ALBERT

Description: Introduction:Low-dose radiation therapy (LD-RT) is a therapeutic option in indolent non Hodgkin B-cell lymphomas (iNHL), usually in the palliative setting. If most iNHL are highly sensitive to radiation therapy, with good local control obtained with a dose of 4 Gy in 2 fractions, little is known about the efficacy and outcome of repetitive courses of LD-RT. We report here the results of a study cohort of repetitive LD-RT in iNHL. Methods : We retrospectively reviewed the records of all iNHL patients treated by two or more courses of LD-RT at Gustave Roussy, between January 1990 and December 2015. Patients received LD-RT as palliative treatment for low-bulky disease, patient's comfort or painful adenopathy. Clinical data, histological types, outcome and treatment lines were collected. Overall survival was the time between lymphoma diagnosis and death from any cause. Last LD-RT follow-up period was the time between the last LD-RT session and latest news. Results: Thirty-five pts were analyzed. Among them, 24 pts (69%) had Follicular Lymphoma (FL), 6 pts (17%) Marginal Zone Lymphoma (MZL), 3 pts (9%) had B-cell primitive Cutaneous Lymphoma Follicular Type (CL-FL) and 2 pts (6%) Nodular Lymphocyte Predominant Hodgkin Lymphoma (NLPHL). At lymphoma diagnosis, median age was 57 years [range 20-80]. Ann Arbor stage was I-II in 18 pts (51%), and III-IV in 17 pts (49%). Patients received a median of 4 therapeutics lines (range 2-11), and 2 LD-RT courses (range 2-6). Median overall survival was 146 months [29-298 months]. Four patients had died: 2 of disease progression and 2 others from concomitant illness (1 cardiac disease and 1 hepatocellular carcinoma). No patient had experienced transformation to diffuse large B cell lymphoma after RT-LD treatments. In the vast majority of cases (31/35; 89%), the LD-RT were successively performed to lymphoma relapse outside irradiation fields. Exclusive repetitive courses of LD-RT without chemotherapy were received by 8/35 (23%) of patients; while 24/35 (69%) patients received repetitive LD-RT alternately with immunotherapies or chemotherapies; and 3/35 (9%) others repetitive LD-RT alternately with standard dose RT. After the second course of LD-RT, 12/35 (34%) patients were managed in watch and wait approach, 6/35 (17%) received another LD-RT and 17/35 (49%) patients had experienced a progressive disease and were treated with immunotherapy or chemotherapy or standard dose radiotherapy. The LD-RT was the last treatment modality in 18/35 (51%) patients with histological types distributed in FL (n=10), MZL (n=5) and CT-FL (n=3). With a median last LD-RT follow up of 32 months [7-177 months], 23/35 (66%) patients remained in complete remission, 9/35 patients (26%) had experienced progressive disease and 3/35 (9%) patients had obtained stable disease. Conclusion: As palliative treatment modality, the repetitive low dose radiation therapy 4 Gy in two fractions could provide alternative option treatment in iNHL. This study support further investigations of this simple, well tolerated and not costly therapy in iNHL, especially in the context of new immunotherapeutic agent's area. Disclosures Michot: Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Ribrag:ArgenX: Research Funding; Esai: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Infinity: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Pharmamar: Membership on an entity's Board of Directors or advisory committees; NanoString: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees.

Description: Introduction: The BEAM (carmustine (BCNU), etoposide, aracytin and melphalan) standard conditioning regimen in autologous stem-cell transplantation is widely used since 1990 in patients with relapsed/refractory non Hodgkin's lymphoma (NHL) and Hodgkin's lymphoma (HL) who remain sensitive to salvage therapy. Recently, a phase Ib-II feasibility study using bendamustine rather than BCNU in the same indication was reported, but was not really compared to BEAM concerning safety. We report herein a safety analysis of Bendamustine-EAM (BeEAM) with a control BEAM counterpart paired cohort (2/1). Methods: We performed a case control retrospective study of patients with NHL and HL who underwent high-dose chemotherapy (HDC) with BeEAM regimen between January 2015 and December 2015. We matched each BeEAM patient with two patients having the same age, sex and number of treatment lines who underwent BEAM and ASCT between January 2008 and December 2014. No patient presented significant comorbidity. The BEAM regimen consisted in BCNU on day -6 (300mg/m2) cytarabine daily from day -6 to day -3 (200mg/m2 every 12 hours), etoposide daily from day -6 to day -3 (100mg/m2 every 12 hours) and melphalan on day -2 (140mg/m2). A similar scheme was adopted in BeEAM arm with bendamustine on day -6 and -5 (100mg/m2/d) replacing BCNU. Autologous stem cells were reinfused on day 0. Unfractionated heparin was used with Bendamustine to prevent veno-occlusive disease. Pegfilgrastim 6mg was injected subcutaneously on day 4. Febrile neutropenia was treated according to ESMO guidelines. Results: One hundred and two patients (68 BEAM and 34 BeEAM) were analyzed. Median age was 48 years in both arms. 61.8% of patients were male and 38.2% female. A median number of 4.4 x106 CD34 were reinfused in the two groups. The median time to neutrophils recovery (〉 0.5 x109) was similar between the two arms (9.06 vs 8.86 days, p=0.3). Grade 3 or greater diarrhea according to Commun Terminology Criteria for Adverse Events (CTCAE v4.03) classification was significantly more frequent in BeEAM patients (44 vs 13.2%, p=0.001). Median time to hospital discharge was significantly longer in BeEAM group (23 vs 20.8 days, p=0.0047). The median loss of weight during hospitalization was significantly greater in BeEAM patients (3.3 vs 1.9 kg, p=0.014). The median number of days with fever 〉38°C and with intravenous antibiotics was significantly higher in BeEAM group (6.06 vs 3.38, p

Description: Introduction: Therapy-related Myeloid Neoplasms (TRMN) arise after cytotoxic chemotherapy and/or radiotherapy administered for a prior neoplasm and have dismal outcome. Inherited predisposition or direct induction of fusion transcripts can be responsible of TRMN. Recent evidence suggests that patient with Clonal Hematopoiesis of Indeterminate Potential (CHIP) may have an increased risk of TRMN. In gynecological and breast cancers, CHIP mutations have been described in around 25 % of patients (Coombs et al., Cell Stem Cell 2017). In this setting, we aimed to identify the impact of CHIP-associated mutations in overall survival of TRMN. Methods: In this retrospective study, we included patients with TRMN diagnosed and/or treated at Gustave Roussy Cancer Center between January 2004 and December 2018 if they had a previous breast or gynecological cancer, DNA samples available at TRMN diagnosis and a signed informed consent. We performed a targeted 77 genes mutational analysis using Next Generation Sequencing (NGS), using Haloplex technique (Agilent), sequencing on MiSeq (Illumina). If any somatic mutation associated with hematological malignancies could define CHIP, the most frequent genes mutated in original CHIP papers are: ASXL1, ASXL2, ATM, BCOR, CBL, CHEK2, DNMT3A, IDH1, IDH2, JAK2, PPM1D, SF3B1, SRSF2, TET2 and TP53. These will define "CHIP-associated mutations" at TRMN diagnosis. According to results, patients were classified into "CHIP-associated mutations" or "no-CHIP" categories (no mutations detected or all the other mutations detected at TRMN diagnosis). Moreover, patients were also classified into 3 subgroups according to a modified genetic ontogeny-based classifier (Lindsley et al., Blood 2015): "P53/PPM1D" subgroup, "MDS and AML with MDS mutations" subgroup, and "de novo/pan-AML" subgroup. Survival analyses were performed using GraphPad software. Results: 77 patients were identified: 49 therapy-related AML (t-AML) (64%) and 28 therapy-related MDS (t-MDS) (36%). Median age at TRMN diagnosis was 62 years [36-86] and median time interval between primary cancer and TRMN was 5.1 years. Primary cancers were breast (70%), ovarian (23%), endometrial (4%) and cervical (3%) cancers. Patients were treated with radiotherapy alone (13%), cytotoxic agent alone (19%), or chemotherapy/radiotherapy (68%). The most frequently mutated genes at TRMN diagnosis were: TP53 (31%), DNMT3A (19%), NRAS (13%), TET2 (12%), NPM1 (10%), PPM1D (9%), PTPN11 (9%) (Fig1A). 10% of patients had no gene mutation detected. According to 2017 ELN risk stratification, genetic risk for t-AML was favorable, intermediate and adverse in 19 (39%), 14 (28%) and 16 patients (33%), respectively. According to IPSS score, 86% of the t-MDS patients were classified as High risk/Intermediate 2 and 14% as Intermediate 1/Low risk. Treatment options included best supportive care for 16 patients (21%), low dose chemotherapy for 26 patients (34%), or intensive chemotherapy/allogenic transplant for 34 patients (45%). Based on Lindsley's modified classifier median overall survival for "P53/PPM1D", "MDS" and "de novo/pan-AML" subgroups were 12, 17 and 25 months, respectively (p=0.009) (Fig1B). "CHIP-associated mutations" were detected in 53 patients (69%) with no significant impact on overall survival (Fig1C). Interestingly, age at TRMN diagnosis in patients with "CHIP-associated mutations" vs patients with "no-CHIP" was higher (65 vs 56 years old, p=0.002) and the time interval between cancer diagnosis and TRMN was longer (6.6 [0.9-38.1] vs 2.9 [1.2-8.5] years, p〈 0.001) (Fig1D). CHIP emergence was not correlated with type of cancer's treatment or with number of treatment lines. "P53/PPM1D" subgroup was more frequent in patients treated with 2 lines or more for their primary cancer than in patients who received only 1 line of treatment (50% and 25% respectively, p=0.03). Conclusion: TRMN occurring after gynecological or breast cancers are of bad prognosis, especially for P53 and PPM1D mutated patients. Our results show that CHIP related mutations are found in a large percentage of patients and could be responsible for emergence of TRMN, especially in older patients. Figure 1 Disclosures de Botton: Forma: Consultancy, Research Funding; Bayer: Consultancy; Daiichi: Consultancy; Novartis: Consultancy; Astellas: Consultancy; Abbvie: Consultancy; Pierre Fabre: Consultancy; Syros: Consultancy; Agios: Consultancy, Research Funding; Janssen: Consultancy; Pfizer: Consultancy; Servier: Consultancy; Celgene: Consultancy, Speakers Bureau. Micol:AbbVie: Consultancy; Jazz Pharmaceuticals: Consultancy.

Description: Adoptive transfer of allogeneic natural killer (NK) cells represents a promising treatment approach against acute myeloid leukaemia (AML). Success of this NK cell immunotherapy is dependent on obtaining high numbers of functional NK cells that have the potential to survive in vivo. The use of umbilical cord blood (UCB) CD34+ cells as a source of allogenic NK cells is an interesting method that can generate a readily available, non-invasive, off the shelf cellular product. We developed a cytokine-based culture method for the generation of NK cell products derived from CD34+hematopoietic progenitor cells (HPC) isolated from fresh UCB units. Immuno-phenotyping of ex vivo expanded NK cells showed typical inhibitory and activating NK receptors except for CD16 and the KIR receptors. UCB-derived NK cells displayed good cytolytic activity against NK-sensitive K562 cells with a percentage of specific lysis varying from 30 to 50%. Cytolysis was directly correlated to CD94 expression since CD94-sorted NK cells were responsible for all the in vitro cytolytic function of differentiated NKs against K562 cells. There was an inconstant susceptibility of patient-derived primary AML cells to UCB-derived NK lysis in vitro with a specific lysis ranging from 0 to 25%. We further characterized UCB-derived NK cells by investigating their toxicity, biodistribution, and anti-leukemic potential in vivo. As adoptive transfer of NK cells is an attractive approach for treating refractory leukemia, immune deficient mice were engrafted with a patient derived AML strain resistant to NK-mediated lysis and doxorubicin. After successful engraftment and randomization, leukemic mice were injected with either UCB- derived NK cells or NK cells from healthy donors (NKhds) or doxorubicin, with one control group that didn't receive any treatment. Mice were sacrificed after 2 weeks of treatment and leukemia load along with NK distribution were evaluated by flow cytometry in the blood, bone marrow (BM) and spleen. There was no evidence of toxicity of UCB-derived or healthy donors NK cells in mice. Both types of cells were preferentially found in the blood and in the spleen, even though NKhds reached much higher levels than UCB-derived NKs. As for treatment efficacy, none of our treatment showed anti-leukemic potential based on the absence of decrease of leukemic cells in BM, blood, and spleen. In vivo microenvironment didn't overcome resistance of the patient derived AML cell to NK lysis or to doxorubicin. Remarkably, all of the UCB derived NK cells founded in vivo expressed the CD94 whereas not more than 20% of the injected cells were positive for this marker. Whether it was by in vivo selection or by in vivo differentiation must be investigated. Interestingly, a small cell population with CD56 and CD34 double staining was distinguished in UCB-derived NK and NK healthy donor treated leukemic mice suggesting in vivo interaction between leukemic and NK cells. Further characterization of this population may help to understand the molecular mechanism of leukemic recognition by NK cells and resistance of leukemic cells to cytolysis. In conclusion, UCB-derived NK generation is feasible. Investigation of the role of CD94 in these cells is needed, as cell sorting by CD94 selection in addition to the CD56 could be an interesting approach in the future to select highly functional expanded NK cells before therapeutic use. Furthermore, infusion of UCB-derived NK cells into immune-deficient mice is achievable and non-toxic. However, in vivo environment didn't overcome primary in vitro resistance of AML cells despite an established interaction. Additional elucidation of AML resistance mechanisms to NK lysis is mandatory before therapeutic application. Disclosures No relevant conflicts of interest to declare.

Description: Background: The oncogenic BRAF V600E mutation activates the MAPK signaling pathway resulting in cell growth and survival of tumor cells carrying the mutation. Targeting BRAF has shown its efficacy in metastatic melanomas. BRAF V600E mutation has been described in multiple myeloma (MM), with an incidence of 4-10% in previous reports. We report here the incidence of BRAF mutations in a series of MM patients treated at our center. Methods: From February 2012 to March 2016, 94 MM patients were tested for BRAF mutations. Eighty-one samples came from patients with relapsed/refractory MM (R/R MM) and 13 samples from newly diagnosed patients. Eighty-eight samples were collected from bone marrow aspirates, 3 from extramedullary plasmacytoma biopsies, 1 from pleural effusion, and 2 from peripheral blood. Targeted sequencing for BRAF mutations using Sanger direct sequencing or targeted next generation sequencing were performed on all samples after CD138 cell (after 2015). In addition, NGS was performed for 2 patients. From 2012 to 2015, we used material obtained from scraping bone marrow smears with 〉 20% plasma cells. Since 2015, we began using immunomagnetic cell-sorting based on the CD138 cell surface marker (Stem Cells®). Results: BRAF mutations were detected in 8 samples out of the 84 that could have been screened (9.5%). When considering only the relapsed/refractory population, the incidence of BRAF mutations was 10.8%. One sample had the inactivating D594N BRAF mutation and the other 7 had the V600E BRAF mutation. The 8 mutated patients had relapsed/refractory MM at the time of analysis. Of note, 5 out of the 8 mutated patients (62.5%) had an IgA MM. Seventy-six samples had no BRAF mutation detectable. Ten samples were not tested because of the small percentage of MM cells on the bone marrow smears. The failure rate was 16% with the scraping technique and 3.4% with the sorting cell technique. Five out of 7 patients harboring the V600E BRAF mutation were treated with a BRAF inhibitor in different clinical trials. One mutated patient could not be treated due to exclusion criteria related to an underlying condition (terminal renal failure requiring chronic hemodialysis) and one patient received allogeneic stem-cell transplantation at relapse. No BRAF mutation was detected in the 13 newly diagnosed patients. Conclusion: Our results show that BRAF screening is feasible in routine practice and can help orienting therapy in relapsed/refractory MM patients. The incidence of 9.5% that we found in our series compares favorably with previously published results, and we observed a slightly increased incidence of 10.8% in the R/R population. The cell sorting technique seems to be more effective than the scraping technique for MM sample DNA acquisition and BRAF mutation screening. Results of BRAF inhibitors therapy in MM are eagerly awaited. Disclosures Lacroix: sanofi: Research Funding; qiagen: Membership on an entity's Board of Directors or advisory committees; roche: Membership on an entity's Board of Directors or advisory committees; astrazeneca: Membership on an entity's Board of Directors or advisory committees. Michot:Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Ribrag:ArgenX: Research Funding; Esai: Membership on an entity's Board of Directors or advisory committees; Infinity: Membership on an entity's Board of Directors or advisory committees; Pharmamar: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees; NanoString: Membership on an entity's Board of Directors or advisory committees.

Description: Therapy-related acute myeloid leukemia/myelodysplastic syndrome (t-AML/MDS) arise after cytotoxic chemotherapy and/or radiotherapy administered for a prior neoplasm and have a dismal outcome (median survival of 8 months in the largest series published including 306 patients (pts), Smith, Blood 2003). Recent registry data suggested a continued increase in survival in AML (Derolf, Blood 2009) and we wondered whether this was also observed in the setting of t-AML/ MDS. All pts with a t-AML/MDS diagnosed and/or treated for their prior neoplasm at Gustave Roussy Cancer Center between July 1986 and 2016 were included in this retrospective study. Data regarding pts' demographics, primary diagnosis and treatment, latency time, cytogenetic, treatment and outcome were collected. The diagnosis of t-AML/MDS was based on the WHO 2016 classification. t-AML were classified based on cytogenetic results as favorable, intermediate and adverse according to international classification, t-MDS based on IPSS score as favorable (Low, Int-1) and adverse (Int-2 and High). 428 pts were analyzed. The median age at diagnosis of t-AML/MDS was 56.4 years with a female predominance (60%). 224/428 (52.3%) pts had t-AML, 204/428 (47.7%) t-MDS. The most common primary malignancies were breast cancer (24%), non-Hodgkin lymphoma (15%), Hodgkin lymphoma (HL) (9%) and ovarian cancer (9%). Occurrence of t-AML/MDS following HL represented 26.6% of t-AML/MDS cases between 1986-96 comparing to 4 % between 2006-16 whereas breast cancer rose from 16% to 45%. Prior treatments included chemotherapy alone in 137/428 pts (32%), radiotherapy alone in 61 pts (14%) and both in 230 pts (54%). At diagnosis of t-AML/MDS, 295 pts (69%) were in complete remission (CR) of their prior neoplasm, 29 (7%) had a stable and 104 (24%) a progressive disease. Median interval between primary cancer and t-AML/MDS was 5 years (4.3 and 5.7 years for t-AML and t-MDS respectively, (p=0.03)). Furthermore, delay to develop t-AML/MDS after radiotherapy alone was longer compare to chemotherapy or both (6.1, 5.1 and 4.3 years, respectively (p=0.0087)). In the t-AML subgroup, 47% of pts presented unfavorable cytogenetic (including complex karyotype (20%) and 11q23 abnormalities (16.9%)), 26% intermediate and 26% favorable cytogenetic (core binding factor mutations (12.7%), t(15;17) (13.3%)). In the t-MDS subgroup, 78% of pts were considered adverse; complex karyotype, chromosome 7 and 5 abnormalities were found in 40.8%, 46.7% and 28.9% respectively. Pts received intensive chemotherapy (including 41 allografts), low dose chemotherapy (including 74 treatments with hypomethylating agents) and best supportive care in 42%, 24% and 34% respectively. The median overall survival (OS) was 10.6 months and the 5-year survival was 19.1% (Figure 1A). The 5-year OS of patients in CR of their prior neoplasm was 25.5% compared to 3.65 and 0% for pts with progressive and stable disease, respectively (p