ALBERT

Description: Key Points Immune responses to FVIII sequence variants encoded by ns-SNPs do not contribute appreciably to inhibitor development in African Americans. African American HA subjects with an intron-22 inversion had a 2- to 3-times-higher inhibitor incidence than whites with the same mutation.

Description: Juvenile and chronic myelomonocytic leukemias (JMML and CMML) are aggressive myeloid malignancies categorized as myelodysplastic syndromes/myeloproliferative neoplasms (MDS/MPN). Chemotherapy has little benefit for MDS/MPN patients, and new therapies are needed. We have used mouse models investigate the potential of signal transduction inhibitors in MDS/MPN, as JMML and CMML are associated with mutations in NRAS, KRAS, PTPN11, CBL, or NF1 that activate Ras signaling. Conditional Mx1-Cre, KrasLSL-D12 (designated KrasD12) mice develop an aggressive and fully penetrant MDS/MPN characterized by leukocytosis, splenomegaly, anemia, and death by 10-16 weeks of age. Mx1-Cre, Nf1flox/- mice (hereafter Nf1Δ/-) undergo conditional loss of Nf1. These mice also develop MDS/MPN, but the disease is more indolent. We and others have investigated inhibition of effector networks downstream of Ras, such as the Raf/MEK/ERK (MAPK) and phosphotidylinositol-3 kinase (PI3K)/Akt pathways. We previously showed that the MEK inhibitor PD0325901 induced sustained hematologic improvement in both KrasD12 and Nf1Δ/- mice. We also have reported that the class I PI3K inhibitor GDC-0941 improves hematologic function and prolongs survival in KrasD12 mice. However, GDC-0941 and other PI3K inhibitors attenuate both PI3K/Akt and Raf/MEK/ERK pathways due to effects of PI3K upstream of Ras. Therefore, the benefit from GDC-0941 could have been due to its modulation of Raf/MEK/ERK signaling. Here, we specifically test the importance of Akt signaling in MDS/MPN in KrasD12 and Nf1 mouse models using the allosteric inhibitor MK-2206. This compound binds to the interface of the PH and kinase domains of Akt1, Akt2, and Akt3, and does not inhibit any of 250 other kinases at 1 µM. MK-2206 induced substantial improvement in both KrasD12 and Nf1Δ/- mice. Mice treated with MK-2206 had pronounced reduction in leukocytosis, reticulocytosis and splenomegaly, increased hemoglobin concentration, and prolonged survival. MK-2206 had no hematologic effects in control WT mice, indicating some selectivity against aberrant hematopoiesis. Importantly, MK-2206 inhibited Akt but not Raf/MEK/ERK or Jak/STAT signaling. This demonstrates that canonical PI3K/Akt signaling plays an important role in Ras-driven MDS/MPN. Furthermore, combined inhibition of MEK and Akt with PD0325901+MK-2206 yielded a greater improvement in splenomegaly than either agent alone in both KrasD12 and Nf1Δ/- models. Akt has multiple effectors relevant to hematopoiesis and leukemia. Of these, mTOR is of particular interest for targeted cancer therapy. Therefore, we tested the response of KrasD12 mice to rapamycin, a partial inhibitor of mTOR with preferential activity against the mTORC1 complex. KrasD12 mice demonstrated variable responses to rapamycin, with approximately half undergoing a complete and durable hematologic response and the remainder having no response. Together, these studies further implicate PI3K/Akt signaling as a pathogenic effector downstream of Ras in MDS/MPN and support the idea that inhibitors targeting this pathway may have a role in treatment of JMML or CMML. Disclosures No relevant conflicts of interest to declare.

Description: Introduction We previously reported on the generation of highly activated/expanded natural killer cells (ENKs) after coculture with K562 cells modified to express membrane bound IL15 and 41BB-ligand. These cells have potent antimyeloma properties in vitro, in a NGS mouse model, and are safe when given to advanced multiple myeloma (MM) patients. (Szmania et al, J Immunother 2015) A potential obstacle to the effectiveness of ENK-based immunotherapy of MM is the evasion of immune recognition. We have generated 4 MM cell lines (OPM2, JJN3, ANBL6, and INA-6) which are resistant to ENK-mediated lysis to study mechanisms of resistance. These lines were derived from parental lines by repeated challenge with ENKs and maintained resistance long term when cultured without further exposure to ENKs.(Garg et al, Blood 2012, 120:4020) We have shown by stable isotope labeling with amino acids in cell culture-mass spectrometry, gene expression profiling (GEP), and flow cytometry that ICAM3 is downregulated in the ENK-resistant version of OPM2 (OPM2-R) compared to the parental OPM2. (OPM2-P; Garg et al, Blood 2013, 122:3105) We investigated OPM2-P and OPM2-R by whole exome sequencing (WES) and RNA sequencing (RNAseq) with a focus on ICAM3, evaluated ICAM3 cell surface expression on patient myeloma cells, and studied the importance of ICAM3 expression on ENK functionality. Methods DNA and RNA were extracted from OPM2-P and OPM2-R cells using the Qiagen AllPrep kit. WES libraries were prepared with the Agilent qXT and Agilent SureSelect Clinical Research Exome kits with additional baits covering the Ig and MYC loci. RNAseq libraries were prepared using the Illumina TruSeq stranded mRNA kit. Samples were sequenced 100bp PE on an Illumina HiSeq2500. Samples for WES were sequenced to a mean coverage of 〉120x and RNAseq to a target of 〉100M reads. WES data were aligned to the Ensembl GRCh37/hg19 human reference using BWA mem. Somatic variants were called MuTect. RNAseq data were analyzed using Tuxedo Suite. Data were aligned to the Ensembl GRCh37/hg19 human reference using TopHat with Bowtie2. Transcriptome reconstruction, quantification and differential analysis was performed using CuffLinks. ENK-mediated lysis of myeloma cells was measured by 4 hour chromium release assay in the presence of isotype or ICAM3 blocking antibody. Bone marrow aspirates were obtained from MM patients after informed consent in accordance with the Declaration of Helsinki. Primary myeloma cells were selected with CD138-coated immunomagnetic beads and ICAM3 expression was assessed by flow cytometry gated on viable CD138 positive cells. Results There was no mutation in ICAM3 in OPM2-R by WES, but RNAseq found a significant reduction in ICAM3 RNA in OPM2-R compared to OPM2-P (p

Description: Introduction : Multiple myeloma (MM) patients with t(14;16)(p16;q32) leading to high expression of C-MAF mRNA and protein have a poor prognosis and have not benefitted from the recent advances in MM therapy. Our previous work showed that posttranslational modification up-regulated C-MAF (ASH Abstract # 281, ASH 2013) and MAF-B proteins (ASH Abstract #2091, 2014) conferred innate resistance to proteasome inhibitors (PI). In order to identify the molecular mechanism underlying this resistance in high C-MAF and MAFB-expressing patients, the present study investigated the downstream targets regulated by C-MAF in MM cells and analyzed the clinical association of target genes using a C-MAF-loss and gain of function cell line model as well as primary plasma cells from MM patients. Methods : To elucidate the molecular mechanism underlying the resistance of MAF expressing cells to PIs, we silenced the C-MAF gene in 3 MM cell lines that highly expressed C-MAF mRNA and protein using lentiviral shRNA. Annexin V staining and analyzed by FACS were used to determine the effect of manipulating MAF on PI-induced apoptosis and activation of the caspase family. Gene expression prolife (GEP) was utilized to determine downstream target genes regulated by MAF. To identify the proteins associated with C-MAF, we used mass spectrometry (MS)-based quantitative proteomics with stable isotope labeling by amino acids in cell culture (SILAC). Results: Our results demonstrated that apoptotic cell numbers were higher in shMAF cells, compared to shControl cells when cultured in the presence of serial concentrations of bortezomib (BZB) or carfilzomib (CFZ), indicating that silencing MAF expression enhances PI-induced apoptosis. The Annexin V + cells numbers were also higher in shMAF compared to shControl when cells were treated with BZB or CFZ in co-culture with bone marrow stromal cells. Furthermore, knockdown of C-MAF expression augmented the PI-induced activation of the caspase family with an increase in the cleavage fragments of caspases-3, -7, -8, PARP and LaminA/C. In contrast, PI-induced apoptotic numbers were lower in XG1/MAF, a stably induced MAF overexpressing MM cell line, than in XG1/EV, the control cell line carrying an empty vector when cultured in serial concentrations of BZB and CFZ. Similarly, the caspase cleavage fragments, PARP and lamin A/C were decreased in XG1/MAF compared with XG1/EV in response to BZB and CFZ. These results suggest that MAF confers MM resistance to PI by preventing the PI-induction of apoptosis. To further identify the downstream target genes regulated by C-MAF, we analyzed the gene expression between shMAF and shControl MM cells by microarray analysis and identified more than 150 genes differentially expressed. Of them, ATP2C1, CCNE2 , CCND2, ELK4 , IGFBP3, SUPT16 and TUBA1A were significantly (p=5.62e-5) lower in shMAF than shControl cells indicating that these genes might be regulated by C-MAF. Furthermore, expression of these genes in primary CD138 plasma cells were significant higher (p

Description: Introduction: Despite the enormous progress in MM therapy brought about by the rapid development of many novel agents, many patients end up with limited treatment options. We have previously reported on the efficacy and safety of metro16 in RRMM (Papanikolaou, Haematology 2013). Here we are reporting on an extension of such treatment to 28 d (metro28). Patients and Methods: The treatment consisted of a cycle of 28d continuous iv infusions of ADR and DDP each at 1mg/m2/d, along with thalidomide 50 to 100mg/d x 28; bortezomib 0.8 to 1.0mg/m2 on days 1, 4, 7, 10, 13, 16, 19, 22, 25, 28; DEX 8 to 12mg on days 1-4, 7-10, 13-16, 19-22, 25-28; some patients also received vincristine 0.07mg flat dose by CI for 28 days. This was off-protocol therapy that patients provided written informed consent for. The IRB permitted data retrieval and analysis. Results: 150 patients were identified, virtually all had received prior tandem transplants, bortezomib, lenalidomide, carfilzomib and pomalidomide. The median age was 64yr; B2M was elevated 〉=3.5mg/L in 48%, abnormal cytogenetics (CA) were present in 86%, and 44% had GEP70-defined high risk MM. Figure 1 portrays clinical outcomes. As of April 2015, 60 patients had died, and the 2-yr OS estimate was 45% (Figure 1A); the 6-mo PFS estimate was 31% although 15% had no progression at 18mo (Figure 1B). Analysis by GEP70 and GEP5 risk revealed 18-mo OS estimates of 80% among the 53 patients with low risk in both models, whereas the presence of high risk (HRMM) in either model conferred a significantly reduced 18-mo OS estimate of 25% (p

Description: Introduction Over the last 15 years gene expression profiling (GEP) has been used to define myeloma molecular subgroups and to determine clinical prognosis. Two major molecular subgroup classifications have been used: the UAMS which determines 7 subgroups and the TC classification based on the presence of IgH translocations and expression of D group cyclins. For prognosis, although a number of different GEP signatures have been defined, the widely used GEP70 identifies 15% of patients with high risk (HR) disease who have a median PFS and OS of 1.75 and 2.83 years. An ideal classification system would identify clinically relevant subgroups with distinct etiology and biology using standardized techniques. We have examined a large group of patients characterized at multiple genetic levels to optimize the diagnostic approach of newly diagnosed patients going forward. Materials and methods Study subjects included 1349 cases enrolled in Total Therapy trials (median follow up 7.5 years). Gene expression profiling was used to determine GEP70 risk status, molecular subgroup by UAMS and TC classifications, and to devise a new and extended TC classification (TC10). Interphase FISH associated with IgH translocations and 1q+ and 17p- were used to build GEP proxies. Data from mutational analysis generated by the FoundationOne targeted sequence panel was also incorporated. Results were validated on the UK MRC MyelomaIX and Hovon65/GMMG-4 studies. Results An initial agnostic analysis of GEP data using sparse k-means clustering verified the existence of TC based groups. Six groups were identified that corresponded overwhelmingly with known TC subgroups; CCND1-t(11;14), D1-HRD, D2-HRD, MMSET, MAF/CCND2, and CCND3. Further comparisons between the molecular subgroup and TC classifier revealed that the UAMS 7 subgroups clustered strongly within one predominant TC group: CD-1 and CD-2 to t(11;14), HY to D1, LB and PR to D2, MF to t(14;16) or t(14;20), and MS to t(4;14). As the UAMS molecular subgroups are largely contained within the TC framework, we aimed to extend the TC by developing the TC10. To extend the known TC subgroups, unsupervised clustering was applied to the 3 largest subgroups [t(11;14), D1, and D2] to determine the strongest single divisor within each respective subgroup. The dominant feature within the t(11;14) cases was CD20 expression, while the D1 and D2 subgroups both split according to RRAS2. CD20 is associated with PAX5 and VPREB3 expression, and RRAS2 is associated with decreased PTP4A3 and increased TNFAIP3 and BIRC3 expression. RRAS2 activation within D1 subgroup and CD20 activation within t(11;14) cases corresponds to an increased time to response to induction therapy suggesting they constitute important biological subgroups. The TC10 combines the known etiologic subgroups of the TC with functionally relevant subdivisions to create 10 novel subgroups: t(11;14) CD20+/-, D1: RRAS2+/-, D2: RRAS2+/-, t(4;14), t(14;16), t(14;20), and t(6;14). Analysis of mutational data revealed that RRAS2 and CD20 activation within the D1, D2, and t(11;14) subgroups reduced the number of mutations in the MAPK pathway. Further mutational analysis revealed that median mutational load was highest in t(14;16) and lowest in D2: RRAS2+ subgroups. The GEP70 score identifies 15% of patients with HR disease and is specific for this purpose. In an analysis of risk assessment methods, we compared GEP detected adverse lesions [t(4;14), t(14;16), t(14;20), 17p- and 1q+] with the GEP70 and revealed that GEP70 HR identified samples have lower OS rates than cases with more than one adverse lesion (validated in external sets). GEP70 HR segregates non-uniformly across molecular subgroups as over 40% of all HR cases are found in the TC10 t(4;14), t(14;16), and t(14;20) subgroups. GEP70 HR cases also have a higher mutational load than low risk cases. Furthermore, GEP70 HR is uniquely associated with 1q+ and 17p- as cases with at least one of these adverse lesions are 4.9 times as likely to be GEP70 HR as cases with neither. Conclusion GEP profiling has a central role in simplifying and standardizing the molecular subgroup designation and risk stratifying of MM patients. The GEP70 risk score reliably identifies HR cases and outperforms FISH in risk assessment, even in validation data sets. The TC10 provides a classification system that improves upon previous methods by defining both etiological and functionally meaningful subgroups. Disclosures Stein: University of Arkansas for Medical Sciences: Employment. Davies:University of Arkansas for Medical Sciences: Employment; Celgene: Consultancy; Janssen: Consultancy; Millenium: Consultancy; Onyx: Consultancy. Heuck:University of Arkansas for Medical Sciences: Employment; Celgene: Consultancy; Janssen: Other: Advisory Board; Millenium: Other: Advisory Board; Foundation Medicine: Honoraria. Weinhold:University of Arkansas for Medical Sciences: Employment; Janssen Cilag: Other: Advisory Board. Chavan:University of Arkansas for Medical Sciences: Employment. Thanendrarajan:University of Arkansas for Medical Sciences: Employment. Epstein:University of Arkansas for Medical Sciences: Employment. Yaccoby:University of Arkansas for Medical Sciences: Employment. Zangari:University of Arkansas for Medical Sciences: Employment; Novartis: Research Funding; Onyx: Research Funding; Millennium: Research Funding. van Rhee:University of Arkansa for Medical Sciences: Employment. Kaiser:Janssen: Honoraria; Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; BristolMyerSquibb: Consultancy; Chugai: Consultancy. Sonneveld:Janssen-Cilag, Celgene, Onyx, Karyopharm: Honoraria, Research Funding; novartis: Honoraria. Goldschmidt:Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Onyx: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen-Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Millenium: Honoraria, Research Funding, Speakers Bureau; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Chugai: Honoraria, Research Funding, Speakers Bureau; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Barlogie:University of Arkansas for Medical Sciences: Employment. Morgan:Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; MMRF: Honoraria; CancerNet: Honoraria; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; University of Arkansas for Medical Sciences: Employment; Weismann Institute: Honoraria; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Description: Introduction: Despite major advances in MM therapy with the inclusion of novel agent combinations for induction prior to and after autotransplant-supported high-dose melphalan, the 15% of patients with GEP-defined HRMM continue to fare poorly with PFS and OS not exceeding 2 and 3 years, respectively. This poor outcome has not been improved with less dose-intense and more dose-dense Total Therapy 5. Having previously reported on 16-day metronomic therapy with low-dose doxorubicin (DOX) and cisplatin (DDP) plus VTD (Papanikolaou, Haematologica), we explored further extension of such metronomic treatment to 28 days (metro-28) also in newly diagnosed HRMM patients. Patients and Methods: All patients signed a written informed consent and data analysis was approved by our IRB. In the outpatient setting, a single cycle of metro-28 comprised DOX and DDP each at 1.0mg/m2/d for 28d by continuous infusion (CI), along with VTD (bortezomib 1.0mg/m2 on days 1-4, 7-10, 13-16, 19-22, 25-28; DEX 12mg on days 1-4, 7-10, 13-16, 19-22, 25-28; thalidomide 50-100mg/d x 28d; some patients also received vincristine [VCR] at a flat daily dose of 0.07mg/d x 28d by CI. Results: Fourteen patients were initiated on metro-28. Their characteristics included age 〉=65y in 12; albumin 5.5mg/L in 7; cytogenetic abnormalities [CA] were present in 10; GEP70 HRMM in 9/13; PR subgroup in 8/13 (Table 1). The median follow up is 11mo. As portrayed in Figure 1A, no patient has died; the 6mo PFS estimate was 85% (Figure 1B); responses included CR in 3/14, VGPR in 7/14 and PR in 10/14 (Figure 1C); and the PR duration estimate at 6mo is 80% (Figure 1D). Of interest, GEP70 scores morphed to low risk in 3/13. Vascular density (CD34) decreased markedly in most patients evaluated. Toxicities were minor; myelosuppression was virtually absent; alopecia was not encountered. Subsequent salvage therapies included repeat metro-28, combination chemotherapy (PACMED) and autotransplants. Conclusion: We conclude that metro-28 is a promising and safe strategy for elderly patients with HRMM, and we hypothesize an anti-angiogenic mechanism of action in addition to direct anti-MM effects. Table 1. Patient characteristics Factor n/N (%) Age 〉= 65 yr 12/14 (86%) Albumin 〈 3.5 g/dL 8/14 (57%) B2M 〉= 3.5 mg/L 9/12 (75%) B2M 〉 5.5 mg/L 7/12 (58%) Hb 〈 10 g/dL 10/14 (71%) Cytogenetic Abnormalities 10/14 (71%) CA within 1 Year of Therapy 10/14 (71%) CA within 90 Days of Therapy 9/14 (64%) GEP 70-Gene High Risk 9/13 (69%) GEP PR Subgroup 8/13 (62%) GEP Proliferation Index 〉= 10 7/13 (54%) GEP Centrosome Index 〉= 3 7/13 (54%) n/N (%): n- Number with factor, N- Number with valid data for factor Figure 1. Figure 1. Disclosures Thanendrarajan: University of Arkansas for Medical Sciences: Employment. Alapat:University of Arkansas for Medical Sciences: Employment. Zangari:University of Arkansas for Medical Sciences: Employment; Onyx: Research Funding; Millennium: Research Funding; Novartis: Research Funding. Schinke:University of Arkansas for Medical Sciences: Employment. Heuck:Millenium: Other: Advisory Board; Janssen: Other: Advisory Board; Foundation Medicine: Honoraria; Celgene: Consultancy; University of Arkansas for Medical Sciences: Employment. van Rhee:University of Arkansa for Medical Sciences: Employment. Rosenthal:Cancer Research and Biostatistics: Employment. Epstein:University of Arkansas for Medical Sciences: Employment. Yaccoby:University of Arkansas for Medical Sciences: Employment. Davies:Janssen: Consultancy; Onyx: Consultancy; University of Arkansas for Medical Sciences: Employment; Millenium: Consultancy; Celgene: Consultancy. Morgan:MMRF: Honoraria; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; CancerNet: Honoraria; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Weismann Institute: Honoraria; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; University of Arkansas for Medical Sciences: Employment. Barlogie:University of Arkansas for Medical Sciences: Employment.

Description: Introduction Achieving complete remission (CR) improves survival outcomes in multiple myeloma (MM). Applying more sensitive flow-cytometry or PCR based minimal residual disease (MRD) testing further improves prognostication and MRD negativity confers better outcomes among CR patients. In our Total Therapy (TT) trials, gene expression profiling based risk (GEP) identifies 15% patients with high risk (HR) MM whose clinical course is characterized by early relapsing disease and prognosis has remained grim despite advances in therapy. Of interest is whether high-risk (HR) patients fail to achieve MRD negativity leading to relapse. We have determined response rates across the total therapy (TT) trials and in this study we report the impact of molecular negativity by MRD assessment in patients who have achieved at least very good partial response (VGPR) or CR at 4-8 months and 12-24 months, respectively, after enrollment in TT3b-TT5 from 2005-2009. We use next-generation sequencing (NGS)-based MRD assessment (Adaptive Biotechnologies) which is sensitive to 1 MM cell in 106 normal cells. Furthermore, we evaluate MRD status in long term relapse-free survivors (〉6 years) to determine the importance of MRD status in prolonged remission. Materials and methods Study subjects include 591 patients enrolled in TT3b-TT5 to determine VGPR/nCR/CR by EBMT/IMWG criteria and to analyze PFS and OS of HR and SR patients defined by our GEP model. For MRD testing, we identified 102 patients from the TT3b-TT6 protocols that achieved at least VGPR and had bone marrow (BM) sample available at 4-8 months and 12-24 months after enrollment. Of these 102 patients, 14 patients had HR disease and 88 had SR. Six of 14 HR patients already had clinical relapse between 12-24 months, but were included to determine MRD at 4-8 months. Of all 102 patients, 51 are currently still in CR 6-9 years after protocol enrollment and most recent flow-cytometric MRD status was evaluated. NGS-based MRD Assessment: Genomic DNA was amplified using locus-specific primer sets for immunoglobulin heavy-chain complete (IGH-VDJH) and incomplete (IGH-VDH) as well as for immunoglobulin κ locus (IGκ). The amplified products were subjected to sequencing and clonal gene rearrangements were analyzed. Final MRD measurement was calculated at a sensitivity level of 1 cancer cell per 1 million cell equivalents. Flow Cytometry based MRD assessment: Erythrocyte-lysed BM samples were immunophenotyped by use of 8-color (CD138/CD38/CD19/CD45/CD27/CD81/CD56/CD20) staining technique. Myelomatous Plasmacells (PCs) were separated from healthy PCs by different phenotype expression and MRD was deemed negative when no myelomatous PCs were detectable at a sensitivity of

Description: Background: Tumor cells exhibit elevated ROS levels and cancer stem cells have lower ROS levels than their normal counterparts [Kobayashi et al., 2011]. However, little is known about the effect of ROS levels on multiple myeloma (MM) plasma cell (PC) chemosensitivity. Previously we found that ROS levels are variable in CD138-selected MMPCs from different cases at diagnosis. However, ROS levels in MMPCs from focal lesions (FL, n=42) are lower than PCs from random bone marrow (BM) aspirates (n=192, p=0.02). ROS levels also vary among MM molecular subgroups, with MF and CD2 subgroups having low ROS levels, suggesting that the microenvironment and genetic factors affect their oxidative capacities. We hypothesized that ROS level may reflect the cancer stemness properties of MM cells. We first sought to establish a model to investigate MM cells with different ROS levels and then determine if this property confers cancer stemness and chemoresistance. Methods: We analyzed gene expression profiles (GEP) of primary myeloma cells from newly diagnosed patients with low (n=48) and high (n=48) ROS level by gene set enrichment analysis. We compared viability, proliferation, cell cycle status, drug efflux ability, chemosensitivity, and clonogenic capacity of MM cell lines H929 and U266 with high and low ROS levels, sorted by FACS using H2DCFDA. We also investigated whether MMPCs with low or high ROS levels would differ in their sensitivity to BBI-608, a drug that is reported to target STAT3 driven gene transcription and cancer stemness properties in solid cancers [Li et al., 2015] and is currently being tested in hematologic cancers [BBI608-103HEME]. Results: Pathways enrichment analysis revealed that primary MM cases with low ROS had increased oxidative phosphorylation, proteasome activity, and nucleotide excision repair compared to cases with high ROS that had increased glycolysis, and chemokine and calcium signaling pathways. To explore these differences we separated MMCLs into cells not sorted by ROS level (non-ROS sorted), ROSLo, and ROSHi groups. After sorting, H929 cells retained significant differences in ROS levels for 48-72 hr and U266 cells returned to basal ROS levels within 24hr. ROSLo H929 cells had significantly reduced cell number (p

Description: Introduction: Black patients with acute myeloid leukemia (AML) have inferior overall survival relative to White patients. Few studies have evaluated differences in induction mortality and none has assessed the contribution of severity of illness at presentation to the disparity in survival. Our primary objectives were to compare induction mortality and acuity of presentation among Black relative to White patients and to assess whether any disparity in induction mortality is the consequence of differences in presentation acuity. In addition, we explored the interaction between Black race and public insurance on induction mortality with use of single referent models. Methods: Using a retrospective cohort of children (ages 0 to 18 years) from 2004 to 2014 with new-onset AML diagnosed and treated at free-standing pediatric hospitals who contribute inpatient information to the Pediatric Health Information System administrative database (PHIS), we evaluated inpatient mortality over two courses of standard induction chemotherapy. We examined race (Black versus White) as the primary exposure and insurance was considered with race using a common reference group. We also considered Intensive Care Unit (ICU)-level resource use during the first 72 hours following the initial AML admission as a surrogate for acuity at presentation and a potential mediator of the association between race and induction mortality. Results: 1,122 patients (183 Black, 939 White) with AML who received standard induction chemotherapy were included. Induction mortality for Blacks was substantially higher than for Whites (cHR= 2.31, 95% CI: 1.01, 5.42). Blacks also had a significantly higher risk of requiring any ICU-level care within the first 72 hours after initial presentation compared with Whites (cHR= 1.52, 95% CI: 1.02, 2.24).The association between race and induction mortality was attenuated following adjustment for ICU-level care within the initial 72 hours after admission, (aHR=1.42, 95% CI: 0.67, 2.99). Publicly insured patients experienced greater induction mortality than privately insured patients regardless of race. Induction mortality rates for Black and White patients were more similar among the privately insured and were increasingly disparate among the publicly insured. Conclusion: Our findings suggest that Black patients with AML present with more acute illness at initial diagnosis, accounting for up to 63% of the relative excess induction mortality. Identifying factors impacting acuity of illness at presentation and associated with public insurance may help to identify opportunities for intervention and thus narrow the current racial disparities in pediatric AML survival. Table 1. Inpatient Induction Mortality and ICU level Care by Race Outcome, Follow-up Period Overall (N=1122) n (%) Black (n=183) n (%) White (n=939) n (%) cHR (95% CI) aHRa (95% CI) Induction Death 27 (2.4%) 8 (4.4%) 19 (2.0%) 2.31 (1.01, 5.42) 1.42 (0.67, 2.99) Any ICU Level Care in first 72 hrs 135 (12.0%) 31 (16.9%) 104 (11.1%) 1.52 (1.04, 2.24) ICU involving 〉1 system in first 72 hrs 47 (4.2%) 18 (9.8%) 29 (3.1%) 3.35 (1.84, 6.12) Any ICU Level Care in Induction 237 (21.1%) 48 (26.2%) 189 (20.1%) 1.30 (0.99, 1.71) 1.09 (0.74, 1.61) ICU involving 〉1 system in Induction 99 (8.8%) 22 (12.0%) 77 (8.2%) 1.42 (0.85, 2.38) 0.92 (0.54, 1.57) a adjusted for ICU acuity score within the first 72 hours of index admission Figure 1. Independent and joint effects of Black race and public insurance on induction mortality Figure 1. Independent and joint effects of Black race and public insurance on induction mortality Disclosures Wilkes: Alex's Lemonade Stand Foundation: Research Funding; Healthcare Research and Quality: Research Funding. Fisher:Merck: Research Funding; Pfizer: Research Funding. Epstein:Medicus Economics: Consultancy. Aplenc:Sigma Tau: Consultancy.