Multiple Myeloma Tumor Burden Research Articles

Plasma cell–derived mtDAMPs activate the macrophage STING pathway, promoting myeloma progression

AbstractMitochondrial damage–associated molecular patterns (mtDAMPs) include proteins, lipids, metabolites, and DNA and have various context-specific immunoregulatory functions. Cell-free mitochondrial DNA (mtDNA) is recognized via pattern recognition receptors and is a potent activator of the innate immune system. Cell-free mtDNA is elevated in the circulation of trauma patients and patients with cancer; however, the functional consequences of elevated mtDNA are largely undefined. Multiple myeloma (MM) relies upon cellular interactions within the bone marrow (BM) microenvironment for survival and progression. Here, using in vivo models, we describe the role of MM cell–derived mtDAMPs in the protumoral BM microenvironment and the mechanism and functional consequence of mtDAMPs in myeloma disease progression. Initially, we identified elevated levels of mtDNA in the peripheral blood serum of patients with MM compared with those of healthy controls. Using the MM1S cells engrafted into nonobese diabetic severe combined immunodeficient gamma mice, we established that elevated mtDNA was derived from MM cells. We further show that BM macrophages sense and respond to mtDAMPs through the stimulator of interferon genes (STING) pathway, and inhibition of this pathway reduces MM tumor burden in the KaLwRij-5TGM1 mouse model. Moreover, we found that MM-derived mtDAMPs induced upregulation of chemokine signatures in BM macrophages, and inhibition of this signature resulted in egress of MM cells from the BM. Here, we demonstrate that malignant plasma cells release mtDNA, a form of mtDAMPs, into the myeloma BM microenvironment, which in turn activates macrophages via STING signaling. We establish the functional role of these mtDAMP-activated macrophages in promoting disease progression and retaining MM cells in the protumoral BM microenvironment.

Effects of teclistamab and talquetamab on soluble BCMA levels inpatients with relapsed/refractory multiple myeloma.

B-cell maturation antigen (BCMA) is selectively expressed as a surface protein on plasma cells and is a validated target for multiple myeloma (MM). A soluble form of BCMA (sBCMA) is elevated in the sera of patients with MM. The bispecific antibodies teclistamab (BCMA×CD3) and talquetamab (G protein-coupled receptor family C group 5 member D [GPRC5D]×CD3) are in clinical development as therapies for MM. Using data from patients with relapsed/refractory MM (N=300) who participated in phase 1 studies of teclistamab (NCT03145181) or talquetamab (NCT03399799), baseline serum sBCMA levels were quantitatively analyzed relative to response to treatment, percentage of bone marrow plasma cells (BMPCs), and cytogenetic risk. By cycle 3 day 1 of treatment, sBCMA levels declined from baseline in 50 (88%) of 57 responders and 49 (98%) of 50 responders to teclistamab and talquetamab, respectively. Depth of response correlated with the magnitude of sBCMA reduction. In contrast, sBCMA increased in 33 (80%) of 41 patients and 24 (49%) of 49 patients who did not respond to teclistamab or talquetamab, respectively. Baseline sBCMA levels correlated with the percentage of BMPC; patients with extramedullary plasmacytomas who had low levels of BMPC (≤10%) tended to have high baseline sBCMA levels (≥400 ng/mL), based on limited data. Baseline sBCMA levels and changes in sBCMA levels at cycle 3 day 1 were similar in patients with high- and standard-risk cytogenetics treated with teclistamab or talquetamab. These results support the use of sBCMA as a potential surrogate marker of tumor burden and treatment response in MM.

The deubiquitinase OTUD1 regulates immunoglobulin production and proteasome inhibitor sensitivity in multiple myeloma

Serum monoclonal immunoglobulin (Ig) is the main diagnostic factor for patients with multiple myeloma (MM), however its prognostic potential remains unclear. On a large MM patient cohort (n = 4146), we observe no correlation between serum Ig levels and patient survival, while amount of intracellular Ig has a strong predictive effect. Focused CRISPR screen, transcriptional and proteomic analysis identify deubiquitinase OTUD1 as a critical mediator of Ig synthesis, proteasome inhibitor sensitivity and tumor burden in MM. Mechanistically, OTUD1 deubiquitinates peroxiredoxin 4 (PRDX4), protecting it from endoplasmic reticulum (ER)-associated degradation. In turn, PRDX4 facilitates Ig production which coincides with the accumulation of unfolded proteins and higher ER stress. The elevated load on proteasome ultimately potentiates myeloma response to proteasome inhibitors providing a window for a rational therapy. Collectively, our findings support the significance of the Ig production machinery as a biomarker and target in the combinatory treatment of MM patients.

Monocyte or white blood cell counts and β2 microglobulin predict the durable efficacy of daratumumab with lenalidomide.

Daratumumab is one of the most widely used treatments for relapsed/refractory multiple myeloma (MM) patients. However, not all patients achieve a lasting therapeutic response with daratumumab. We hypothesized that a durable response to daratumumab could be predicted by the balance between the MM tumor burden and host immune status. We conducted a retrospective study using the real-world data in the Kansai Myeloma Forum (KMF) database. We retrospectively analyzed 324 relapsed/refractory MM patients who were treated with daratumumab in the KMF database. In this study, 196 patients were treated with daratumumab, lenalidomide, and dexamethasone (DLd) regimen and 128 patients were treated with daratumumab, bortezomib, and dexamethasone (DBd) regimen. The median age at treatment, number of prior treatment regimens and time-to-next-treatment (TTNT) were 68, 4 and 8.02 months, respectively. A multivariate analysis showed that the TTNT under the DLd regimen was longer with either higher monocyte counts (analysis 1), higher white blood cell (WBC) counts (analysis 2), lower β2 microglobulin (B2MG < 5.5 mg/L) or fewer prior regimens (<4). No parameters were correlated with TTNT under the DBd regimen. We propose a simple scoring model to predict a durable effect of the DLd regimen by classifying patients into three categories based on either monocyte counts (0 points for ⩾200/μl; 1 point for <200/μl) or WBC counts (0 points for ⩾3500/μl; 1 point for <3500/μl) plus B2MG (0 points for <5.5 mg/L; 1 point for ⩾5.5 mg/L). Patients with a score of 0 showed significantly longer TTNT and significantly better survival compared to those with a score of 1 or 2 (both p < 0.001). To confirm this concept, our results will need to be validated in other cohorts.

Aberrant ENPP2 expression promotes tumor progression in multiple myeloma

Ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2) has been recently linked to tumor development. However, its role in modulating multiple myeloma (MM) disease progression remains unclear. Here, we demonstrated that CD138+ cells isolated from MM patients presented with higher expression of ENPP2 compared with CD138- cells. Treatment of MM cells with IL-6 resulted in ENPP2 upregulation. ENPP2 overexpression promoted proliferation, inhibited apoptosis, increased lysophosphatidic acid (LPA) generation, and upregulated osteoclastogenesis mediator expression in MM cells. In contrast, ENPP2 inhibition induced apoptosis, suppressed proliferation and survival, decreased LPA generation and downregulated osteoclastogenesis mediator expression. In an MM xenograft mouse model, ENPP2 knockdown significantly reduced MM tumor burden by inhibiting cell proliferation and inducing apoptosis. Furthermore, ENPP2 knockdown decreased the levels of LPA, osteoclastogenesis mediators in sera of mice with MM. Our findings revealed the tumor-promoting role of ENPP2 in MM, thus providing new molecular evidence for targeting the ENPP2-LPA axis in MM therapy.

Circulating Plasma Cells Are the Most Powerful Prognostic Marker in Transplant Ineligible Multiple Myeloma with 2% As a New Cut-Off for Primary Plasma Cell Leukemia

Background:Tumor burden in multiple myeloma (MM) is routinely evaluated in the bone marrow, though its prognostic value is not proven. There is an increasing interest in liquid biopsies due to its minimally invasive nature and more comprehensive evaluation of tumor burden. Growing evidence supports the quantification of circulating plasma cells (cPCs) measured by multiparameter flow cytometry (MFC) as a powerful diagnostic biomarker suitable for risk stratification of newly diagnosed transplant eligible MM (Garces et al, EHA 2021). Nevertheless, there are virtually no data regarding prognostic impact of cPCs in MM patients ineligible for transplantation. Primary plasma cell leukemia (pPCL) is a rare and most aggressive monoclonal gammopathy with dismal outcomes defined by more than 20% of cPCs and/or absolute cPCs count of ≥2 x10 9/L. Recently, there have been many efforts to redefine these criteria as lower number of cPCs probably portends equally poor prognosis.Aims:To evaluate prognostic significance of cPCs in a large cohort of transplant ineligible (Tx-ineligible) newly diagnosed MM patients and to define cut-offs for risk stratification. Moreover, to establish cut-off identifying ultra high risk MM patients mimicking the prognosis of pPCL and propose new definition of pPCL.Methods:Circulating PCs were measured by 8 color flow cytometry in 402 Tx-ineligible MM patients (including n=7 pPCL) diagnosed between 2012 and 2019 at University Hospitals Brno and Ostrava, Czech Republic. Patients were treated in real-world setting and the clinical analysis was performed retrospectively based on data from the Czech Registry of Monoclonal Gammopathies. Median follow-up was 20.5 months. The intermediate cutoff was identified using ROC analysis considering overall survival (OS). Moreover, data from the large published cohort of pPCL patients treated in real-world setting were used to find a new cut-off identifying these ultra-high-risk pPCL-like patients (Jurczyszyn et al. BJH,2018).Results:Circulating PCs were detected in peripheral blood (PB) of 303/402 (75%) patients. In 303 patients with detectable cPCs the median percentage was 0.06% with range 0.0008% - 79%. The median limit of detection of MFC technique was 0.006 (sensitivity 10e-5). Patients stratification into 3 subgroups according to quantification of cPCs (low: ≤ 0.2%; intermediate: 0.2 - 2% and high: >2.0%) resulted in significantly different OS (36.5 vs. 28.1 and 13.6 months; p < 0.0001) and progression free survival (PFS) (17.9 vs. 14.7 and 3.4 months; p < 0.0001). Patients with no detected cPCs (0%) did not separate from subgroup >0% to <0.2% (p >0.05) suggesting that next-generation flow cytometry (NGF) with sensitivity 10e-6 is needed for the identification of this favorable prognostic group. In order to demonstrate that patients with more than 2% of cPCs have similarly poor outcome as pPCL patients, we compared those with 2% - 20% (n=15) vs. those with >20% (n=7) of cPCs. The outcomes were practically identical with median PFS of 3.1 vs. 4.2 months (p = 0.23) and median OS of 13.6 vs. 14.6 months (p=0.23).Next, we analyzed patient´s characteristics in association with the level of cPCs (low, intermediate and high) and we demonstrated that there was significantly higher proportion of patients with ISS III stage (38%, 52% and 82%), elevated LDH level (8%, 19% and 55%) and high risk cytogenetics (12%, 21% and 36%) hand in hand with increasing number of cPCs. CPCs were identified as the most powerful prognostic marker in univariate (HR=2.7 for OS and HR=6.3 for PFS; p=0.001) and multivariate analysis (HR=4.2 for OS and HR=5.8 for PFS; p<0.001), including ISS, LDH and FISH cytogenetics.Conclusion:The quantification of cPCs in PB of newly diagnosed MM is the most powerful prognostic factor as we demonstrated on a large cohort of transplant ineligible patients. We defined 2% of cPCs as a new cut-off for ultra high risk myeloma resembling behavior of primary PCL. We propose this 2% cut-off for redefinition of pPCL criteria that warrants further investigation in prospective setting. To identify subgroup with especially favorable outcome with no detectable cPCs, NGF with high sensitivity of 10e-6 is needed. Quantification of cPCs by MFC is easy, fast, affordable and worldwide available procedure providing highly relevant prognostic information that might be implemented into routine clinical practice. [Display omitted] DisclosuresJurczyszyn: Janssen-Cilag, Amgen: Honoraria, Speakers Bureau. Castillo: Abbvie: Consultancy, Research Funding; BeiGene: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Janssen: Consultancy; Roche: Consultancy; TG Therapeutics: Research Funding. Hajek: Janssen: Consultancy, Honoraria, Research Funding; Pharma MAR: Consultancy, Honoraria; BMS: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.

Genetic Inhibition of PD-L1 on Myeloma Cells Improves Survival in Contrast to the Treatment with Checkpoint Inihibitors

Checkpoint inhibition (CPI) has shown dramatic improvements in overall survival in many malignant diseases. However, in multiple myeloma (MM) the results were disappointing resulting in an early termination of clinical trials. Despite the advantages in therapy the disease remains incurable.Methods: We analyzed the efficiency and immunological mechanisms of PD-1/PD-L1 blockade using KaLwRij mice that develop MM upon injection of 5T33 myeloma cells. Treatment of mice started d19 post inoculation. Bone marrow (BM) and spleen cells were analyzed by flow cytometry for the phenotype of immune cells.Results: Comprehensive immunophenotyping including the analysis of T and NK cell subpopulations revealed no differences of early MM disease stage compared to healthy control groups. Treatment of mice with mAbs blocking PD-1 or PD-L1 had no effect on tumor growth and survival. It was demonstrated that HDAC inhibitors beside their direct effect on malignant cells may increase the immunogenicity of malignant cells by improving the presentation of tumor antigens and modulate the immunological composition of the tumor microenvironment (TME). The pan-HDAC inhibitor panobinostat that is approved for the treatment of myeloma patients inhibited the development of myeloma in treated mice. Surprisingly, the combined application of the anti-PD-1 blocking antibody with panobinostat reduced the anti-myeloma effect of the compound and resulted in decreased survival.By analyzing the phenotype of immune cells in the different populations, we found in the panobinostat treated animal group an increase in the CXCR4 expressing CD4+ NKT cells. Additionally, the CD8+ T cells expressing CD1d and CXCR4 decreased compared to the other groups in the spleen. CD1d is a MHC like receptor for glycolipids activating NKT cells, whereas CXCR4 is a BM homing receptor and linked to metastasis and tumor aggressiveness. We found a gradually increase of CXCR4+ NKT cells in the BM corresponding to MM disease progression. Interestingly, we observed a shift in the CD4+/CD4- NKT cell ratio during disease progression, whereby the CXCR4+ CD4- NKT cells seem to be associated with advanced tumor growth, while the increase of CXCR4+ CD4+ NKT are associated with prolonged survival as observed in in the panobinostat treated group.To further analyze the role of PD-L1 expression on myeloma cells we generated a PD-L1 KO of the 5T33 cell line using the CRISPR/Cas9 technology. We found no differences in the expression of surface molecules such as MHC class I and II, co-stimulatory or adhesion molecules, proliferation and migration of the genetically engineered cells in comparison to the mock control. Interestingly, mice inoculated with the 5T33 PD-L1 KO cells showed a significant longer survival compared with the 5T33 mock injected, indicating that blocking of the PD-L1 molecule on myeloma cells plays an important role in the pathogenesis of MM and its direct blocking on malignant cells rather than in the TME might have an impact on the clinical efficiency. When analyzing the spleen of the mock vs PD-L1 KO myeloma inoculated mice, we found the same downregulation of CXCR4 and CD1d on CD8+ T cells in the PD-L1 KO myeloma group as observed in the panobinostat treated group with extended survival.In addition, we used NOD. scid. Il2Rγc null (NSG) mice to proof that the survival prolongation is a result of the immunological response to PD-L1 and that the myeloma cells are not otherwise effected in their tumor cell properties in vivo. NSG mice experience the same tumor burden post 5T33 mock and PD-L1 KO challenge, assuming that the previous observed survival prolongation is exclusively dependent on the PD-L1 tumor- immune cell interaction.Conclusion: We found that PD-1 blockade might negatively affect and inhibit the therapeutic efficacy of HDAC inhibitors such as panobinostat. Genetic down regulation of PD-L1 on the myeloma cells enables a significant improvement and longer survival. These results give new insights into the complexity of the action of CPI in the treatment of malignant diseases which might help to develop combinatorial approaches of checkpoint inhibitors in clinical trials. Furthermore, the increase of CD4- CXCR4 expressing NKT cells in the BM might be used as biomarker to monitor MM disease progression, whereas the increase of the CD4+/CD4- NKT cell ratio in the BM might be associated with the shrinkage of MM tumor burden. DisclosuresBrossart: BMS: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; MSD: Honoraria.

Quantitative whole-body MR imaging for assessment of tumor burden in patients with multiple myeloma: correlation with prognostic biomarkers.

To assess the quantification of tumor burden in multiple myeloma (MM) patients using whole-body magnetic resonance imaging (MRI) and to identify the correlation between MRI parameters and prognostic biomarkers. We retrospectively analyzed 95 newly diagnosed MM patients treated at our hospital from June 2018 to March 2020. All patients underwent whole-body MRI examination, including diffusion-weighted whole-body imaging with background body signal suppression (DWIBS), modified Dixon chemical-shift imaging (mDIXON), and short TI inversion recovery (STIR) sequences. The MRI presentation was used to determine MM infiltration patterns and calculate apparent diffusion coefficient (ADC) and a fat fraction (FF). The one-way ANOVA and Kruskal-Wallis test were used to compare the differences of these values between DS, ISS, and R-ISS stages in different MM infiltration patterns. Spearman correlation test was used for correlation analysis of ADC and FF against prognostic biomarkers, and two independent sample t-test was used to evaluate the differences of ADC and FF in different free light-chain ratio groups. The MRI presentation was classified into normal pattern (36 patients; 37.9%), diffuse (27 patients; 28.4%), and focal (32 patients; 33.7%) infiltration patterns. Statistically significant ADC and FF differences between different DS, ISS, and R-ISS stages were observed in normal/diffuse infiltration patterns but not in focal infiltration patterns. The ADC and FF of the normal/diffuse infiltration pattern showed correlations with hemoglobin, β2-microglobulin, bone marrow plasma cells, flow cytometry of bone marrow cells, and serum monoclonal protein. In contrast, ADC in focal infiltration patterns was negatively correlated with β2-microglobulin and C-reactive protein. The FF of patients with a normal/diffuse infiltration pattern was higher in the low free light-chain ratio group than that in the high free light-chain ratio group (P=0.023). Our observations indicate that quantitative whole-body functional MRI examination may serve as an effective complement to imaging diagnosis based on morphology and provide further information on the tumor burden of patients with MM.

PK/PD modeling analysis for dosing regimen selection of isatuximab as single agent and in combination therapy in patients with multiple myeloma.

This analysis describes the pharmacokinetic/pharmacodynamic (PK/PD) modeling framework that supported selection of the isatuximab (anti‐CD38 monoclonal antibody) dosing regimen alongside its early clinical development in patients with relapsed/refractory multiple myeloma (RRMM). The PK/PD mathematical model characterized the variations of patient serum M‐protein concentrations, the primary marker of tumor burden in multiple myeloma (MM). Three separate PK/PD models were built sequentially as data became available from phase I clinical trials. The primary PK/PD analysis was initiated using monotherapy phase I study data (n = 122), followed by analysis of data collected from phase Ib combination studies with lenalidomide and dexamethasone (Rd, n = 40) and then with pomalidomide and dexamethasone (Pd, n = 31). Using the PK/PD model, abnormal “myeloma” protein (M‐protein) profiles under different isatuximab dosing regimens were simulated. Overall, simulations revealed that regimens which included a loading period of four weekly administrations followed by administration every 2 weeks thereafter (QW4‐Q2W), reduced M‐protein levels more than a Q2W regimen without a loading period. For isatuximab monotherapy, a 20 mg/kg dose induced greater reduction in serum M‐protein levels compared with doses equal or lower than 10 mg/kg. For isatuximab in combination with either Rd or Pd, simulations yielded no substantial benefit in terms of M‐protein reduction between isatuximab 10 mg/kg and 20 mg/kg. These PK/PD analyses supported the use of isatuximab 10 mg/kg QW4‐Q2W in combination with Pd in the phase III trial.

Phase 2 Study of Pembrolizumab during Lymphodepleted State after Autologous Hematopoietic Cell Transplantation in Multiple Myeloma Patients

Background: Patients (pts) with multiple myeloma (MM) who do not achieve complete response (CR) with primary therapy, including autologous hematopoietic cell transplantation (AHCT), have worse long term outcomes. Preclinical and clinical evidence suggests that the immune checkpoint programmed death-1 (PD-1) receptor/PD-1 ligand axis plays an important role in suppressing immune surveillance against MM, especially after AHCT. Thus, we hypothesized that the administration of pembrolizumab, an anti-PD-1 antibody, to disrupt the PD-1/PD-L1 axis to augment T and NK cell function during the lymphodepleted state post-AHCT (lack of competitors: cytokine sink) when PD-1 expression on effector cells peaks and MM tumor burden nadirs, may result in therapeutic efficacy in MM. Patients and Methods: We conducted a phase 2, two-site, single-arm study of post-AHCT pembrolizumab in MM pts who had not achieved pre-AHCT CR to induction therapy. Pembrolizumab was administered 200 mg IV fixed dose every 3 weeks, starting day+14 after AHCT (upon engraftment) for 9 doses (day+180). The primary endpoint was CR rate after last treatment in pts receiving ≥2 doses (evaluable pts). Bone marrow (BM) minimal residual disease (MRD) testing was checked using flow cytometry in each institution (sensitivity 1 in 30,000 - 1 in 200,000). Pts aged 18-70 years with MM within 12 months after diagnosis, of any molecular risk group and with Results: Thirty-two pts were enrolled but 3 withdrew consent prior to receiving first dose. For the 29 pts who received pembrolizumab, there were 19 males and 10 females and the median age was 59 years (range, 49-70). Four pts (14%) had light chain MM. ISS stages were stage I and II in 10 pts each (34%) and stage III in 9 pts (33%). All pts except 1 received a triple-regimen primary therapy, with a median of 4 cycles (range, 2-7). Twenty pts (69%) achieved partial response (PR) and 9 (31%) very good PR pre-AHCT. The median post-transplant day of initiation of pembrolizumab was 14 days (range, 13-19). The median number of doses received was 9 (range, 1-9). Among these 29 pts, grade 3 toxicities leading to discontinuation included infusion reaction and colitis in 1 pt each and grade 2 radiculopathy in another pt. Overall, another 12 immune-mediated adverse reactions that did not lead to discontinuation were grade 1-2 events including 4 infusion reactions, 2 colitis, 2 skin rashes, 2 hypothyroidism and grade 3 events of acute kidney injury and hepatitis (1 pt each). There were no primary or secondary graft failures, and the median time to neutrophil and platelet engraftment days were 11 days (range, 6-13) and 12 days (range, 8-39), respectively. Twenty-six pts were considered evaluable for response assessment. Twenty-four pts started lenalidomide maintenance at the median of 103 (range, 60-154) days post-AHCT. For the other 2 pts, one had progressive disease at day 100 and the other withdrew consent. Of the 23 pts who had a complete end-of-treatment (EOT) response evaluation by serology alone (non-CR) or both serology and bone marrow (BM) study (morphology and flow cytometry) (CR); the CR rate was 31% (7/23). In the remaining 3 pts, 2 pts with serologic CR did not have a confirmed BM at EOT study while 1 pt had no EOT evaluation. The MRD-negative rates by BM flow cytometry were 44% (11/25) at day100 and 67% (12/18) at EOT. With the median follow-up of 11.3 months (range, 3-24), none of 7 pts achieving CRs has relapsed; 3 progressed by EOT and one did by 8 months after initial EOT response. The estimated 1-year overall survival and progression-free survival are 100% and 87% (95% CI; 74%-100%), respectively. There was no correlation between the numbers of T cells, NK cells or B cells in the grafts infused back and clinical response (CR and BM MRD-negative disease) (Figure). Conclusions: The addition of pembrolizumab in the early post-AHCT period was safe, as was the combined administration with standard-of-care low-dose lenalidomide maintenance. Pembrolizumab resulted in CR rate of 31% at 6 months, including a very encouraging 67% rate of BM MRD-negative state, in these patients not in CR pre-AHCT. Concurrent administration of pembrolizumab and low-dose lenalidomide maintenance is able to deepen the responses observed after AHCT. Disclosures Pawarode: AMGEN: Research Funding; MERCK: Research Funding. D9Souza: Merck: Research Funding; Prothena: Research Funding; Celgene: Consultancy. Dhakal: Celgene: Honoraria. Hamadani: Sanofi Genzyme: Research Funding, Speakers Bureau; Takeda, Otsuka, MedImmune, Merck, ADC Therapeutics: Research Funding; Celgene, Cellerant, Jansen, MedImmune: Consultancy. Shah: Oncosec: Equity Ownership; Exelixis: Equity Ownership; Geron: Equity Ownership; Jazz Pharmaceuticals: Consultancy. Hari: Janssen: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Sanofi: Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Honoraria, Research Funding; Novartis: Consultancy; Spectrum: Consultancy, Research Funding; Takeda Pharmaceuticals: Consultancy, Honoraria, Research Funding.

Total diffusion volume in MRI vs. total lesion glycolysis in PET/CT for tumor volume evaluation of multiple myeloma.

This study compared the tumor burden and prognostic impact of total diffusion volume (tDV) and total lesion glycolysis (TLG) in the same patients with newly diagnosed multiple myeloma (NDMM) simultaneously. We also examined the relationship between these imaging tumor volumes (TVs) and plasma cell (PC) TV in bone marrow (BM) specimens. We retrospectively reviewed the data of 63 patients with newly diagnosed multiple myeloma (NDMM) from April 2016 to March 2018. tDV was calculated from whole-body diffusion-weighted imaging and TLG was calculated from the average standard uptake value and the metabolic tumor volume, respectively. Cellularity of BM hematopoietic tissue and the percentage of BM PCs were used as a reference of PC volume in the BM. The Spearman correlation coefficient between tDV and TLG was moderate (ɤs = 0.588, p < 0.001) when PET false-negative patients were excluded. There were positive correlations between the BM plasma cell volume (BMPCV) and the imaging TVs (ɤs = 0.505, vs. tDV; and 0.464, vs. TLG). Patients with high tDV and high TLG, as determined by the receiver operating characteristic curve, had worse survival; moreover, patients with both high tDV and high TLG showed the worst prognosis (median progression-free and overall survival: 13.2 and 28.9 months, respectively). Although tDV and TLG each reflected the total TV, in several cases, tDV and TLG were discrepant due to the biological features of each MM. It is important to use both modalities for complementary assessment of total tumor burden and biological characteristics in MM. • Total diffusion volume (tDV) and total lesion glycolysis (TLG) reflect the total tumor volume and have prognostic value in patients with multiple myeloma (MM). • tDV and TLG could assess MM from different biological perspectives and should be considered for each patient individually.

Total Diffusion Volume Estimated By Whole-Body Diffusion Magnetic Resonance Imaging for the Assessment of Tumor Burden in Multiple Myeloma

Introduction: Whole-body diffusion-weighted magnetic resonance imaging (wbDWI) is an emerging imaging technique for assessing the tumor burden of patients with multiple myeloma (MM). Total diffusion volume (tDV) is a potential parameter to quantify the tumor volume of bone marrow (BM) images obtained by wbDWI. The apparent diffusion coefficient (ADC) value, also calculated with wbDWI, has the potential to accurately evaluate the treatment response of MM; however, correlation between tDV and the treatment response of MM remains poorly studied. Methods: We retrospectively selected patients with newly-diagnosed, symptomatic MM who were treated at Kameda Medical Center, Japan, from January 2016 to November 2019; we analyzed the tDV obtained by wbDWI before and after treatment. We used the BD score software (PixSpace, Japan) to analyze tDV. Briefly, the regions of interest, including areas with abnormally high signal intensity of diffusion weighted imaging considered to contain myeloma lesions in the BM or extramedullary tissue, were automatically obtained with a b-value of 900 s/mm2. Physiological areas with high signal intensity were removed manually. Threshold values for distinguishing diseased areas from the background were manually adjusted to cover all bone lesions. Diagnosis and response were assessed using the International Myeloma Working Group criteria. We compared tDV before and after treatment, and in each response category. Comparison of continuous variables between the two categories and between three or more sets were analyzed using Wilcoxon test and Kruskal-Wallis test, respectively. All statistical analyses were performed using R (version 4.0.2; R Foundation, Vienna, Austria). Results: We analyzed 42 patients (18 men; median age, 71.5 years; interquartile range [IQR]: 65.0-77.0) whose pre- and post-treatment tDV values were available. The type of myeloma at diagnosis included immunoglobulin G (IgG) (n=21, 50%), immunoglobulin A (IgA) (n=12, 28%), and light chain alone (n=9, 21.4%). Three patients (7.1%) underwent wbDWI when they achieved partial response (PR); 6 patients (14.2%) achieved very good partial response (VGPR), 3 patients (7.1%) achieved complete response (CR), and 30 patients (71.4%) achieved stringent complete response (sCR). In all patients, median tDV decreased significantly before and after the treatment (median, 464.24 mL [IQR: 152.99-929.23] and 108.2 mL [IQR: 27.76-368.85], respectively; p&amp;lt;0.001) (Figure 1). Median tDV after treatment trended to be progressively low for patients who achieved PR, VGPR, CR, and sCR (tDV: 381.33 mL, 218.85 mL, 19.26 mL, and 63.97 mL, respectively; p=0.12). However, these values were not statistically significant. We also compared the tDV before and after treatment for patients who achieved less than CR (PR and VGPR) and those who achieved CR or better (≥CR). The median tDV of the patients who achieved less than CR was 298.24 mL [IQR: 147.88-381.33] and for those who achieved ≥CR was 62.65 mL [IQR: 19.26-279.79] (p=0.02) (Figure 2). We also compared the decreasing rate of tDV before and after treatment between patients who achieved less than CR and those who achieved ≥CR. The median decreasing rate of tDV was 61.0% 72.0% for patients who achieved less than CR and ≥CR, respectively. Although this result was not statistically significant, the data suggests that patients who achieved a deeper response tended to have a larger decreasing rate of tDV. Conclusions: Our data shows that tDV correlates with the depth of treatment response, and thus, is a potentially useful marker for evaluating the treatment response in patients with MM. Disclosures No relevant conflicts of interest to declare.

Monocytosis at Diagnosis Is Associated with Decreased Overall Survival in Patients with Newly Diagnosed Multiple Myeloma

Introduction : Multiple myeloma (MM) is an incurable hematologic malignancy caused by the neoplastic proliferation of plasma cells within the bone marrow. The tumor microenvironment within the bone marrow consists of a diverse array of immune cells which regulate MM cell proliferation and survival. Specifically, tumor-associated macrophages (TAMs) derived from peripheral blood monocytes migrate to the bone marrow where they support MM cell growth and promote resistance to apoptosis. Several studies suggest that the amount of bone marrow TAMs directly correlates with disease activity and worse clinical outcomes in MM. Considering that peripheral blood absolute monocyte count (AMC) could reflect the burden of bone marrow TAMs, we sought to determine the prognostic significance of AMCs at diagnosis of MM. Methods: Using the integrated nationwide VA electronic health records and VA Corporate Data Warehouse, patients were identified by International Classification of Diseases (ICD) codes for MM. Patients were required to have at least 3 visits with an MM ICD code on separate days, have received at least one MM drug with the exception of corticosteroids after the date of diagnosis and have 2 or more absolute monocyte counts measured by automated or manual differential at separate visits within 18 months before and 7 days after MM diagnosis. We further confirmed the date of diagnosis using the VA Cancer Registry. We excluded patients with aplastic anemia, myelodysplastic syndrome, chronic myelomonocytic leukemia and other myeloproliferative neoplasms. In this cohort, monocytosis was defined based on the institution's cut-off as a sustained absolute increase in monocyte count of 0.8 (800/mm3) or greater with extreme monocytosis being greater than 1.25 (1250/mm3). Patients were stratified into three groups according to AMC at diagnosis, namely AMC &amp;lt;0.8, AMC 0.8 - 1.25 and AMC &amp;gt;1.25. To avoid extreme values significantly affecting the analysis, the Kruskal-Wallis test was used to compare continuous variables between the groups. Chi-squared tests were used to compare categorical variables. Hazard estimates for the prognostic analysis were obtained via adjusted Cox proportional hazard models. Overall survival was estimated using the Kaplan-Meier method and log-rank tests were used to compare the survival of each group. Results: A total of 5,265 patients with MM were included in the final analysis with a median follow up of 2.9 years (range 1.3 - 5.2). At diagnosis of MM, 39.3% of patients presented with monocytosis. Patients with monocytosis were younger and more likely to have abnormal levels of prognostic markers known to be associated with increased tumor burden and worse outcomes in MM. Notably, patients with monocytosis had higher median levels of β2-microglobulin and lactate dehydrogenase (p &amp;lt;0.001) and lower median albumin levels (p &amp;lt;0.001) than those without monocytosis. Patients with extreme monocytosis had the highest median β2-microglobulin (5.01, range 3.30 - 6.98, p &amp;lt;0.001) and the lowest median albumin levels (2.9, range 2.4 - 3.4, p &amp;lt;0.001) in the entire cohort. Patient characteristics according to AMC at diagnosis are shown in Table 1. On univariate analysis, the AMC predicted overall survival when analyzed as a trichotomized variable. The median overall survival was 4.6, 3.5 and 3.1 years in the AMC &amp;lt;0.8, AMC 0.8-1.25 and AMC &amp;gt;1.25 groups respectively (p &amp;lt;0.0001) [Figure 1]. When adjusting for known prognostic factors associated with outcomes in MM, the AMC was significantly associated with inferior overall survival (AMC 0.8 - 1.25 HR 1.230, p &amp;lt;0.008; AMC &amp;gt;1.25 HR of 1.312, p &amp;lt;0.003). Conclusion : The results of our study suggest that an elevated absolute monocyte count at MM diagnosis is associated with worse overall survival independent of known poor prognostic factors. The role of peripheral blood monocyte count as a surrogate biomarker for the state of the bone marrow microenvironment in MM warrants further in vitro, prospective studies. Disclosures Munshi: AbbVie: Consultancy; Amgen: Consultancy; Legend: Consultancy; Adaptive: Consultancy; Janssen: Consultancy; C4: Current equity holder in private company; OncoPep: Consultancy, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees, Patents &amp; Royalties; Karyopharm: Consultancy; Takeda: Consultancy; BMS: Consultancy.

Prospective Comparison of 18F-Choline Positron Emission Tomography/Computed Tomography (PET/CT) and 18F-Fluorodeoxyglucose (FDG) PET/CT in the Initial Workup of Multiple Myeloma: Study Protocol of a Prospective Imaging Trial.

BackgroundThe International Myeloma Working Group recommends the use of 18-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) for treatment response evaluation, as it is superior to magnetic resonance imaging (MRI). However, at initial staging, the sensitivity of FDG-PET remains inferior to that of MRI. Therefore, there is a need for an imaging technique that could have a sensitivity equal to that of MRI at diagnosis and could serve to evaluate therapy. 18F-choline has shown increased sensitivity when compared with 18-FDG, with about 75% more lesions detected in patients with relapsed or progressive multiple myeloma (MM).ObjectiveOur primary objective is to prospectively compare the detection rate of bone lesions by 18F-choline PET/CT (FCH-PET) and FDG-PET in newly diagnosed MM. Our secondary objectives are to assess the accuracy of both PET modalities for the detection of bone lesions and the diagnosis of diffuse disease, to assess the detection rate of extramedullary lesions.MethodsWe will prospectively include 30 patients in a paired comparative accuracy study. Patients with de novo MM will undergo FCH-PET, FDG-PET, and whole-body MRI (WB-MRI) within a 3-week period. WB-MRI will be composed of conventional sequences on the spine and pelvis and of whole-body diffusion axial sequences. The following 6 skeletal areas will be defined: skull, sternum/costal grid, spine, pelvis, superior limbs, and inferior limbs. The number of focal lesions, their respective localization, and intensity of uptake will be retrieved for each skeletal area. Readings will be performed blinded from other imaging techniques. The reference standard will be WB-MRI. Focal lesions present on PET/CT but not on WB-MRI will require a decision made with a consensus of experts based on clinical and imaging data. The number of bone lesions and number of extramedullary lesions will be compared using the Wilcoxon test. The accuracy of FCH-PET and FDG-PET will be compared using the McNemar test.ResultsThe study started in September 2019, and enrollment is ongoing. As of June 2020, 8 participants have been included. Data collection is expected to be completed in June 2021, and the results are expected to be available in December 2021.ConclusionsThis study will assess if FCH-PET is superior to FDG-PET for the evaluation of MM tumor burden. This will pave the way for future prospective evaluations of the prognostic value of 18-FCH for treatment response evaluation in MM patients. Additionally, this work may provide new perspectives for better assessment of the risk of smoldering MM progressing to MM.Trial RegistrationClinicalTrials.gov NCT03891914; https://clinicaltrials.gov/ct2/show/NCT03891914International Registered Report Identifier (IRRID)DERR1-10.2196/17850

Targeted Disruption of Bone Marrow Stromal Cell-Derived Gremlin1 Limits Multiple Myeloma Disease Progression In Vivo.

In most instances, multiple myeloma (MM) plasma cells (PCs) are reliant on factors made by cells of the bone marrow (BM) stroma for their survival and growth. To date, the nature and cellular composition of the BM tumor microenvironment and the critical factors which drive tumor progression remain imprecisely defined. Our studies show that Gremlin1 (Grem1), a highly conserved protein, which is abundantly secreted by a subset of BM mesenchymal stromal cells, plays a critical role in MM disease development. Analysis of human and mouse BM stromal samples by quantitative PCR showed that GREM1/Grem1 expression was significantly higher in the MM tumor-bearing cohorts compared to healthy controls (p < 0.05, Mann–Whitney test). Additionally, BM-stromal cells cultured with 5TGM1 MM PC line expressed significantly higher levels of Grem1, compared to stromal cells alone (p < 0.01, t-test), suggesting that MM PCs promote increased Grem1 expression in stromal cells. Furthermore, the proliferation of 5TGM1 MM PCs was found to be significantly increased when co-cultured with Grem1-overexpressing stromal cells (p < 0.01, t-test). To examine the role of Grem1 in MM disease in vivo, we utilized the 5TGM1/KaLwRij mouse model of MM. Our studies showed that, compared to immunoglobulin G (IgG) control antibody-treated mice, mice treated with an anti-Grem1 neutralizing antibody had a decrease in MM tumor burden of up to 81.2% (p < 0.05, two-way ANOVA). The studies presented here demonstrate, for the first time, a novel positive feedback loop between MM PCs and BM stroma, and that inhibiting this vicious cycle with a neutralizing antibody can dramatically reduce tumor burden in a preclinical mouse model of MM.

Tumor load in patients with multiple myeloma: β₂-microglobulin levels versus low-dose whole-body CT.

Beta-2-microglobulin is a serum marker of tumor burden in multiple myeloma (MM). Our aim was to correlate serum β₂-microglobulin levels in patients with MM to tumor burden determined by low-dose whole-body CT (LDWBCT). A total of 52 patients with newly diagnosed, untreated MM who underwent LDWBCT were included. LDWBCT scans were assessed by two musculoskeletal radiologists in consensus for focal lesions. The Durie and Salmon PLUS staging system was used for staging patients in stages I-III. β₂-microglobulin was also subdivided into stages I-III on the basis of the multiple myeloma International Staging System (ISS). Using the Durie and Salmon PLUS staging system criteria for image evaluation, we were able to identify stage I MM in 17 patients, stage II MM in nine patients, and stage III MM in 26 patients. Eight of nine patients with stage II MM and 16 of 26 patients with stage III MM had normal β₂-microglobulin levels. Thus, 24 of 35 patients (68.6%) had 5 or more focal lesions and false-negative β₂-microglobulin levels. Serum β₂-microglobulin levels alone may not indicate the full extent of tumor burden in a significant subset of myeloma patients.

Quantification of tumor burden in multiple myeloma by atlas-based semi-automatic segmentation of WB-DWI

BackgroundWhole-body diffusion weighted imaging (WB-DWI) has proven value to detect multiple myeloma (MM) lesions. However, the large volume of imaging data and the presence of numerous lesions makes the reading process challenging. The aim of the current study was to develop a semi-automatic lesion segmentation algorithm for WB-DWI images in MM patients and to evaluate this smart-algorithm (SA) performance by comparing it to the manual segmentations performed by radiologists.MethodsAn atlas-based segmentation was developed to remove the high-signal intensity normal tissues on WB-DWI and to restrict the lesion area to the skeleton. Then, an outlier threshold-based segmentation was applied to WB-DWI images, and the segmented area’s signal intensity was compared to the average signal intensity of a low-fat muscle on T1-weighted images. This method was validated in 22 whole-body DWI images of patients diagnosed with MM. Dice similarity coefficient (DSC), sensitivity and positive predictive value (PPV) were computed to evaluate the SA performance against the gold standard (GS) and to compare with the radiologists. A non-parametric Wilcoxon test was also performed. Apparent diffusion coefficient (ADC) histogram metrics and lesion volume were extracted for the GS segmentation and for the correctly identified lesions by SA and their correlation was assessed.ResultsThe mean inter-radiologists DSC was 0.323 ± 0.268. The SA vs GS achieved a DSC of 0.274 ± 0.227, sensitivity of 0.764 ± 0.276 and PPV 0.217 ± 0.207. Its distribution was not significantly different from the mean DSC of inter-radiologist segmentation (p = 0.108, Wilcoxon test). ADC and lesion volume intraclass correlation coefficient (ICC) of the GS and of the correctly identified lesions by the SA was 0.996 for the median and 0.894 for the lesion volume (p < 0.001). The duration of the lesion volume segmentation by the SA was, on average, 10.22 ± 0.86 min, per patient.ConclusionsThe SA provides equally reproducible segmentation results when compared to the manual segmentation of radiologists. Thus, the proposed method offers robust and efficient segmentation of MM lesions on WB-DWI. This method may aid accurate assessment of tumor burden and therefore provide insights to treatment response assessment.

Serum Neutrophil Gelatinase-Associated Lipocalin Independently Predicts for Renal Response in Myeloma Patients with Severe Renal Impairment

Introduction: Severe renal impairment (RI) is a common complication of multiple myeloma (MM). Effective anti-MM therapy and supportive care can restore renal function in several patients, but we lack biomarkers that could predict renal outcomes. Neutrophil gelatinase-associated lipocalin (NGAL) is one of the earliest and most robust markers of acute kidney injury while serum Cystatin C (CysC) reflects renal function more accurately than creatinine and correlates with both tumor burden and renal function in MM. GDF-15 is secreted by bone marrow stromal cells and high serum levels indicate a poor treatment prognosis and a high risk of progression to dialysis in AL amyloidosis. Our aim was to evaluate serum NGAL, CysC, and GDF-15 in MM patients with severe RI defined as eGFR&lt;30 ml/min/1.73m2, by CKD-EPI formula, including those on dialysis, as biomarkers for prediction of renal responses. Patients &amp; Methods: MM patients who presented with myeloma-related severe RI in the Department of Clinical Therapeutics (Athens, Greece) were included in this study. NGAL, CysC and GDF-15 were measured in the same frozen serum sample collected before the administration of any therapy. Serum NGAL was measured using ELISA (BioPorto Diagnostics A/S, Gentofte, Denmark), while CysC was measured using a latex particle-enhanced nephelometric immunoassay (Dade Behring-Siemens Healthcare Diagnostics, Liederbach, Germany). GDF-15 was measured by a novel immunoassay, the Elecsys GDF-15 assay (Roche Diagnostics International Ltd, Rotkreuz, Switzerland), which is based on the sandwich immunoassay principle using biotin-streptavidin technology. Patients on dialysis received dialysis with regular membranes. IMWG renal response criteria were used. Results: In total, 101 newly diagnosed MM patients with severe RI were included in this analysis. Median creatinine was 4.6 mg/dl (range 2-&gt;10 mg/dl), median eGFR (ml/min/1.73 m2) was 11.3 (1.3-29.8) and dialysis was required in 35 (35%) patients. The median age was 71 years; median involved FLC was 4740 mg/L; hypercalcemia was found in 24%, LDH ≥ULN in 44% and high-risk cytogenetics in 25% of patients, while 98% of patients were classified as having ISS-3 and 49% as having R-ISS-3 disease stage. Treatment was bortezomib-based in all patients (in 21% VD and in 79% a triplet: VCD or VTD). Median NGAL levels were 182.3 ng/mL (range 20-550 ng/ml) and of CysC were 3.3 mg/L (0.9-8 mg/L); there was a strong correlation between NGAL and CysC levels (R2=0.653, p&lt;0.001). There was also a strong correlation of both CysC and NGAL with eGFR (for CysC: R2=0.615, p&lt;0.001 and for NGAL: R2=0.464, p&lt;0.001). Both NGAL and CysC levels were higher in patients requiring dialysis vs those not in dialysis (median NGAL: 308 vs 166 ng/mL, p&lt;0.002 and median CysC: 4.99 vs 2.83 mg/L, p=0.008). No significant correlations were observed with GDF-15 and the parameters tested. Renal response (Rrenal) was achieved in 63% of patients, including 45% with major Rrenal. Importantly, 40% of patients who required dialysis became dialysis independent. Median time to Rrenal was one month and median time to dialysis independence was two months. Lower levels of NGAL (p=0.0003) and CysC (p=0.03) were associated with higher probability of major Rrenal among patients with severe RI. Baseline eGFR, levels of sFLC and GDF-15 did not correlate with major Rrenal. None of these markers was associated with probability of dialysis independence among patients who required dialysis. Based on ROC analysis, in patients with severe RI (but not on dialysis), NGAL &lt;138 ng/ml was strongly associated with major Rrenal (80% vs 20% at 3 months, p&lt;0.001; Figure). Regarding CysC, levels &lt;2.9 mg/L were associated with higher probability and shorter time to major Rrenal (p=0.012). In multivariate analysis performed in patients not on dialysis, that included age, NGAL, CysC and eGFR, only NGAL &lt;138ng/ml remained a significant prognostic factor associated with major Rrenal (HR 5, 95% CI 2-18, p=0.002). Conclusions: Serum levels of NGAL were independent predictors of major renal response in MM patients with severe RI. Thus, serum NGAL could identify MM patients with severe RI who should be treated with more aggressive therapies and more effective and rapidly acting antimyeloma regimens. The absence of any predictive value of GDF-15 in MM-related severe RI in contrast to AL amyloidosis patients reflect the different biology of RI in these plasma cell dyscrasias. Figure Disclosures Dimopoulos: Sanofi Oncology: Research Funding. Kastritis:Amgen: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Genesis: Honoraria; Prothena: Honoraria; Pfizer: Honoraria; Takeda: Honoraria. Papassotiriou:Roche: Employment. Gavriatopoulou:Amgen: Honoraria; Janssen: Honoraria, Other: Travel expenses; Genesis: Honoraria, Other: Travel expenses; Takeda: Honoraria, Other: Travel expenses. Terpos:Janssen: Honoraria, Other: Travel expenses, Research Funding; Amgen: Honoraria, Research Funding; Celgene: Honoraria; Takeda: Honoraria, Other: Travel expenses, Research Funding; Genesis: Honoraria, Other: Travel expenses, Research Funding; Medison: Honoraria.

Plasma cell-free DNA chromosomal instability score as early predictor to monitor tumor burden in response to therapeutic in multiple myeloma patients.

8036 Background: Multiple myeloma (MM) is a plasma cell malignancy characterized by chromosomal instabilities (CIN). Here we investigate the potential of cell-free DNA CIN as non-invasive biomarker to predict early response for MM treatments. Methods: In this prospective study, we recruited 11 relapsed/refractory (RRMM) and 19 newly diagnosed (NDMM) patients at Changzheng Hospital. Plasma samples were collected after finished two cycles or one month (RRMM) of therapy, with matched ones before the current regimen. cfDNA was extracted, followed by CIN analyses by using a customized bioinformatics workflow, ultrasensitive chromosomal aneuploidy detector (UCAD). Criteria for response and progression were according to the IMWG (Durie BG et al. 2006). Results: 7 (23%) patients (5 RRMM and 2 NDMM) showed high cfDNA CIN regard as strong positive after two cycles of treatment. Plasma cfDNA CIN profiling found complex clonal evolution compared two cycles to baseline. Multiple genomic regions, including chr7, 17p (TP53), 12q and 3p, were involved in clonal evolution. The degree of cfDNA CIN correlated with myeloma stage and overall survival. Remarkably, of the 5 heavily treated RRMM patients and 1 primary refractory newly diagnosed patient, 3 died within 60 days after the last time of cfDNA detection. Nine patients (30%) of patients showed positive cfDNA CIN after two cycles of treatment, which response rate was 11% (n=1) with SD, 33% (n=3) with MR, and 56% with PR, respectively. Fourteen patients with 5 RRMM and 9 NDMM were detected marginal or negative cfDNA CIN after two cycle’s treatment. The overall response rate in 14 patients was 100%, including 14.3% with a complete response, 14.3% with a very good partial response (VGPR), 57.1% with a PR, and 14.3% with a MR. Of these patients, 3 RRMM who received with more than six lines of therapy, showed positive cfDNA CIN. Subsequently, these three heavily treated RRMM patients have chance to enroll the chimeric antigen receptor T-Cell immunotherapy (CAR-T) therapy (enrolled NCT03093168). Surprisedly, all of them benefit from the CAR-T therapy to improve responses dramatically, meanwhile, the dynamics of total cfDNA concentration correlated with tumor burden to negative. Conclusions: We provide evidence that cfDNA level correlates with tumor burden and response rate in MM. For heavily pre-treated advanced RRMM patients with cfDNA CIN positive were benefit from the CAR-T therapy. Therefore, serial plasma cfDNA analysis is a robust and sensitive tool for monitoring response to therapy.

Lack of a Spleen Signal on Diffusion Weighted MRI Is Associated with High Tumor Burden and Poor Prognosis in Multiple Myeloma

▪Background: Whole body medical imaging is an integral component of Multiple Myeloma (MM) evaluation as it reveals bone disease and/or focal lesions. While established imaging techniques, such as PET-CT, are powerful in delineating focal pathologies, they suffer from low specificity when it comes to assessment of diffuse bone marrow (BM) signals. Especially in patients treated with G-CSF, discrimination between malignant and non-malignant BM signals is barely possible. Diffusion-weighted MRI with background suppression (DWIBS) is a novel functional MRI technique which measures water diffusion in vivo. Basically, diffusion of water is more restricted in tissues with high cellularity, making DWIBS a promising tool to investigate malignant diffuse signals. Initially, we searched for an internal reference to classify the level of BM infiltration. We selected the spleen, since it is the abdominal organ with the highest restriction on DWIBS. Unexpectedly, we observed significant heterogeneity in the spleen signal itself including a subgroup of MM patients with total loss of the signal on DWIBS. Perplexingly, these patients suffered from high tumor burden and poor outcome. Here, we report on our strategy to elucidate this phenomenon and to evaluate its clinical value.Methods: We investigated 295 transplant-eligible newly diagnosed MM patients and 72 cases with monoclonal gammopathy of undetermined significance (MGUS). This study was approved by the institutional review board (#205415). DWIBS was performed on a 1.5 Tesla Philips Achieva scanner. The spleen signal was assessed on DWI and ADC maps by two experienced investigators in consensus read. The Kaplan-Meier method was used for survival analyses. Molecular characterization included fluorescence in situ hybridization, and gene expression profiling of CD138-enriched plasma cells (PCs). Differential gene expression was performed using a threshold of 2-fold expression difference. Wilcoxon or Fisher's exact tests were used to compare the median of a continuous variable or the distribution of discrete variables across groups, respectively. Correlation coefficients were determined using Spearman's rank correlation.Results: Absence of the spleen signal on DWIBS was a frequent phenomenon in MM, seen in 71/295 (24%) patients. In all of these patients asplenia was excluded using alternative imaging techniques. Lack of a signal was highly positively associated with tumor-load parameters, such as the degree of BMPC infiltration (P=1x10-10) or International Staging System (ISS) 3 (P=0.0001). In contrast, it was not associated with age, gender, or the tumor progression markers gain(1q) and del(17p). Patients with absence of spleen signal experienced unfavorable outcome (hazard ratio of 1.8 for PFS and OS). In order to further investigate the biological underpinnings of this phenomenon we performed a differential gene expression analysis of purified CD138 MM cells. No differentially expressed genes were found between patients with and without spleen signal, suggesting that the absence of the spleen signal mainly reflected increased tumor burden rather than specific tumor features. As a proof of concept, we addressed the spleen signal in individuals with MGUS, and longitudinally in MM patients who presented with absence of the spleen signal at diagnoses. Indeed, in all 78 individuals with MGUS the signal was preserved, and the majority of MM patients showed re-appearance of the spleen on DWIBS during treatment as the tumor burden declined. Interestingly, re-appearance of the spleen was helpful to distinguish between malignant and non-malignant hyperintensities in the BM, making the spleen signal a promising parameter for MM follow-up investigations.Conclusions: Due to the low frequency of abnormalities affecting the spleen, this organ is often overlooked during abdominal examinations using cross-sectional imaging techniques. Here we show that the spleen signal on DWIBS provides clinically useful information on tumor burden in MM. Moreover, it opens the avenue to distinguish between malignant and reactive BM hypercellularity on imaging in patients receiving treatment. Our observation strongly supports the hypothesis, that the spleen signal is suppressed by a high BMPC involvement. Yet, the reasons for this phenomenon are still elusive. [Display omitted] DisclosuresRoy Choudhury:University of Arkansas for Medical Sciences: Employment, Research Funding. Epstein:University of Arkansas for Medical Sciences: Employment. Barlogie:Dana Farber Cancer Institute: Other: travel stipend; International Workshop on Waldenström's Macroglobulinemia: Other: travel stipend; ComtecMed- World Congress on Controversies in Hematology: Other: travel stipend; European School of Haematology- International Conference on Multiple Myeloma: Other: travel stipend; Celgene: Consultancy, Research Funding; Millenium: Consultancy, Research Funding; Myeloma Health, LLC: Patents & Royalties: : Co-inventor of patents and patent applications related to use of GEP in cancer medicine licensed to Myeloma Health, LLC; Multiple Myeloma Research Foundation: Other: travel stipend. Davies:Amgen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria. Morgan:Janssen: Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding.