ALBERT

Description: Individual microbial species are known to occupy distinct metabolic niches within multi-species communities. However, it has remained largely unclear whether metabolic specialization can similarly occur within a clonal bacterial population. More specifically, it is not clear what functions such specialization could provide and how specialization could be coordinated dynamically. Here, we show that exponentially growing Bacillus subtilis cultures divide into distinct interacting metabolic subpopulations, including one population that produces acetate, and another population that differentially expresses metabolic genes for the production of acetoin, a pH-neutral storage molecule. These subpopulations exhibit distinct growth rates and dynamic interconversion between states. Furthermore, acetate concentration influences the relative sizes of the different subpopulations. These results show that clonal populations can use metabolic specialization to control the environment through a process of dynamic, environmentally-sensitive state-switching.

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: 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: Gene expression profiling (GEP)-defined high-risk de novo multiple myeloma (HI-MM) has a dismal prognosis with median PFS and OS stagnating at 2 and 3 years, respectively, despite the incorporation of novel agents into our Total Therapy (TT) trials. Having seen encouraging results in relapsed-refractory MM with an extended 16-day metronomic therapy (Papanikolaou, Haematologica 2014), we tested this approach in a small cohort of untreated patients with HI-MM. METRO emphasizes targeting neo-angiogenesis and other components of the bone marrow micro-environment while avoiding cytokine surges with recovering hematopoiesis following myelotoxic therapy. Patients and Methods: 10 previously untreated patients with HI-MM, who were either ineligible or unwilling for our Total Therapy protocols received a single cycle of METRO. Therapy comprised of SC bortezomib 1.0mg/m2 (0.8mg/m2 in case of grade 〉2 peripheral neuropathy) on days 1, 4, 7, 10, 13, 16, 19, 22, 25 and 28 schedule, PO dexamethasone 12mg (8mg in case of prior intolerance of higher dose or diabetes mellitus) on days 1 to 4, 7 to 10, 13 to 16, 19 to 22 and 25 to 28, PO Thalidomide 100mg (50mg in case of peripheral neuropathy grade 〉2) and continuous IV infusions of doxorubicin and cisplatin at 1.0mg/m2 daily for 28 days. Cisplatin was dose-reduced for Cr 〉2mg/dL and omitted for Cr 〉3mg/dL. Arsenic tri-oxide was given at a fixed dose of 0.01mg/kg on the days after bortezomib. Laboratory monitoring for response and toxicities were done on a Monday-Wednesday-Friday schedule. Maximal responses, based on current IMWG definitions, were measured within 30 days of completion of cycle 1, and at least monthly thereafter. KM curves were current as of 07/31/14. The Institutional Review Board granted permission for our retrospective data review, the results of which are presented here. Results: Patient characteristics included age 〉=65 in 8, 5 male, 5 female with ISS III in 5 patients. Metaphase cytogenetic abnormalities (CA) were detected in 7 patients. GEP70 based high risk MM was present in all 10 patients, and GEP proliferation (PR) subgroup was dominant in 8 out of 10 patients. All 10 patients achieved at least PR, including 3 qualifying for VGPR and 4 for CR. Bone Marrow responses were equally encouraging in that 8 of 10 patients qualified for complete morphologic negativity including 4 with no minimal residual disease (MRD) by 8-color flow cytometry. Of 8 patients with FDG-avid PET-CT focal lesions, 6 achieved PET-CT CR; all patients showed decreases in SUV-max and SUV-diff (background SUV). Number and/or apparent diffusion coefficient (ADC) mapping of focal lesions and background marrow on diffusion-weighted MRI improved in all 7 evaluable patients. GEP70 risk morphed from high risk to low risk in 3/4 evaluable patients. Pre and post serologic, urinary and radiologic responses are shown in Figure 1. The median follow-up time for the population was 3.2 months (98 days). All 10 patients are alive from 1 to 7 months, and 1 suffered progression (Figure 2). Overall tolerance was good. Non-hematologic grade 3/4 SEs included fatigue, electrolyte abnormalities (20%), dyspnea, hypotension, LE edema and transaminitis (10%). Conclusion: Primary 28-day metronomic therapy is highly effective and well-tolerated in patients with previously untreated HI-MM. Further prospective studies with longer follow-up are currently being devised in an attempt to improve outcomes in this population. Figure 1: Individual responses Figure 1:. Individual responses Figure 2 Figure 2. Figure 3 Figure 3. Disclosures van Rhee: Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Millenium: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees. Zangari:Norvartis: Membership on an entity's Board of Directors or advisory committees; Onyx: Research Funding; Millennium: Research Funding.

Description: Background: The definition of high risk smoldering multiple myeloma (HR-SMM) is in flux. There are several models using serologic, bone marrow and radiologic data that predict for time to progression (TTP) to clinical myeloma (CMM). Lenalidomide and Dexamethasone in HR-SMM is reported to delay onset of end organ damage and improve overall survival, stressing the clinical utility of early intervention. We previously reported a GEP70 based score cutoff (-0.26, serum M spike ≥3g/dL, and involved SFLC 〉25 mg/dL identified a subset of patients with 67% risk of progression at 2 years. With longer follow up, we now examine whether unique gene probe sets can be identified at the AMM stage that portend an earlier time to therapy (TTT). Patients and Methods: We identified 105 patients with AMM who had baseline GEP data on our S0120 protocol, after IRB approval for retrospective data review, and evaluated each of 54,675 Affymetrix gene probes for their potential to predict TTT. Probes were ranked by their q-values; we found 40 probes with q-value 〈 0.05 and 7 probes with q-value 〈 0.01; the top probe had a q-value of 0.00066. Scores based on the number of significant probes at these cut-points were computed by subtracting the sum of the expressions of the up-regulated probes from the sum of the expressions of the down-regulated probes, then dividing by the total number of probes. Results: In the GEP40 model, an optimal cut-point for risk of progression was identified at 7.05. The 3-year TTT probability was 83% with scores 〉=7.05 and only 11% for patients with values under this threshold (Figure 1A; p65 (HR: 2.3), Albumin3g/dl (HR: 4.99), BM plasmacytosis〉=10% (HR: 12.2), GEP70〉-0.26 (HR: 3.4), GEP40〉=7.05 (HR: 16.41), GEP proliferation index 〉 -0.26 (HR: 2.8), GEP PR subgroup (HR: 9.4) and GEP PolyPC 〉11.6 (HR: 0.22) to be significant. In the multivariate model, GEP40〉=7.05 was the most significant (HR: 13.7), followed by SFLC〉10mg/dl and M-protein〉3g/dl. GEP40 score positively correlated with proliferation index (R: 0.804), and showed no correlation with GEP polyPC score (R: -0.156). Next, we used recursive partitioning on data from 72 patients and identified 23 patients with GEP40 score 〉=7.05 of whom 22 suffered TTT by 3 years (87%). Among the remaining 49 patients with GEP40 59 years identified 24 patients, of whom 11 suffered progression with a 3 year TTT estimate of 25%. In the 25 patients with GEP40

Description: Progression of Asymptomatic Monoclonal Gammopathies to Myeloma requiring treatment -Clinical Multiple Myeloma (CMM)- is an important issue in current clinical investigation toward secondary prevention, i.e. treating high-risk AMG. Several predictive models have been published, including one that incorporated gene expression profiling (GEP) of plasma cells (PC) where a GEP70 score ≥0.26 was linked to higher AMG-CMM progression in a multivariate model (Dhodapkar, Blood 2014). We have applied 2-parameter flow cytometry of DNA and cytoplasmic immunoglobulin (FDC) of bone marrow aspirates as part of baseline staging of all patients with plasma cell dyscrasia. A modification introduced in August 2006 on the doublet discrimination method increased accuracy and reproducibility of FDC results considerably and allowed for the detection of a plasma cell population with a low CI17% (HR: 6.76, P

Description: Despite the increasing availability of novel agents for RRMM, many patients run out of treatment options due to disease refractoriness and/or progressive toxicities, both hematological (cytopenia) and non-hematological (cachexia, asthenia, renal and cardiopulmonary).Patients who are refractory to both proteasome inhibitors and immunomodulatory agents (IMIDs) carry an ominous prognosis with a median overall survival (OS) of 9 months (Leukemia, Kumar 2013). We previously reported on the efficacy and toxicity profiles of a metronomically scheduled chemotherapy regimen administered over the course of 16 days (Haematologica, Papanikolaou 2014). Encouraged by the results of that treatment, we have since modified metronomic chemotherapy to cover a 28-day period (metro28). The regimen consisted largely of bortezomib 1.0mg/m2 on days 1, 4, 7, 10, 13,16,19, 22,25,28; along with dexamethasone 12mg on days 1-4, 7-10, 13-16, 19-22, 25-28; thalidomide 100mg days 1-28, continuous 28 day infusion of doxorubicin 1.0mg/m2 (0.5mg/m2 for Ejection Fraction 2mg/dL), with the addition of arsenic trioxide (ATO) at 0.1mg/kg on each day after bortezomib; and lately vincristine (VCR) at 0.07mg daily dose by continuous infusion for 28 days. Patient characteristics included age 〉65yr, 46%; female, 33%; cytogenetic abnormalities at any time prior to therapy (CA), 86%; Gene Expression Profile (GEP) 70-defined high risk, 44%. The median number of prior therapies was 8 (1-64), prior transplants, 83% (1, 2, 〉=3), 20%, 32%, 31%; prior exposure to novel agents including Carfilzomib/Pomalidomide was 81%. Thrombocytopenia prior to treatment was present in 75% including thrombocytopenia = 8 mg/L 54/122 (44%) Creatinine 〉= 2 mg/dL 15/122 (12%) Hb 〈 10 g/dL 57/122 (47%) LDH 〉= 190 U/L 36/121 (30%) Platelet Count 〈 150 x 10^9/L 92/122 (75%) Cytogenetic abnormalities 105/122 (86%) CA within 1 year of therapy 84/121 (69%) CA within 90 days of therapy 65/119 (55%) New Kit GEP Sample 91/91 (100%) GEP 70 High Risk 40/91 (44%) GEP 5 High Risk 39/91 (43%) GEP CD-1 subgroup 5/91 (5%) GEP CD-2 subgroup 17/91 (19%) GEP HY subgroup 14/91 (15%) GEP LB subgroup 6/91 (7%) GEP MF subgroup 9/91 (10%) GEP MS subgroup 9/91 (10%) GEP PR subgroup 31/91 (34%) GEP proliferation index 〉= 10 28/91 (31%) GEP centrosome index 〉= 3 26/91 (29%) n/N (%): n- Number with factor, N- Number with valid data for factor ND: No valid observations for factor Figure 1A Figure 1A. Figure 1B Figure 1B. Figure 1C: OS by GEP5 Figure 1C:. OS by GEP5 Disclosures Zangari: Norvartis: Membership on an entity's Board of Directors or advisory committees; Onyx: Research Funding; Millennium: Research Funding. van Rhee:Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Millenium: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees. Morgan:Celgene Corp: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Myeloma UK: Membership on an entity's Board of Directors or advisory committees; International Myeloma Foundation: Membership on an entity's Board of Directors or advisory committees; The Binding Site: Membership on an entity's Board of Directors or advisory committees; MMRF: Membership on an entity's Board of Directors or advisory committees.

Description: Prolonged survival of myeloma patients on Total Therapy regimens including IMIDS and novel agents has been associated with increased incidence of treatment-related myelodysplastic syndrome and acute leukemia (t-MDS/AL). MDS cytogenetic abnormalities (MDS-CAs) are often observed prior to t-MDS/AL development. Among 1,080 patients on TT2 and TT3 protocols, MDS-CA occurred in 11% and t-MDS/AML in 3%. Risk features of MDS-CA included TT3b treatment, age ³65 yr, male sex, elevated B2M, and MM relapse. Lower doses of CD34 HSCs applied with first transplants raised the probability of MDS-CAs, and lower CD34 HSCs dosing was an independent contributor to MDS-CAs and clinical t-MDS (Usmani et al., 2013). Although patients with MDS-CAs do not always develop t-MDS/AML, the phenomenon, also recognized in other tumors, requires understanding and development of preventive measures. We analyzed gene expression profiling (GEP) of bone marrow core biopsies at diagnosis from patients enrolled in our TT2 (n=88) and TT3 (n=263; training set) trials to identify genes associated with time to MDS-CA. Univariate Cox regression identified BCL11A as the only gene with expression associated with the time to development of MDS-CA (q 5.5 mg/L, GEP70-defined high risk and low BCL11A expression, while female sex was linked with longer time to MDS-CA development. In multivariate analysis, females retained independent prognostic significance for time to MDS-CA (HR-0.41, p 0.006), as did B2M 〉 5.5 mg/L (HR-1.95, p=0.038) and BCL11A expression