Subset Of Multiple Myeloma Patients Research Articles

G-Quadruplex Forming DNA Sequence Context Is Enriched around Points of Somatic Mutations in a Subset of Multiple Myeloma Patients.

Multiple myeloma (MM) is the second most common hematological malignancy, which remains incurable despite recent advances in treatment strategies. Like other forms of cancer, MM is characterized by genomic instability, caused by defects in DNA repair. Along with mutations in DNA repair genes and genotoxic drugs used to treat MM, non-canonical secondary DNA structures (four-stranded G-quadruplex structures) can affect accumulation of somatic mutations and chromosomal abnormalities in the tumor cells of MM patients. Here, we tested the hypothesis that G-quadruplex structures may influence the distribution of somatic mutations in the tumor cells of MM patients. We sequenced exomes of normal and tumor cells of 11 MM patients and analyzed the data for the presence of G4 context around points of somatic mutations. To identify molecular mechanisms that could affect mutational profile of tumors, we also analyzed mutational signatures in tumor cells as well as germline mutations for the presence of specific SNPs in DNA repair genes or in genes regulating G-quadruplex unwinding. In several patients, we found that sites of somatic mutations are frequently located in regions with G4 context. This pattern correlated with specific germline variants found in these patients. We discuss the possible implications of these variants for mutation accumulation and specificity in MM and propose that the extent of G4 context enrichment around somatic mutation sites may be a novel metric characterizing mutational processes in tumors.

Optimizing Autologous Stem Cell Transplantation in Multiple Myeloma: The Impact of Intensive Chemomobilization

Most transplant-eligible multiple myeloma (MM) patients undergo autologous peripheral blood stem cell collection (PBSC) using G-CSF with on-demand plerixafor (G±P). Chemomobilization (CM) can be used as a salvage regimen after G±P failure or for debulking residual tumor burden ahead of autologous peripheral blood stem cell transplantation (ASCT). Prior studies utilizing cyclophosphamide-based CM have not shown long-term benefit. At our center, intensive CM (ICM) using a PACE- or HyperCVAD-based regimen has been used to mitigate "excessive" residual disease based on plasma cell burden or MM-related biomarkers. Given the lack of efficacy of non-intensive CM, we sought to determine the impact of ICM on event-free survival (EFS), defined as death, progressive disease, or unplanned treatment escalation. We performed a retrospective study of newly diagnosed MM patients who collected autologous PBSCs with the intent to proceed immediately to ASCT at our center between 7/2020 and 2/2023. Patients were excluded if they underwent a tandem autologous or sequential autologous-allogeneic transplant, had primary plasma cell leukemia, received non-ICM treatment (i.e., cyclophosphamide and/or etoposide), or had previously failed G±P mobilization. To appropriately evaluate the impact of ICM among those who potentially could have received it, we utilized a propensity score matching (PSM) approach whereby ICM patients were compared to a cohort of non-CM patients matched on pre-ASCT factors most strongly associated with the receipt of ICM. Of 451 patients identified, 61 (13.5%) received ICM (PACE-based, n=45; hyper-CVAD-based, n=16). Post-ICM/pre-ASCT, 11 patients (18%) required admission for neutropenic fever and/or infection. Among 51 evaluable patients, the overall response rate was 31%; however, 46 of 55 evaluable patients (84%) saw a reduction in M-spike and/or involved FLCs. Among those evaluated with longitudinal peripheral blood flow cytometry (n=8), 5 patients (63%) cleared circulating blood PCs post-ICM. Compared to patients mobilized with non-CM, ICM patients collected a slightly greater median number of CD34+ cells (10.8 vs 10.2 × 10⁶/kg, p=.018). The median follow-up was 30.6 months post-ASCT. In a PSM multivariable analysis, ICM was associated with significantly improved EFS (HR 0.30, 95% CI 0.14 to 0.67, p=.003), but not improved OS (HR 0.38, 95% CI 0.10 to 1.44, p=.2). ICM was associated with longer post-ASCT inpatient duration (+4.1 days, 95% CI, 2.4 to 5.8, p<.001), more febrile days (+0.96 days, 95% CI 0.50 to 1.4, p<.001), impaired platelet engraftment (HR 0.23, 95% CI 0.06 to 0.87, p=.031), more bacteremia (OR 3.41, 95% CI 1.20 to 9.31, p=.018), and increased antibiotic usage (cefepime: +2.3 doses, 95% CI 0.39 to 4.1, p=.018; vancomycin: +1.0 doses, 95% CI 0.23 to 1.8, p=.012). ICM was independently associated with improved EFS in a matched analysis involving MM patients with excessive disease burden at pre-ASCT workup. This benefit came at the cost of longer inpatient duration, more febrile days, greater incidence of bacteremia, and increased antibiotic usage in the immediate post-ASCT setting. Our findings suggest that ICM could be considered for a subset of MM patients, but its use must be weighed carefully against additional toxicity.

Distinct phenotype of neutrophil, monocyte, and eosinophil populations indicates altered myelopoiesis in a subset of patients with multiple myeloma.

Hematologic malignancies, including multiple myeloma (MM), promote systemic immune dysregulation resulting in an alteration and increased plasticity of myeloid cell subsets. To determine the heterogeneity of the myeloid cell compartment in the peripheral blood of patients with MM, we performed a detailed investigation of the phenotype and function of myeloid subpopulations. We report that a subset of MM patients exhibits a specific myeloid cell phenotype indicative of altered myelopoiesis characterized by significant changes in the properties of circulating granulocytic, monocytic, and eosinophilic populations. The subset, referred to as MM2, is defined by a markedly elevated level of CD64 (FcγRI) on the surface of circulating neutrophils. Compared to healthy controls or MM1 patients displaying intermediate levels of CD64, neutrophils from MM2 patients exhibit a less differentiated phenotype, low levels of CD10 and CXC chemokine receptor 2 (CXCR2), increased capacity for the production of mitochondrial reactive oxygen species, and an expansion of CD16neg immature neutrophil subset. Classical and patrolling monocytes from MM2 patients express elevated levels of CD64 and activation markers. MM2 eosinophils display lower levels of C-C Chemokine receptor 3 (CCR3), Toll-like receptor 4 (TLR4, CD284), and tissue factor (TF, CD142). The MM2 (CD64high) phenotype is independent of age, race, sex, and treatment type. Characteristic features of the MM2 (CD64high) phenotype are associated with myeloma-defining events including elevated involved/uninvolved immunoglobulin free light chain (FLC) ratio at diagnosis. Detailed characterization of the altered myeloid phenotype in multiple myeloma will likely facilitate the identification of patients with an increased risk of disease progression and open new avenues for the rational design of novel therapeutic approaches.

External Validation of the Saved Score for Venous Thromboembolism Risk Stratification in Patients with Multiple Myeloma Receiving Immunomodulatory Drugs

Background: High thrombogenicity is a well-established phenomenon in multiple myeloma (MM) and can lead to high morbidity and mortality. The risk of venous thromboembolism (VTE) is particularly elevated in the subset of MM patients treated with immunomodulatory drug (IMiD) containing regimens. The National Comprehensive Cancer Network (NCCN) and International Myeloma Working Group (IMWG) have implemented VTE risk assessment models, and recommend prophylactic anticoagulation based on individual patient, disease, or treatment-related factors specifically for MM patients receiving IMiD. With the recent incorporation of the SAVED (PMID 31319391) scoring system in the NCCN guidelines for VTE risk stratification, we sought to externally validate this risk assessment model and evaluate its predictive ability in our MM patient cohort. Methods: We retrospectively evaluated 501 adult patients with either newly diagnosed or relapsed/refractory MM who were newly initiated on an IMiD-based regimen between 2010 and 2019. Patients already on therapeutic or prophylactic anticoagulation were excluded from the study (aspirin was allowed). The SAVED score was calculated at IMiD initiation according to the following five parameters: surgery within last 90 days (+2), Asian race (-3), prior VTE history (+3), age ≥80 years (+1), dexamethasone use (+2 for high dose [>160 mg/30-day], or +1 for standard dose [120-160 mg/30-day]). Patients with SAVED scores of <2 and ≥2 were considered of low and high risk, respectively, for developing VTE after IMiD initiation. Time to VTE was estimated by Kaplan-Meier failure method. Logistic regression model was used to test components of the SAVED score with 1-year VTE status. Model discrimination was assessed via c statistic by calculating the area under the receiver operating characteristic curve (AUC) with bootstrapped 95% confidence interval (CI). Results: The median age of our cohort at the IMiD initiation was 64 years (range 34-89); 264 (53%) patients were male and 378 (75%) were white. IgG and IgA heavy chain subtypes were present in 280 (56%), and 105 (21%) patients respectively, whereas 110 (22%) patients had light chain disease only. The International Staging System (ISS) was I, II, III, and unknown for 161 (32%), 149 (30%), 136 (27%), and 55 (11%) of patients, respectively. Most patients (94%) received lenalidomide, whereas 14 (3%) and 15 (3%) patients were treated with thalidomide and pomalidomide, respectively. The distribution of SAVED covariates is shown in Table 1. The cumulative incidence of VTE at 12-month after IMiD initiation for the entire cohort was 16% (95% CI: 13-20%). Among the 84 patients who developed new VTE after IMiD initiation, 60 (71%) had lower extremity deep venous thrombosis (DVT), 10 (12%) had pulmonary embolism, 9 (11%) had both DVT and PE, and 5 (6%) had upper extremity DVT. The SAVED score stratified 112 patients (22%) as high-risk and 389 patients (78%) as low-risk (similar proportion as the original derivation study). Cumulative incidence of VTE at 6 and 12 months was 31% and 41% in the high-risk group, versus 6% and 9% in the low-risk group, respectively (Figure 1). Each point of SAVED score increase was associated with significantly higher chance of VTE within 1-year post IMiD initiation with excellent discrimination (odds ratio = 2.10 [95% CI 1.71-2.56], c statistic (AUC) = 0.76 [95% CI 0.70-0.81]). Conclusion: Our results successfully externally validated the SAVED model to predict IMiD-associated VTE in a real-life cohort of patients with MM. Patients with SAVED score of ≥2 had significantly higher incidence for VTE. This indicates that more aggressive pharmacologic thromboprophylaxis is needed in this specific patient population. The SAVED score outperformed several other similar VTE prediction models that were also validated in the current cohort (c statistic=0.68 for IMPEDE [PMID: 33997961] and 0.62 for PRISM [PMID: 35772005]). In contrast to other risk models that focused on all patients with MM, the SAVED score was derived and now validated specifically for patients starting IMiD as this it is the only cancer population with a strong guideline recommendation for ambulatory thromboprophylaxis. Future prospective validation would be useful to determine the predictive nature of SAVED while evaluating safety and efficacy of thromboprophylaxis with direct oral anticoagulant in high-risk patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Cellular Immunotherapies for Multiple Myeloma: Current Status, Challenges, and Future Directions.

Multiple myeloma (MM) remains incurable due to relapse, although the use of proteasome inhibitors, immunomodulatory drugs, CD38-targeting antibodies, and autologous stem cell transplantation (auto-SCT) significantly improve the clinical outcomes of patients with newly diagnosed MM. In recent years, the introduction of chimeric antigen receptor T-cell (CAR T-cell) therapy has brought hope to patients with refractory and relapsed MM. The graft-versus-myeloma effect of allogeneic SCT provides the possibility for curing a subset of MM patients. In this review, we summarize the recent advances and challenges of cellular immunotherapies for MM, focusing on auto-SCT, allogeneic SCT, and CAR T-cell approaches. We also discuss future directions, and propose a specific algorithm for cellular therapies for MM and probability of minimal residual disease-directed therapy.

TRIM33 Loss in Multiple Myeloma Impairs the DNA Damage Response Resulting in Sensitivity to PARP and ATR Inhibitors

IntroductionChromosomal instability is a hallmark of Multiple Myeloma (MM), with most patients displaying cytogenetic abnormalities which can arise due to DNA damage response (DDR) defects. TRIM33 is an E3 ligase and transcriptional co-repressor located on chromosome 1p13.2, a region frequently deleted in MM. Previous studies have shown that TRIM33 plays a role in the DDR and can regulate chromosomal stability, but its precise function remains unknown. In this study we investigated the impact of TRIM33 loss in MM on genomic stability and DDR pathways and whether this could be exploited therapeutically.MethodsThe CoMMpass dataset (IA15 release) was screened to identify patients with copy number (CN) loss of TRIM33 and this was correlated with overall survival (OS) and structural variants. TRIM33 shRNA knockdown models were established in JJN3 and U266 cells. The effect on DDR signalling was determined by western blotting and immunofluorescence. The Selleckchem DNA Damage/Repair Compound Library was screened on the JJN3 model in a high-throughput manner using the CellTox™ Green cytotoxicity assay. Validation of selected compounds was performed using CellTiter® Glo viability assay or clonogenic assays. Combination indices (CI) were calculated using CompuSyn software.ResultsData on CN, OS and structural variants were available for 730 newly diagnosed MM patients and of these, 69 (9.5%) were identified to have a CN loss of TRIM33. These patients have poorer OS compared to those without TRIM33 loss (52.3 months vs 72.6 months; p<0.0001). Moreover, they exhibit a significantly higher median number of structural variants (deletions, duplications, inversions, and translocations; 38 vs 26; p<0.0001), indicative of increased chromosomal instability.Our data in MM cell lines has shown that TRIM33 is rapidly recruited to chromatin within 5 minutes of induced DNA damage. TRIM33 knockdown led to an increase in 53BP1 foci formation and endogenous γH2AX (P<0.001) indicating unrepaired DNA double-strand breaks (DSBs) typical of a DDR defect. In response to these DSBs both ATM and ATR kinases were activated as demonstrated by increased pKAP1 Ser824 and pCHK1 Ser345 respectively (p<0.001). Additionally, we observed a reduction in RAD51 (p<0.05) indicative of a potential defect in the DSB repair pathway homologous recombination (HR).To identify therapeutic vulnerabilities relating to TRIM33 loss, we performed a high-throughput screen to assess sensitivity to 160 unique DNA damaging compounds. TRIM33 knockdown cells exhibited increased sensitivity to 27 compounds across a range of drug classes. Additional studies confirmed that compared to control cells, TRIM33 knockdown sensitized cells to the PARP inhibitor Olaparib and ATR inhibitors BAY-1895344 and VE-821. Further investigation with VE-821 demonstrated that whilst treatment induced PARP cleavage and DSBs in both control and knockdown cells within 48 hours, knockdown cells exhibited significantly more pCHK1 Ser345 inhibition (p<0.01). Furthermore, combining VE-821 with bortezomib yielded synergistic effects in TRIM33 knockdown cells across a range of doses (CI range 0.57-0.9) while no synergy was observed in control cells (CI>1 for all combinations).ConclusionWe have identified a subset of MM patients with TRIM33 loss who display high-risk disease characterized by chromosomal abnormalities and defective DDR. Alongside this we have identified PARP and ATR inhibitors as therapeutic vulnerabilities in cell line models of TRIM33 loss. Moreover, we demonstrate that ATR inhibition increases the efficacy of bortezomib in TRIM33 knockdown cells. Further investigation into these compounds could lead to novel therapies for patients with TRIM33 loss. DisclosuresNo relevant conflicts of interest to declare.

Real-World Outcomes for Standard-of-Care Treatments in Patients with Relapsed/Refractory Multiple Myeloma

Background: With the advent of immunomodulatory agents (IMiDs), proteasome inhibitors (PIs) and, more recently, anti-CD38 monoclonal antibodies (mAbs), prognosis of patients with multiple myeloma (MM) has improved considerably. Unfortunately, even with these 3 major MM drug classes, most patients ultimately relapse and require further therapy. There remains an incomplete understanding of how patients who have received extensive therapy and with relapsed/refractory multiple myeloma (RRMM) are treated in routine clinical practice, as no standard-of-care exists for these patients, and what the outcomes are in this real-world setting.Objective: This study aims to evaluate the outcomes of patients with triple-class (IMiD, PI and anti-CD38 mAb) and triple-line exposed RRMM using real-world data from patients in Belgium.Methods: A multicenter, observational study, involving 7 non-academic and academic Belgian centers, was conducted based on a retrospective chart review of adult RRMM patients who started subsequent treatment from March 2017 through May 2021 after having received ≥3 lines of therapy including at least an IMiD, a PI, and anti-CD38-directed therapy (tri-exposed). Data were captured in an electronic case report form (Castor EDC). Patients with an ECOG performance status of ≥2, who received prior CAR-T treatment or prior BCMA-targeted therapy, or with a known active or prior history of CNS involvement (or with clinical signs thereof), were excluded. All treatment lines initiated after becoming eligible were used in the analysis. Specifically, all treatment lines for patients meeting the eligibility criteria more than once in their entire follow-up were included as separate observations, with date of treatment initiation as specific baseline for each treatment line. Cox proportional hazards models were fitted to explore the prognostic value with Overall Survival (OS), Progression Free Survival (PFS), and Time to Next Therapy (TTNT).Results: A total of 112 patients with 237 eligible treatment lines were included in the analysis; median follow-up was 16.6 months. In 45% of the initiated treatment lines, patients were refractory to 4 or 5 therapies, 62% had received ≥5 prior lines, 22% had extramedullary disease and in 48% of observations the time to progression in prior line was shorter than 4 months. After patients became tri-exposed, more than 50 unique treatment regimens were initiated, with the following being the most common: carfilzomib + dexamethasone (14%), pomalidomide + dexamethasone + chemotherapy (8%), and ixazomib + lenalidomide + dexamethasone (6%). Additionally, 4% of included observations were exposed to anti-BCMA agents. Overall, the following treatment classes were the most frequently started: PI only (19%), PI + IMiD combinations (17%), and regimens including anti-CD38 antibodies (15%). Median OS was 9.79 months [95% CI: 7.79; 12.22], median PFS was 3.42 months [95% CI: 2.79; 4.27], median TTNT was 3.61 months [95% CI: 3.09; 4.57]. Higher refractory status (p<0.001), being male (p=0.001), older age (p<0.001), shorter duration of prior lines (p<0.001), shorter time to progression in prior line (p=0.025), and higher LDH levels (p<0.002) were prognostic for worse outcomes for both OS (Figure 1) and PFS.Conclusions: This retrospective chart review of patients with tri-exposed RRMM in Belgium shows that real-world outcomes in terms of OS, PFS and TTNT are poor for these patients, with a median OS of <10 months. A wide variety of treatment regimens used in clinical practice confirm the absence of a clear standard-of-care in this patient population. The literature also confirms that these poor outcomes observed in Belgium, for this subset of MM patients, are similar in other countries. These real-world data highlight the high unmet medical need in this patient population and critical need for new and effective treatment options.MD and MCV contributed equally to this work. [Display omitted] DisclosuresDelforge: Amgen, Celgene, Janssen, Sanofi: Honoraria, Research Funding. Vekemans: Amgen: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; BMS-Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen Pharmaceutica: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees. Depaus: Takeda: Consultancy; Novartis: Consultancy; Janssen: Consultancy; Celgene: Consultancy. Meuleman: iTeos Therapeutics: Consultancy. Strens: Realidad bvba: Consultancy. Van Hoorenbeeck: Janssen: Current Employment. Moorkens: Janssen-Cilag: Current Employment. Diels: Janssen: Current Employment. Ghilotti: Janssen-Cilag SpA, Cologno Monzese, Italy: Current Employment. Dalhuisen: Janssen: Current Employment. Vandervennet: Janssen: Current Employment.

P-080: Single-cell transcriptomic analysis of bone marrow NK cells reveals loss of activated cytotoxic NK cells in Multiple Myeloma

<h3>Background</h3> Multiple Myeloma (MM) disease progression and therapy response are influenced by cues of the microenvironment including tumor control by a cytotoxic immune response. Natural killer (NK) cells are notable mediators of the cytotoxic immune response to MM, and important effector cells in recent immune-mediated therapies. NK cells are drivers of antibody-dependent-cellular cytotoxicity in therapies based on anti-CD38 monoclonal antibodies such as Daratumumab. Classically, NK cells are divided based upon CD56 expression into a cytokine-producing CD56bright subset, releasing cytokines such as IFN-γ, TNF-a and GM-CSF, and a cytotoxic CD56dim subset. However, accumulating evidence suggests much larger heterogeneity in the NK cell compartment and modulation of these NK cell subsets could impact response to NK cell-driven immunotherapies. Here, we used single-cell RNA sequencing to investigate the heterogeneity and MM-driven alterations of the NK cell compartment in the bone marrow of newly diagnosed MM patients undergoing first-line Daratumumab-containing therapy. <h3>Methods</h3> We performed single-cell RNA sequencing of the CD38+ and CD38- fractions of viably frozen bone marrow aspirates from 19 newly diagnosed MM patients and 5 non-cancer control patients. NK cells were identified in silico by transcription of KLRF1, KLRD1, GNLY and NKG7 resulting in a single-cell transcriptomic dataset of 30,373 NK cells from MM patients and 8,865 NK cells from control patients. <h3>Results</h3> After integration of the datasets, bone marrow NK cells formed eight distinct transcriptomic clusters. Conventional CD56bright and CD56dim NK-cells were identified by increased transcription of GZMK or GZMB, respectively. The GZMK+CD56bright NK cells contained both a cluster of naive and a cluster of activated NK cells. The GZMB+CD56dim NK cells consisted of 5 subclusters. To identify MM-induced alterations in NK cell subsets we compared GZMK+CD56bright vs GZMB+CD56dim cluster composition and distribution between control and MM patients. Control bone marrow was dominated by GZMB-transcribing cytotoxic CD56dim NK cells, represented by a low cytokine-producing GZMK+CD56bright vs cytotoxic GZMB+CD56dim ratio. In contrast, MM bone marrow was characterized by heterogeneity in the ratio of cytokine-producing GZMK+CD56bright vs cytotoxic GZMB+CD56dim NK cells. A subset of patients presented with a complete reversal of this ratio. This altered composition was due to a loss of cytotoxic GZMB+CD56dim NK cells, and more specifically a loss of NK cells with a transcriptome suggesting recent activation. <h3>Conclusion</h3> Here we present a transcriptomic overview of NK cells in MM bone marrow at the single-cell level. A subset of MM patients has a loss of activated cytotoxic GZMB+CD56dim NK cells, suggestive of reduced cytotoxic anti-tumor responses. Current analyses are focused on clinical implications of NK cell alterations in response to Daratumumab-based therapies.

NS-64: Lessons learned: patient management and outcomes of Multiple Myeloma patients during the COVID-19 pandemic in New York City

New York City (NYC) was an epicenter of COVID-19 pandemic in the United States during spring 2020. During March–May 2020, approximately 203,000 confirmed COVID-19 cases were reported among NYC residents including 54,211 (26.6%) known to have been hospitalized and 18,679 (9.2%) who died. NYC had a higher confirmed coronavirus case volume than any country in the world, besides the United States. Here, we discuss how advanced practitioners Providers (APPs) and Health Care Support Staff in the Multiple Myeloma (MM) program at Mount Sinai Hospital in NYC modified patient care guidelines during the COVID-19 pandemic in an effort to balance the need for social distancing and optimizing patient care. Cognizant of these key factors during the pandemic, oncology APPs, nurses, physicians, and staff at Mount Sinai convened to determine how patients should be best managed. Important considerations in addition to remission status and type of therapy (IV/SC or oral) included the patient’s performance status and chemotherapy oral adherence. Based on these, the group determined: (1) Newly diagnosed patients or relapsed patients with significant cytopenias who need parenteral therapy should continue with the current therapy, but the number of visits should be decreased to limit exposure to other HCPs. If the patient has had a good response to treatment, consider changing to an oral regimen. (2) Patients in remission (complete remission, very good partial remission) should consider switching therapy to an oral regimen (immunomodulatory agents, proteasome inhibitors, dexamethasone, and/or Histone deacetylase inhibitors). (3) Stem cell harvest and stem cell transplant should be delayed if the patient continues to respond to the current therapyOutcomesDuring this time we had 58 MM patients diagnosed with COVID-19, 36 were hospitalized and 22 were managed at home. The median age was 67 years; 52% of patients were male and 63% were non-White. Hypertension (64%), hyperlipidemia (62%), obesity (37%), diabetes mellitus (28%), chronic kidney disease (24%), and lung disease (21%) were the most common comorbidities. In the total cohort, 14 patients (24%) died. Older age (>70 years), male sex, cardiovascular risk, and patients not in complete remission (CR) or stringent CR were significantly (p<0.05) associated with hospitalization. Although several demographic factors and comorbidities increased the risk of hospitalization and mortality, myeloma therapy did not influence outcomes. In fact, survival was comparable to the overall population of New York during the pandemic. Our data supports the need to maintain proactive management of MM patients by balancing their need for therapy with the increased risk of hospitalization and death in a subset of MM patients with COVID-19.

P-054: TRIM33 loss in Multiple Myeloma is associated with defective DNA repair and sensitivity to PARP inhibition

<h3>Background</h3> Chromosomal instability is a hallmark of Multiple Myeloma (MM) with most patients displaying cytogenetic abnormalities which can often act as prognostic indicators. Such abnormalities can arise due to defects in the DNA Damage Response (DDR). TRIM33 is an E3 ligase and transcription co-repressor located on chromosome 1p13.2, a region frequently deleted in MM. Previous studies have shown that TRIM33 is involved in PARP-dependent DDR and regulation of chromosomal stability. Here, we investigated the influence of TRIM33 loss in MM, focusing on its role in the DDR and whether this could be exploited therapeutically. <h3>Methods</h3> The CoMMpass dataset (IA15 release) was screened to identify patients with copy number (CN) loss of TRIM33 and this was correlated with survival, structural variants and common cytogenetic abnormalities. TRIM33 shRNA knockdown models were established in JJN3 and U266 cells for in vitro studies. Protein expression and interactions were assessed by co-immunoprecipitation, western blotting and/or immunofluorescence. Clonogenic survival assays were used to assess response to Olaparib. <h3>Results</h3> Previously we identified a subset of MM patients with TRIM33 loss and identified that these patients exhibit significantly more chromosomal structural variants (deletions, inversions, duplications and translocations) (p<0.0001) and a significantly poorer overall survival (p<0.0001). Additionally, we have determined the frequency of common recurrent primary and secondary cytogenetic abnormalities for these patients. No recurrent primary cytogenetic abnormalities were associated with TRIM33 loss. However, high-risk secondary chromosome 1 aberrations were associated with loss of TRIM33, both del(1p) and gain(1q) (p<0.0001 and p=0.0474 respectively). In vitro, TRIM33 knockdown resulted in increased formation of 53BP1 foci and increased γH2AX expression (p<0.001) indicating unrepaired DNA damage typical of a DDR defect. Following induced DNA damage using 2Gy irradiation (IR), TRIM33 is recruited to chromatin within 5 minutes, with levels returning to basal by 30 minutes. The chromatin remodelling enzyme ALC1 is known to regulate sensitivity to PARP inhibition and TRIM33 is required for its timely removal from sites of damage. TRIM33 transiently interacts with ALC1 within 15 minutes of 2Gy IR. TRIM33 knockdown did not affect ALC1 expression. However, knockdown did sensitize MM cells to the PARP inhibitor Olaparib reducing the IC50 from 1.7µM to 780nM. <h3>Conclusion</h3> Here, we show that a subgroup of MM patients have TRIM33 loss and these patients have high-risk disease and poor outcome. We show that TRIM33 is recruited to damaged chromatin where it regulates ALC1 activity. Therefore, TRIM33 loss results in a DDR defect leading to chromosomal abnormalities. However, TRIM33 loss-associated DDR defects can be exploited therapeutically using Olaparib which is currently approved for the treatment of BRCA1/2 mutated breast and ovarian cancers.

Bone marrow plasma level of decorin may be associated with improved treatment outcomes in a subset of multiple myeloma patients

Decorin is a small leucine-rich proteoglycan rich in extracellular matrix with potential antitumor activity. However, the role of decorin in hematological malignancies remains unclear, especially in the case of multiple myeloma (MM), a bone marrow (BM) stroma-dependent plasma cell neoplasm. We measured decorin levels in BM plasma samples from 270 patients with newly diagnosed MM (NDMM) using enzyme-linked immunosorbent assays. Patients were divided into high decorin (H-DCN, > 18.99ng/mL) and low decorin (L-DCN <9.76ng/mL) groups. Patients in the H-DCN group had more advanced-stage disease, including more osteolysis terms of higher levels of C-terminal telopeptides of type I collagen (0.69±0.55 vs. 0.49±0.36ng/mL; P=0.028), than those in the L-DCN group. Decorin levels correlated positively with hepatocyte growth factor (HGF) levels in BM plasma samples from NDMM patients (Pearson correlation coefficient, 0.226; P<0.001). Patients with low HGF (<0.79ng/mL) but high decorin levels (≥12.95ng/mL) had a higher treatment response rate (90.5% vs. 54.5%, respectively; P=0.015) and improved overall survival (not reached vs. 53 months; P=0.0148) than those with lower decorin levels (<12.95ng/mL). Multivariate analysis confirmed that a high decorin level was an independent predictive factor for treatment response and survival in patients with low HGF levels. Our findings suggest that decorin may exert protective effects in this subset of MM patients.

Vulnerability Variables Among Octogenarians Myeloma Patients: A Single-Center Analysis in 110 Patients

In the era of an aging population an increasing number of patients are diagnosed with multiple myeloma (MM) at an age of ≥80 years. The frailty of the octogenarian population as a result of coexisting comorbidities and age-related organ impairment is an additional management challenge and often excludes this population from enrollment in clinical trials. The aim of our study was to analyze the disease characteristics, frailty scores at diagnosis and treatment toxicity profile in relation to patient outcomes. All newly diagnosed symptomatic MM patients ≥80 years of age who received at least one dose of treatment were included in the study.Among 827 consecutive, newly diagnosed, symptomatic MM patients who were treated in the Department of Clinical Therapeutics (Athens, Greece) after 01/01/2000, 110 were ≥80 years old (13.3%); their median age at diagnosis was 83 years (range 80-92 years) and 55% were male. Performance status (PS) was 1-2 in 40% and 3-4 in 60% of the patients respectively. Anemia (Hb <10 g/dl) was present in 64.5% of patients, increased serum LDH (>250 IU/L) in 12%, hypercalcemia (calcium ≥11 mg/dL) in 19% and osteolytic bone lesions in 73% of patients. Serum creatinine was ≥2 mg/dl in 30% and eGFR (per CKD-EPI equation) was <60 ml/min/1.73m2 in 69% and in 36% of patients was <30 ml/min/1.73m2. Per ISS, 56% had stage-3, 35% had -2 and 9% had disease stage-1, respectively. Cytogenetics were available for 66 patients and high-risk cytogenetics were present in 9 (12%) patients.First line treatment was bortezomib-based in 30% of patients, IMiD-based in 62%, while 8% received conventional chemotherapy. Overall response to first line treatment (≥PR) was 62% (sCR 9%, CR 4%, VGPR 30% and PR 57%). At the time of diagnosis 41% of patients required hospitalization due to disease related complications, while 60% were hospitalized at least once during their disease course. Low eGFR was associated with need for hospitalization at the time of diagnosis (p<0.001). Treatment emerging toxicities were common; 63% of our patients experienced at least one grade ≥3 event and 14% had grade 5 events. Grade ≥3 hematological toxicity was reported in 24% and grade ≥2 neuropathy in 23% of patients; 28% of patients experienced at least one grade ≥ 3 infectious episode during their disease course.According to IMWG geriatric assessment, due to the age of the patients (≥80), all of them were categorized as frail. Using the G8 geriatric screening tool, 73% of patients had a score £ 11, per the CIRS-G scale 52% had score >17, and the Charlson Comorbidity Index was ≥2 in 49%; on the IADL scale 42% had a score £5 and on the ADL scale 18% scored £4 while the VES-13 score was ≥6 in 58% of patients.Median survival was 21 months (95% CI 9-33 months) and early mortality (at 2 months from treatment initiation) was 20%. Several factors were associated with inferior survival in univariate analysis, including female gender (p=0.042), ISS stage-3 (p=0.015), LDH ≥250 IU/L (p=0.001) and eGFR <30 ml/min/1.73m2 (p<0.001) at the time of diagnosis, while, the presence of anemia, hypercalcemia, high risk cytogenetics, and the use of bortezomib vs IMiDs-based therapy were not. Factors such as the need for hospitalization at the time of diagnosis (p<0.001) and the presence of grade 3 or 4 adverse events during the disease course (p=0.003) had a negative impact on survival. A higher score on the G8-screening tool (p=0.002) and VES13 (p=0.008) were also associated to shorter OS. However, VES13 was available only in 38 patients. In the multivariate analysis, eGFR <30 ml/min/1.73m2 (p=0.003) and LDH ≥250 IU/L (p=0.048) were independently associated with poor OS, while there was a trend for a significant association for the G8 geriatric screening tool (p=0.08) and need for hospitalization at the time of diagnosis (p=0.065).In conclusion, the octogenarian MM patient population is a distinct frail subset of MM patients and its management should be individualized considering both disease characteristics and functional status. Renal impairment at diagnosis and high LDH correlated independently with OS in this group of patients. The G8 and VES13 geriatric assessment tools can provide additional information. The need for hospitalization at diagnosis is an ominous sign. This vulnerable patient population needs detailed assessment, individualized treatment and intensive supportive care to avoid hospitalization due to complications and to reduce early mortality. DisclosuresTerpos:Genesis/Celgene: Honoraria, Other: DMC member, Research Funding; Takeda: Honoraria, Other: SC member; GSK: Honoraria; BMS: Honoraria; Amgen: Honoraria, Other: SC member, Research Funding; Abbvie: Honoraria; Janssen: Honoraria, Research Funding. Kastritis:Prothena: Honoraria; Janssen: Honoraria; Genesis pharma: Honoraria; Takeda: Honoraria; Amgen: Honoraria, Research Funding. Dimopoulos:Novartis: Consultancy, Honoraria; Amgen Inc, Celgene Corporation, Janssen Biotech Inc, Onyx Pharmaceuticals, an Amgen subsidiary, Takeda Oncology: Consultancy, Honoraria, Other: Advisory Committee: Amgen Inc, Celgene Corporation, Janssen Biotech Inc, Onyx Pharmaceuticals, an Amgen subsidiary, Takeda Oncology; Genesis Pharma: Research Funding.

CD33 Is Expressed in a Significant Subset of Multiple Myeloma Patients in the US and May Represent a Viable Therapeutic Target

Background: CD33 (sialic acid binding Ig-like lectin 3; SIGLEC3) is a type I transmembrane receptor glycoprotein that may function as a sialic acid-dependent cell adhesion molecule. CD33 is generally viewed as a marker of myeloid lineage, and its expression on lymphoid lineage cells is thought to be aberrant and rare, reported to be present in only 1% - 8% of non-myeloid samples, with estimated similarly low expression in multiple myeloma plasmocytes specifically. Some data indicate that CD33 may be expressed on multiple myeloma (MM) plasmocytes in a much larger proportion of MM patients than previously thought, and that when expressed it may confer worse prognosis. However, these data were obtained from established MM cell lines and small series of patient samples. For the first time, we have attempted to obtain data on CD33 expression on MM plasmocytes from patient samples in a large US library.Methods: Data from the IA9 release of the MMRF CoMMpass study was downloaded from the MMRF Researcher Gateway. The per patient visit clinical table was filtered to include only patient visits with numerical calls in the “CD33_detected” field, which indicates whether CD33 was detected on the surface of MM plasmocytes by flow cytometry. Data in the per patient visit clinical table was further filtered to include only “Baseline” and “Confirm Progression” visits. Any patient with a baseline visit was considered in the analysis of baseline, newly-diagnosed MM patients. Only patients with both a baseline and at least one confirm progression visit were considered in the analysis of patients with CD33 data at two or more timepoints. For each visit, the percent CD33+ plasma cell calls (CD33_PC_Percent) were assigned a bin as follows: <20%, 20-39%, 40-59%, 60-79%, 80-100% CD33 positive plasma cells. High expression of CD33+ was defined as CD33 detected in ≥40% of MM plasmocytes whereas very high expression was defined as CD33 detected in ≥60% of MM plasmocytes.Results: The IA9 CoMMpass release includes 995 samples that have corresponding CD33 data, representing 865 unique patients. Samples were collected at baseline for newly-diagnosed MM patients (n=865) and at relapse when patients had progression events (n=59 patients, 66 samples). Among newly-diagnosed MM patients at baseline 25% (n=219) of patient tumors expressed CD33 based on flow cytometry analysis, whereas 75% (n=646) did not. Among patients with CD33+ MM, the proportion of CD33+ plasmocytes detected were as follows: <20% cells: 11.0% of samples (n=24); 20%-39% of cells: 27.4% of samples (n=60); 40%-59% of cells: 20.5% of samples (n=45); 60%-79% of cells: 10.5% of samples (n=23); 80-100% of cells: 30.6% of samples (n=67). At baseline, 61.6% (n=135) of patients exhibited a high percentage of CD33+ plasmocytes (≥40% of MM plasmocytes expressing CD33) and 41% (n=90) of patients exhibited a very high percentage of CD33+ plasmocytes (≥60% of MM plasmocytes expressing CD33).Similarly to newly diagnosed patients, among the 59 patients that had samples available at baseline and at least one progression visit, 27.1% (n=16) were CD33+ at relapse. Plasmocytes from 62.7% (n=37) were CD33- at both baseline and progression, 20.3% (n=12) were CD33+ at both baseline and progression, 10.2% (n=6) were CD33+ at baseline and CD33- at progression, and 6.8% (n=4) were CD33- at baseline and CD33+ at progression. Among the 12 patients that were CD33+ at both baseline and progression, 58.3% (n=7) had very high CD33 at both timepoints. Of the other 5 patients, 25% (n=3) exhibited an increase in the percent of CD33+ plasmocytes, going from low to high (n=1), low to very high (n=1), and high to very high (n=1), whereas 16.7% (n=2) of patients exhibited a decrease in the percent of CD33+ plasmocytes, going from very high to high (n=1) or very high to low (n=1) at baseline versus relapse.Conclusions: These findings indicate that a substantial proportion of MM patients express CD33 on their myeloma plasmocytes, including relapsed patients. Therefore, CD33 may represent a viable target in multiple myeloma. Relevance of this pattern of CD33 expression may be further enhanced by the presence of a number of CD33 targeting agents already in clinical trials in acute myeloid leukemia, which could allow for a rapid translation into the clinical settings in multiple myeloma. One trial in relapsed and refractory multiple myeloma with a CD33 targeting radiolabeled monoclonal antibody is underway. DisclosuresLevy:Actinium Pharmaceuticals: Equity Ownership; Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Research Funding; Takeda: Consultancy, Speakers Bureau. Cicic:Actinium Pharmaceuticals: Employment, Equity Ownership.

Multiple myeloma: a review of atypical imaging features and other distinct plasma cell disorders that demonstrate similar imaging features.

Multiple osteolytic lesions are a well-recognized and typical imaging feature of multiple myeloma as well as several other plasma cell disorders. Given the high volume of imaging studies obtained of multiple myeloma patients, radiologists will likely encounter a subset of multiple myeloma patients with less common or "atypical" findings during their practice. These atypical findings include osteosclerotic lesions, extramedullary lesions, and amyloid deposition. Similar imaging findings that are considered atypical for multiple myeloma can also be detected in other plasma cell disorders that are distinct from multiple myeloma. For instance, POEMS syndrome is a distinct plasma cell disorder from multiple myeloma, but also can present with osteosclerotic lesions. This article reviews the atypical findings associated with multiple myeloma and also reviews other plasma cell disorders that can have a similar spectrum of imaging findings. Special attention is paid to the musculoskeletal imaging findings.

Strategies to Overcome High-Risk Multiple Myeloma.

Multiple myeloma (MM) patients with high-risk cytogenetics continue to have inferior outcomes despite recent advances in the treatment of MM. As defined by the International Myeloma Working Group, the presence of t(4;14), t(14;16), del(17p), t(14;20) and amplification of 1q are considered to be high-risk chromosomal abnormalities associated with poor survival. Despite the use of immunomodulatory agents, proteasome inhibitors, autologous stem cell transplantation, and anti-CD38 monoclonal antibodies, clinical trials of current therapies have not shown strong statistical evidence of being able to overcome the poor prognosis of high-risk disease. Novel treatment approaches are urgently needed to improve survival in this subset of MM patients.

Real World Experience of Daratumumab: Evaluating Lymphopenia and Adverse Events in Multiple Myeloma Patients.

Multiple myeloma (MM) is an incurable disease with a limited life expectancy of five years from diagnosis. Uncontrolled disease or infections are the main causes of mortality. Daratumumab, a monoclonal antibody against CD38, is approved to treat patients with MM. Its target, CD38, is expressed not only on MM cells but also on common lymphoid precursors and subsets of normal lymphocytes. Daratumumab-induced lymphopenia is common, but its clinical significance is understudied. In this study, we report the baseline characteristics, rates of severe lymphopenia, infections, and clinical trajectory of multiple myeloma patients (n = 100) treated with daratumumab-based regimens at the Ohio State University Comprehensive Cancer Center. We discover high rates of infections, hospital utilization, and severe lymphopenia and identify risks factors for severe lymphopenia, such as low pretreatment absolute lymphocyte count (ALC) values. Severe lymphopenia persists in 23% of patients, resulting in worst survival outcomes. Our data underline the importance of monitoring ALC and consider future use of prophylactic measures or alternative regimens in subsets of MM patients.

Identification of Molecular Mechanisms Responsible for the Development of Extramedullary Disease in Myeloma and Potential Novel Therapeutic Targets Using Transcriptomic and Exome Profiling

Introduction: Multiple myeloma (MM) is the second most common blood cancer, and despite recent treatment advances, the majority of patients will ultimately die due to progression. Extramedullary disease (EMD) is a less frequent manifestation of MM often occurring during the course of disease where tumor plasma cells become independent of bone marrow microenvironment and infiltrate other tissues and organs. The development of EMD is always considered a high-risk feature with poor prognosis. Aim: To identify molecular mechanisms responsible for the development of EMD using DNA and RNA sequencing and thus reveal potentially novel druggable targets. Currently, available anti-myeloma agents are not effective in patients with EMD. Thus it is critical to perform this kind of translational research that will ultimately lead to discovering new treatment strategies in this prognostically poor subset of MM patients. Methods: We collected a unique set of FACS/MACS sorted aberrant plasma cells from 4 freshly obtained EMD samples from relapse and their respective cryopreserved bone marrow samples from diagnosis (available in 3 cases). In the 3 longitudinal ND and EMD samples we analyzed somatic mutations in the whole-exome sequencing data (Illumina, Sure select V6) using a combination of Mutect2 and Strelka2. To analyze differential gene expression, we used Deseq2 R package and Salmon for read mapping and quantification and Cytospace for the pathway enrichment analysis. Besides, to investigate the effect of cryopreservation on the gene expression in longitudinal samples from diagnosis, we compared all 4 EMD samples with 5 fresh unrelated BM samples from ND patients with PET/CT confirmed lack of EMD. Results: Analysis of somatic mutations revealed an only partial overlap of mutated genes between ND and respective EMD sample (23% on average) and 1.49x increased number of variants in EMD. In our samples (ND and EMD), we identified 11 mutated myeloma drivers (Walker 2018) that mostly affected epigenetic processes and MAPK pathway, with the latter being mutated in all three patients in both stages. Interestingly, we identified two genes, both exclusively mutated only in 2/3 EMD samples and not mutated in any ND sample, DNAJC16 and HERC1. DNAJC16 is a member of heat shock proteins (Hsp40; Kampinga 2009). HERC1 is an E3 ubiquitin ligase that regulates p38 signaling and cell migration (Padrazza 2020), and thus, it represents a potentially interesting target for further analysis of EMD development. Differential expression analysis of 3 paired ND-EMD samples revealed 131 deregulated transcripts with the TNFSF9 (CD137L) ligand, HECW1 ubiquitin ligase, and UNC13C gene being top upregulated genes. Among the top downregulated were immunoglobulin genes and S1PR4 signaling receptor, MPO myeloperoxidase, and SLAMF1 (CD150) signaling lymphocytic activation molecule. Analysis of a larger cohort of fresh unpaired 5 ND and 4 EMD samples resulted in more distinct clusters of ND/EMD samples and the total number of 673 deregulated genes (Fig. 1). EMD downregulated genes mostly belong to cell adhesion, chemokine signaling pathway, and neutrophil activation process. Upregulated genes belong to proliferation, cell cycle, and DNA damage signaling pathways. Evaluation of currently available druggable targets revealed that only LAG3 (lymphocyte activation 3 protein) is significantly downregulated in EMD. For three upregulated genes (FGFR3, NTRK3, EZH2), we found interaction with potential cancer inhibitors (Erdafitinib, Entrectinib, Tazemetostat). Notably, the latter agent targeting EZH2 methyltransferase gene has been recently approved by the FDA for follicular lymphoma treatment, and clinical trials for MM are ongoing. Conclusion: Here, we report the preliminary analysis of the genomic and transcriptomic profile of EMD samples with their respective paired samples from the new diagnosis. The genomic alterations proved to be heterogeneous; though, we were able to identify 2 genes exclusively mutated in EMD samples. Moreover, deregulated transcriptome clearly separated ND and EMD stage in unrelated cohorts and pointed to rapid proliferation and increased DNA damage response pathways and pinpointed several potential drug targets. However, our findings warrant a more in-depth analysis on the bigger patient cohort. Acknowledgement: Work was supported by project ENOCH (No. CZ.02.1.01/0.0/0.0/16_019/0000868) Figure 1 Disclosures No relevant conflicts of interest to declare.

Translational Approach of Using Ex Vivo Cytotoxicity and Early Clinical Data to Predict Teclistamab Efficacious Therapeutic Range in Multiple Myeloma Patients

B cell maturation antigen (BCMA; CD269; TNFRSF17) is a surface receptor, expressed on benign and malignant plasma cells. The restricted expression of BCMA on B-cell lineage makes it an ideal target for multiple myeloma (MM) immunotherapy. BCMAxCD3 bispecific antibody (Teclistamab; JNJ-64007957) has been developed to recruit CD3+ T-cells to BCMA+ MM cells. Teclistamab demonstrated potent cytotoxicity in a murine xenograft model and against ex vivo primary MM cells. Teclistamab clinical starting dose in the first in human (FIH) study in relapsed refractory MM patients began at the minimal anticipated biological effect level (MABEL) of 0.3 µg/kg using the EC20 of the in vitro cytotoxicity assay in the purified T cells model with MM.1R cell line. Since the MABEL dose is usually low, it is important to have guidance on the expected therapeutic range in patients. The therapeutic concentrations of teclistamab were estimated from an ex vivo cytotoxicity assay in which samples from MM patients (frozen purified mononuclear cells from bone marrow) were treated with healthy human purified T cells (1:1 ratio) spiked with teclistamab and incubated for 48 hours. The mean and range of EC50 and EC90 values of the cytotoxicity endpoint were estimated. The pharmacokinetic (PK) data following the first cycle doses in the low dose cohorts in the FIH study in MM patients were modeled using 2-compartment model and simulated to predict the doses that will have average and trough serum teclistamab concentrations in the expected therapeutic range (between EC50 and EC90). The predicted doses with average serum concentrations between the mean EC50 (372 ng/mL) and mean EC90 (2287 ng/mL) range were validated with the observed clinical data showing positive response in 2 out of 4 patients at the 38.4 µg/kg intravenous treatment dose level and their average serum teclistamab concentration overlaid within the estimated therapeutic range. This was also confirmed with the teclistamab concentrations in bone marrow samples (measured in a subset of MM patients in the study) where they were found to superimpose in the estimated therapeutic range. In addition, simulations predicted the dose that will have trough levels above the maximum EC90 (6070 ng/mL), to account for patients with high tumor burden, at which the observed clinical data have shown promising response rate and has been selected to be the recommended subcutaneous phase 2 dose of 1500 µg/kg. In conclusion, ex vivo cytotoxicity assay with MM patients bone marrow samples is an informative tool to predict the efficacious serum concentration of immuno-oncology drugs in MM patients. PK modeling and simulations of MM patients' data in FIH study coupled with the ex vivo cytotoxicity estimates provided guidance on the expected recommended phase 2 dose during the escalation phase of the FIH study. Disclosures Girgis: Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Wang Lin:Johnson &amp; Johnson: Current Employment. Pillarisetti:Johnson &amp; Johnson: Current Employment. Banerjee:Janssen: Current Employment. Goldberg:Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Shetty:Johnson &amp; Johnson: Current Employment. Stephenson:Johnson &amp; Johnson: Current Employment, Current equity holder in private company. Hilder:Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Hanna:Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Smit:Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Adams III:Genmab: Current Employment. Sun:Johnson &amp; Johnson: Current Employment, Current equity holder in publicly-traded company. Infante:Janssen: Current Employment. Elsayed:Johnson &amp; Johnson: Current Employment. Sharma:Johnson &amp; Johnson: Current Employment.

POU2AF1 As a Master Regulator of Oncogenic Transcription Factor Networks in Myeloma

Background: Functional genomics studies based on CRISPR and shRNA have documented that multiple myeloma (MM) cells are preferentially dependent (compared to other neoplasias) on a series of TFs, including IKZF1 and IKZF3 (which are targeted by thalidomide derivatives) and others that are not amenable to degradation or small molecule inhibition. Transcriptional co-factors have been therapeutically targeted, for example, inhibitors of BRD4, a co-factor for pTEFB, can be used to down-regulate c-myc. Aim: To identify new transcriptional vulnerabilities in MM with an emphasis on transcriptional co-factors Methods: We integrated results from genome-scale studies using the AVANA library for loss-of-function by gene editing (in 19 MM lines) and the Calabrese library for CRISPR-mediated gene activation (in 5 MM lines) to identify critical transcriptional co-factors (co-TFs). RNA-Seq analysis was used to identify critical pathways affected by POU2AF1 activation and existing ChIP-Seq tracks in MM cells were reanalyzed. Results: POU2AF1 (OCA-B) was the most preferentially essential TF co-factor in MM cell lines vs. non-MM and one of top genes which, upon CRISPR activation in genome-scale studies, increased MM cell fitness in vitro. We further confirmed the role of this gene using focused libraries of sgRNAs against POU2AF1 in vitro and in an in vivo model of MM cell growth in bone marrow-like scaffolds "functionalized" with humanized mesenchymal bone marrow stromal cells to simulate the human BM. CRISPR activation of POU2AF1 is associated with increased MM cell growth. RNA-Seq of POU2AF1 activation in LP1 cells a transcriptional program characterized by upregulation of other genes that are preferentially essential for MM including PRDM1, SUPT7L, UBE2G2 and TSC1; broad-spectrum oncogenic dependencies (e.g KRAS) and genes known or proposed to be involved in the pathophysiology of MM or other neoplasias (e.g. RUNX2, FGFR3, SMO, CREB5, TNFRSF13B, MEF2D, PCGF2). POU2AF1 overexpression was also associated with down-regulation of CDKN1C; of MHC class II molecules and their transcriptional activator CIITA, suggesting that POU2AF1 activation could also contribute to increased MM growth in vivo by allowing escape from immune surveillance. ATAC-Seq data and genome-wide ChIPseq for H3K27Ac in MM cell lines indicate that chromatin surrounding the POU2AF1 locus was highly accessible, concordant with the consistent expression of this TF in MM cell lines and patient-derived cells. CoMMpass data showed that POU2AF1 expression was enhanced in a subset of MM patients at relapse compared to diagnosis. Motif analysis of ChIP-seq data for POU2AF1 identified significant overlap with motifs for TFs relevant to the POU family (e.g. Oct11, Oct2, Oct4); members of the ETS family (e.g. ELF1, Elf4, GABPA); and other TFs with roles in MM including c-myc; IRF4; NF-kappaB, PRDM1, RUNX2 and the POU2AF1 target CREB5. These data suggest a functional interaction between POU2AF1 and other MM-relevant TFs. The transcriptional signature of POU2AF1 activation is enriched for genes downregulated by suppression/inhibition of MM-preferential TFs or epigenetic regulators including IRF4, PRDM1, IKZF1/3 and DOT1L. POU2AF1 binding motifs are also enriched in the promoter regions of MM-preferential dependencies including several MM-preferential TFs. Conclusions: POU2AF1 is essential for MM cells in vitro and in vivo; has a significantly more pronounced and recurrent role as a dependency in MM compared to most other neoplasias; and can further drive MM cell growth, through its ability to interact with several TFs critical for MM, forming multi-protein functional complexes. These results establish POU2AF1 as a central component in the regulatory network of oncogenic TFs in MM and highlight the value of further exploring POU2AF1 as a therapeutic target in MM. Disclosures Downey-Kopyscinski: Rancho BioSciences, LLC: Current Employment. Tsherniak:Cedilla Therapeutics: Consultancy; Tango Therapeutics: Consultancy. Boise:AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genetech: Membership on an entity's Board of Directors or advisory committees. Mitsiades:FIMECS: Consultancy, Honoraria; Ionis Pharmaceuticals, Inc.: Consultancy, Honoraria; Arch Oncology: Research Funding; Janssen/Johnson &amp; Johnson: Research Funding; Karyopharm: Research Funding; TEVA: Research Funding; Takeda: Other: employment of a relative; Fate Therapeutics: Consultancy, Honoraria; Sanofi: Research Funding; Abbvie: Research Funding; EMD Serono: Research Funding.

MM-080: Platelet Hyperactivation in Multiple Myeloma Is Also Evident in Premalignant MGUS Patients and Persists During Treatment Initiation

Context: Thrombotic events are a common complication in multiple myeloma (MM) and smouldering myeloma (SM) patients, and are increasingly being recognized as a feature of MGUS. Platelet hyperactivation has been implicated in this prothrombotic state but remains to be characterized in MGUS patients and MM patients initiating treatment, when thrombotic risk is greatest. Objective: To address this, we evaluated platelet activation status with an extended panel of platelet activation markers (PAC-1, P-selectin, CD63, Annexin V) and platelet responsiveness to agonists (ADP, TRAP-6) in patients and healthy controls (HC). We also assessed additional parameters; reticulated platelet levels, platelet-leucocyte aggregates (PLA), and platelet-associated immunoglobulins (PAIg). Design, Setting & Patients: MGUS (n=19) and SM (n=4) patients who lacked anticoagulant use and underlying bleeding, thrombotic, and renal disorders were recruited from outpatient clinics. Newly diagnosed MM (n=12) patients were recruited using same criteria. A subset of MM patients (n=8) provided samples two months post-initiating treatment. HC (n=16) underwent paraprotein screening to exclude MGUS status. Interventions: No interventions to routine care. MM patients received Bortezomib (Velcade), dexamethasone and Lenalidomide (Revlimid) (VRD) (n=4) or cyclophosphamide (VCD) (n=4), and aspirin thromboprophylaxis (n=5). Main outcome measures: Platelets are hyperactivated and hyporesponsive to agonists in MGUS and SM/MM patients at diagnosis and in patients undergoing treatment. Results: Median platelet activation marker levels were elevated in patients versus HC, significantly for MGUS in terms of PAC-1 (0.72 v 1.7%, p=0.024), CD63 (0.55 v 1.29%, p=0.003), Annexin V (0.47 v 1.61%, p=0.004). In MM undergoing treatment, P-selectin (1.00 v 2.46%, p=0.010), CD63 (0.48 v 1.32%, p=0.020) and Annexin V (0.54 v 1.51%, p=0.038) were significantly elevated versus HC at diagnosis and remained elevated after treatment. MGUS patients had significantly reduced P-selectin levels in response to ADP (34.85 v 10.77%, p=0.005). In MM, hyporesponsiveness was evident versus HC at diagnosis and following treatment in terms of PAC-1 (58.5 v 34.91%, p=0.009) and P-selectin (42.21 v 23.49%, p=0.028). PLA (2.38 v 5.84%, p=0.023) and reticulated platelet levels (1.82 v 24.5%, p=0.031) increased significantly in MM after treatment. PAIg were evident in subsets of MGUS and SM/MM patients. Conclusions: This study reports changes in platelet parameters at various points in MM disease progression. Further investigation of the predictive and prognostic value of platelet dysregulation in such patients is warranted. Funding: Irish Research Council (GOIPG/2017/952) & TRAP Award, College of Medicine & Health, University College Cork.