ALBERT

Description: Background Bortezomib-based combinations, including alkylating agents (VMP or CyBorD) or immunomodulatory drugs (VTD or RVD) have been established as regimens widely used in newly diagnosed MM patients. Bendamustine is a bifunctional alkylating agent effective in relapsed and/or refractory MM patients, and approved in Europe in combination with prednisone for elderly newly diagnosed MM. Since bendamustine may be more efficient than other alkylators, an attractive possibility would be to explore it in combination with bortezomib and prednisone (BVP) in newly diagnosed MM patients both transplant and non transplant candidates. Patients and Methods 60 newly diagnosed MM patients were included in the trial. The first cycle consisted on bendamustine at 90 mg/m2 given IV on days 1 and 4, in combination with bortezomib at 1,3 mg/m2 given IV on days 1, 4, 8, 11, 22, 25, 29 and 32 and prednisone at 60 mg/m2 given PO on days 1 to 4. In the following cycles, bendamustine was given on days 1 and 8, and bortezomib on days 1, 8, 15 and 22 (weekly schedule), and prednisone as it was previously described. Patients younger than 65 years proceeded to peripheral blood stem cell collection (PBSC) using growth factors alone after 4 cycles; HDT-ASCT was performed after 6 cycles. Patients older than 65 years received up to nine 28-day cycles. Results Between May 2011 and July 2012 enrollment was completed (60 pts). Median age was 61 years (range 38-82; 18 pts ≥65), 67% had ISS stage II/III, and 67% had unfavorable cytogenetics: t(4;14), t(14;16), del 17p or 1q gains by FISH. After a median of 6 cycles (2-9), 75% of patients achieved at least PR, including 16% of sCR, 9% CR and 28% of VGPR. Although the differences were not statistically significant, there was a trend to higher CR rate in the group of patients

Description: Background and Objectives: Recently, knowledge of the specific genetic markers responsible for cancer malignancies and their associated signaling pathways have generated many new targets that promise to increase drug efficacy while reducing side effects such as hematotoxicity. Although hematotoxicity is widely assumed to be the result of depleting hematopoietic progenitors, in particular myeloid progenitors, there are no assays to test new compounds in bone marrow samples to investigate this effect. Because these effects are patient-specific, an assay that identifies those patients risk of severe hematotoxicity by certain treatments could help personalizing patient treatment. Here, we show the ability of our flow cytometry-based automated Exvitech© platform to measure depletion analysis of different subsets of CD34+ progenitors and other cell subsets to potentially establish a new assay to screen drug candidates and combinations for hematotoxicity, as well as personalizing therapy to the individual sensitive patient at risk. So that in multiple myeloma (MM) we can identify at the same time CD34+ cells and pathological plasma cells; we could actually measure depletion of both malignant cells and progenitor cells on the same patient sample. Patients and methods: 16 normal bone marrow (NBM) and 4 MM samples were studied. For a proof of concept to test the hypothesis, we have selected two known cytotoxic drugs (cytarabine and clofarabine: N=10NBM) and two novel drugs with low expected cytotoxicity (ruxolotinib and volasertib: N=6NBM). The whole sample was plated into 96-well assay plates containing 8 concentrations of each drug and incubated for 48-hours for NBM and 12h with Bortezomib for MM samples. A multiple staining (CD45v450/Anexin-FITC/CD117-PE/CD34PerCP/CD38APC/CD19APCya7) was used to distinguish between both populations. Drug response was evaluated as a depletion survival index of each cell population relative to the average of 6 control wells in each plate. Results: As expected, nucleoside induces hematotoxicity in most of the studied NBM samples, but not all. Results reflect that cytarabine has similar activity than clofarabine in terms of efficacy (Ymax: 30% vs 23%) but with less potency (EC50: 6µM vs 0.2µM) in the immature population (N=10; Figure 1). This reflects a lower hematological toxicity which is consistent with clinical practice. Interestingly, for both drugs there is a large range of inter-patient variability inside this population in terms of efficacy (cytarabine, range Ymax: 4%-75% and clofarabine, range Ymax: 10%-37%) and potency (cytarabine, range EC50: 1µM-14µM and clofarabine, range EC50: 0.01µM-2µM) suggesting that in a subsets of vulnerable patients, drug doses could be tailored. By contrast, ruxolitinib and volasertib had little effect (Ymax: 80% vs 73%) in the immature population with minimal interpatient variability confirming the low toxicity expected for these novel drugs even at very high concentrations never achieve in vivo (Figure 1). Figure 2 shows bortezomib activity in MM bone marrow samples measuring both the dose response on malignant and myeloid progenitor cells. For Patient 1 the drug depletes myeloid precursor at lower doses than malignant cells, suggestive of severe hematotoxicity before therapeutic benefit can be achieved. Patient 2 shows the opposite case, where bortezomib depletes malignant cells completely without depleting myeloid precursors, suggestive of a good therapeutic index for this individual patient. Conclusions: These preliminary results show that Vivia Exvitech© platform is able to measure hematopoietic progenitors depletion in addition to other cell populations for novel drugs or before patientxs treatment that could contribute to a more selective drug development or a better clinical management of patients. This approach enables screening for hematotoxicty potential new discovery compounds, new drug candidates, and their synergistic combinations, thus supporting drug discovery and development. This assay could be helpful to both hematological and solid tumor drugs. The platform can measure both malignant and progenitor cells in bone marrow samples of MM and Non Hodgkin's Lymphoma. This simultaneous analysis shown for bortezomib could help guiding the clinical response and possible hematological toxicities associated to drug treatments. Figure 1: Figure 1:. Figure 2: Figure 2:. Disclosures Primo: Vivia Biotech: Employment. Ballesteros:Vivia Biotech: Employment. Robles:Vivia Biotech: Employment. Gorrochategui:Vivia Biotech: Employment. Garcia:Vivia Biotech: Employment. Hernandez:Vivia Biotech: Employment. Martinez:Vivia Biotech: Membership on an entity's Board of Directors or advisory committees.

Description: Invasive fungal infections (IFI) remain as an important cause of morbidity and mortality in adult patients with acute leukemia (AL). The incidence of proven or probable mold and yeast infections ranges from 5–24% among this selected high-risk patients. Prophylaxis is a commonly used treatment strategy, because the diagnosis of IFI is difficult and often delayed. A meta-analysis showed that itraconazole was more effective than fluconazole as prophylaxis of IFI, and a recent randomized trial demonstrated that in patients undergoing chemotehrapy for acute myelogenous leukemia (AML) posaconazole prevented IFI more effectively than did either fluconazole or itraconazole. Voriconazole (Vor) is a second-generation triazole with an extended spectrum of activity and is the drug of choice for invasive aspergillosis, the main mould that causes IFI among patients with acute leukemia. The drug is available for intravenous or oral administration and is well tolerated with transient visual disturbances, liver enzyme abnormalities and skin rashes being the most common adverse events reported, that rarely lead to treatment discontinuation. In this pilot study we evaluated Vor as prohylaxis treatment for IFI in 70 consecutive cases that underwent induction chemotherapy treatment for acute leukemia in two Clinical Units of Hematology in Spain. Methods:Oral prophylaxis with VOR at a fixed dose of 200mg/12h after two loading doses of 400mg was started on the first day of chemotherapy and continued until recovery from neutropenia or until ocurrence of an IFI. Patients who were unable to tolerate orally could receive VOR intravenously. Galactomanan was measured twice a week. HR-CT were performed in selected cases. The primary efficacy end point was the incidence of proven/probable IFI. Secondary end points included treatment success (vs failure) and the ocurrence of emergent mould infections. Treatment failure was defined as the ocurrence of proven/probable IFI; receipt of any other systemic antifungal agent for suspected IFI; discontinuation of the study drug for an adverse event; or withdrawal from the study with no additional follow-up. Results:From Jan 06 to Feb 07, 70 consecutive cases of AL that received induction chemotherapy for AML(58) or ALL(12) were included in the study. Median age was 57 (20–75). Median days with 0.1x109/L and 0.5x109/L were 16(10–35) and 19(12–38) respectively. Neutropenic fever ocurred in 62 cases (88,5%) and in 53% of them (33/62) a documented bacteria was isolated from the blood cultures. Galactomanan was possitive in only two cases (2,8%). HR-CT findings consisting with IFI were observed in 3 cases. Proven or probable IFI ocurred in 3 cases (A, flavus, A. fumigatus and Fusarium) (4,2%). Treatment failure as defined ocurred in 10 cases (16%). Conclusions: Prophylaxis with voriconazole was an effective and well tolerated regimen for IFI prevention in adult patients with acute leukemia.

Description: Background: Although great strides were made in the management of MM, our best chances to eradicate this malignancy may lie in preventing its progression.Most current models to predict risk of transformation in SMM are commonly established at diagnosis and not reevaluated over time, because some parameters such as tumor burden or genetic abnormalities require invasive bone marrow (BM) aspirates. It could be hypothesized that periodic monitoring of tumor biomarkers is needed to improve risk-stratification of SMM patients, and so would be new minimally-invasive methods that can replace those performed in BM samples. Such methods should also monitor immune profiles, to identify patients with stable tumor burden/genetics but at risk of progression due to lost immune surveillance. Aim: Determine the level of concordance between the tumor/immune landscape in BM vs peripheral blood (PB) of SMM patients, as well as to evaluate immune profiles together with circulating tumor cell (CTC) numbers and genetic alterations every 6 months in PB, as minimally-invasive methods for identification of SMM patients at risk of developing active MM. Methods: 300 patients are planned to be enrolled in the iMMunocell study that includes 24 sites across 8 European countries. PB samples are collected every 6 months during three years for next-generation flow (NGF) cytometry monitoring of CTCs and immune profiles. Additionally, CTCs and various immune cells are FACSorted to evaluate, every 6 months, their molecular profile in SMM patients with stable vs progressive disease. BM samples are taken at baseline and every 12 months according to patients' choice, in which the same methods described previously for PB are performed. An interim analysis was preplanned to the moment when 150 patients were enrolled. Results: A total of 170 SMM patients were enrolled and we report here data on the first 150. Thus far, 18/150 (12%) patients progressed to MM and according to 20/20/20 criteria, 1 had low, 7 intermediate and 10 had high risk SMM. Only 7/18 cases who progressed had 〉20% BM plasma cells (PC) by morphology. CTCs were detectable in 107/150 (71%) patients at baseline (median of 0.001% [0% - 0.42%] and 0.03 [0 - 21] CTCs/µL of PB). There was no correlation (or only modestly-significant) between the percentage of CTCs and BMPC by morphology (r=0.156, p=0.065) or flow cytometry (r=0.293, p=0.02). Median CTC counts were 0.02, 0.03 and 0.11 in SMM patients with low, intermediate and high risk disease according to 20/20/20 criteria, respectively (p=0.002). Median CTC numbers were significantly different between cases with stable vs progressive disease (0.02 vs 0.11, p=0.005). As compared to those with ≤1 CTC/µL of PB, patients with 〉1 CTC/uL showed significantly higher risk of transformation (8% vs 47%, p0.5 CTCs/uL. Data on the genetic landscape of CTCs analyzed every 6 months from baseline to disease progression will be shown at the meeting. Immune monitoring in patient-paired PB and BM samples at baseline (n=50) uncovered that 48 of 74 innate and adaptive immune cell types measured by multidimensional flow cytometry had similar distribution. Furthermore, we found significant differences in the distribution of three CD8 T cell subsets defined by differential expression of CD28, CD127, PD1, TIGIT, in PB of SMM patients with stable vs progressive disease. In patients with longitudinal PB samples from baseline until progression to active MM (n=7), there was a significant decrease in helper effector memory CXCR3+CCR4+ and cytotoxic CD127+TIGIT+PD1+ T cells, together with a significant increase in adaptive NK cells and Tγδ CD69+ T cells. Conclusions: This is the first study performing CTC and immune monitoring every 6 months in PB samples from patients with SMM. Our results show a significant correlation between CTC counts and stable vs progressive disease, and suggest that CTC kinetics could be complementary to the 20/20/20 criteria for real-time identification of individual SMM patients at risk of developing active MM. Beyond CTC numbers, this study is uncovering key immune cell types associated with disease progression. Disclosures Terpos: Amgen: Honoraria, Research Funding; Genesis: Honoraria, Other: travel expenses , Research Funding; Janssen: Honoraria, Other: travel expenses , Research Funding; Takeda: Honoraria, Other: travel expenses , Research Funding; Celgene: Honoraria; Medison: Honoraria. Raab:Amgen: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees, Research Funding; Heidelberg Pharma: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. Ocio:Sanofi: Consultancy, Honoraria; Secura-Bio: Consultancy; Oncopeptides: Consultancy; Celgene: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Speakers Bureau; Amgen: Consultancy, Honoraria; MDS: Honoraria; GSK: Consultancy; Takeda: Honoraria; Asofarma: Honoraria. Martinez-Lopez:Novartis: Consultancy; Janssen-cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Consultancy, Research Funding; Incyte: Consultancy, Research Funding; Janssen: Consultancy, Honoraria. de la Rubia:Amgen: Consultancy, Other: Expert Testimony; Celgene: Consultancy, Other: Expert Testimony; Janssen: Consultancy, Other: Expert Testimony; Ablynx/Sanofi: Consultancy, Other: Expert Testimony. Hajek:Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharma MAR: Consultancy, Honoraria; BMS: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Oncopeptides: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Ludwig:Celgene: Speakers Bureau; Janssen: Other: Advisory Boards, Speakers Bureau; Bristol Myers: Other: Advisory Boards, Speakers Bureau; Sanofi: Other: Advisory Boards, Speakers Bureau; Amgen: Other: Advisory Boards, Research Funding, Speakers Bureau; Takeda: Research Funding; Seattle Genetics: Other: Advisory Boards. Goldschmidt:Dietmar-Hopp-Foundation: Other: Grants and/or provision of Investigational Medicinal Product:; Chugai: Honoraria, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Incyte: Research Funding; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Molecular Partners: Research Funding; Johns Hopkins University: Other: Grants and/or provision of Investigational Medicinal Product; Mundipharma GmbH: Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Honoraria, Research Funding; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; University Hospital Heidelberg, Internal Medicine V and National Center for Tumor Diseases (NCT), Heidelberg, Germany: Current Employment; GlaxoSmithKline (GSK): Honoraria; Adaptive Biotechnology: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product, Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Merck Sharp and Dohme (MSD): Research Funding. Roccaro:European Hematology Association: Research Funding; AstraZeneca: Research Funding; Transcan2-ERANET: Research Funding; Italian Association for Cancer Research (AIRC): Research Funding; Janssen: Other; Celgene: Other; Amgen: Other. San-Miguel:Amgen, BMS, Celgene, Janssen, MSD, Novartis, Takeda, Sanofi, Roche, Abbvie, GlaxoSmithKline and Karyopharm: Consultancy, Membership on an entity's Board of Directors or advisory committees. Paiva:SkylineDx: Consultancy; Takeda: Consultancy, Honoraria, Research Funding; Roche: Research Funding; Adaptive: Honoraria; Amgen: Honoraria; Janssen: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; Kite: Consultancy; Sanofi: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau.

Description: Background Many proteins with tandem repeats in their sequence have been described and classified according to the length of the repeats: I) Repeats of short oligopeptides (from 2 to 20 amino acids), including structural cell wall proteins and arabinogalactan proteins. II) Repeats that range in length from 20 to 40 residues, including proteins with a well-established three-dimensional structure often involved in mediating protein-protein interactions. (III) Longer repeats in the order of 100 amino acids that constitute structurally and functionally independent units. Here we analyse ShooT specific (ST) proteins, a family of proteins with tandem repeats of unknown function that were first found in Leguminosae, and their possible similarities to other proteins with tandem repeats. Results ST protein sequences were only found in dicotyledonous plants, limited to several plant families, mainly the Fabaceae and the Asteraceae. ST mRNAs accumulate mainly in the roots and under biotic interactions. Most ST proteins have one or several Domain(s) of Unknown Function 2775 (DUF2775). All deduced ST proteins have a signal peptide, indicating that these proteins enter the secretory pathway, and the mature proteins have tandem repeat oligopeptides that share a hexapeptide (E/D)FEPRP followed by 4 partially conserved amino acids, which could determine a putative N-glycosylation signal, and a fully conserved tyrosine. In a phylogenetic tree, the sequences clade according to taxonomic group. A possible involvement in symbiosis and abiotic stress as well as in plant cell elongation is suggested, although different STs could play different roles in plant development. Conclusions We describe a new family of proteins called ST whose presence is limited to the plant kingdom, specifically to a few families of dicotyledonous plants. They present 20 to 40 amino acid tandem repeat sequences with different characteristics (signal peptide, DUF2775 domain, conservative repeat regions) from the described group of 20 to 40 amino acid tandem repeat proteins and also from known cell wall proteins with repeat sequences. Several putative roles in plant physiology can be inferred from the characteristics found.