics if MRD negative was acquired during induction. According to a multivariate analysis, the R-ISS stage was a poor predictor of TTP and OS at various treatment intervals. Therapeutic effectiveness was a newly independent prognostic factor following treatment. Conclusion: A median survival of almost 10 years is possible for MM patients who receive an integrated strategy of induction regimens followed by auto-HSCT and maintenance therapy, which significantly improves prognosis. However, this approach did not significantly benefit high-risk cytogenetic MM patients.

Advanced Risk Stratification in Multiple Myeloma Beyond Traditional FISH: The First Prospective Real-World Evidence for SKY92 Gene Expression Profiling

Background The prognosis of multiple myeloma (MM) has been significantly improved in the last decades. However, high-risk (HR) MM still suffers from early disease progression. Therefore, identifying HR MM patients may be a crucial step to improve the prognosis of this patient group. To date, fluorescence in situ hybridization (FISH) is the most commonly used method for risk stratification in MM. In recent years, gene expression profiling (GEP) approaches, e.g. SKY92, have been developed for detection of HR MM patients. Currently, real-world data of MM risk stratification applying SKY92 alone or in addition to traditional FISH are still missing. In this study, we provide the first prospective real-world evidence of the prognostic value of the combined SKY92 and FISH risk stratification in newly diagnosed (ND) and relapsed/refractory (RR) MM. Methods We prospectively collected bone marrow (BM) samples and clinical data of 147 MM patients. Cytogenetics were analyzed on purified CD138 positive MM cells by FISH, and HR cytogenetics was defined applying the Revised International Staging System (del(17p), t(4;14), and t(14;16)) (Palumbo et al, J Clin Oncol. 2015). SKY92 risk classification was determined with MMprofiler gene expression assay (Kuiper et al, Leukemia 2012). In addition, whole genome sequencing (WGS) was performed, and 150 bp paired-end sequences were generated on the NovaSeq6000 sequencing instruments (Illumina) with 100x coverage. Structural variations (SV) were called using Manta, and copy number variations (CNV) calling was performed with GATK4. Strelka2 variant caller was used for single nucleotide variations (SNV). Variants with global population frequencies >1% were excluded. Results We included 147 patients in our study (NDMM: n=51, RRMM: n=96). SKY92 classification was available for 121 (82.3%) patients. The median follow-up time was 344 days. HR SKY92 was significantly enriched in RRMM (40/76) compared with NDMM (6/45) (P<0.0001). RRMM patients with HR SKY92 showed significantly shorter progression free survival (PFS) (P<0.0001) and overall survival (OS) (P=0.0004) than standard-risk (SR). In NDMM, HR SKY92 also indicated a significantly inferior PFS (P=0.001) in comparison with SR. Of note, samples of 26 (17.7%) patients did not meet the SKY92 quality control criteria due to low BM infiltration (median BM infiltration: 25% vs 55% in other patients, P=0.038). We then combined the SKY92 classification with cytogenetic analyses by FISH, which were performed in 99 patients (NDMM: n=33; RRMM: n=66). We noticed a discrepancy between both risk stratification systems, with 44 (44.4%) and 42 (42.4%) patients being classified as HR by SKY92 and FISH, respectively. In total, 24 (36.3%) RRMM patients and 6 (18.2%) NDMM patients displayed HR in both SKY92 and FISH (double-HR) (Figure). Regarding survival outcome in RRMM, double-HR patients showed the worst PFS (P<0.0001) and OS (P<0.0001) when compared to the other patients (Figure). Noteworthy, RRMM patients with only HR SKY92 demonstrated significantly worse PFS than those with only HR FISH (P=0.007). In addition, double-HR presented a negative prognostic factor for PFS in NDMM (P=0.01). To further elucidate the discrepancy between FISH and SKY92 classifications, we performed WGS in 12 patients who exhibited either HR SKY92 only (n=9) or HR FISH only (n=3). Interestingly, 1 patient with bi-allelic TP53 inactivation (deletion + mutation) and 2 patients harbouring t(4;14) or t(14;16) were determined as SR by SKY92 but as HR by FISH; all the 3 patients were still in remission at the time point of data cut-off (July 2022). Moreover, we found abnormalities at chromosome 1 in 7 out of the 9 patients with HR on SKY92 only (gain1q21: n=6; del1p32: n=2) and, notably, gain1q21 and del1p32 were detected only by WGS in 3 and 2 patients, respectively. The remaining 2 patients did not show any known HR SV/CNV/SNVs, suggesting that HR MM may be associated with some other mechanisms, e.g. epigenetic modifications. Conclusion Altogether, risk classification using SKY92 performed well for detecting HR MM in the clinical routine. Here, we provide the first prospective real-world evidence that advanced risk stratification combining SKY92 and FISH may help to identify the "highest-risk" MM, which requires intensive anti-MM therapy and close monitoring in the course of the disease. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal