ALBERT

Description: Abstract 3878 Poster Board III-814 Introduction Despite recent therapeutic improvements in the management of myeloma it remains an incurable disease. New therapies are therefore required. Pomalidomide (POM, Celgene) is a thalidomide derived immunomodulatory agent with similar preclinical spectrum of activity as lenalidomide. In Phase 1 studies we have previously demonstrated that POM monotherapy is well tolerated by patients with relapsed myeloma determining a maximum tolerated dose of 2mg od or 5mg on alternate days (Schey et al. JCO 2004; Streetly et al. BJHaem 2008). The toxicity profile was acceptable and overall response rates of 54 and 50% were observed with daily and alternate day dosing respectively. The current study examines long term responses, progression and survival outcomes. Patients were entered into the POM daily dosing or alternate day dosing Phase 1 studies between March 2001 and September 2003 and continued to receive treatment with POM until progressive disease (PD) or Grade 3 or greater non-haematological toxicity. Patients with PD were eligible to receive dexamethasone in addition to POM. POM became unavailable in May 2005 and patients still receiving drug were switched to receive lenalidomide. All patients gave informed consent. Results 44 patients received treatment with POM at a dose of 1mg alternate days – 10mg od. A median of 3 (range 1 – 8) prior lines of therapy had been received. POM was received for a median of 9.3 months (1 – 53). Following POM withdrawal 8/44 patients who had not developed PD subsequently received lenalidomide from a median of 30 months after starting POM. Overall responses by IMWG criteria to POM monotherapy were: CR 13.6%, VGPR 4.5%, PR 34%, MR 9%, SD 29.5% and PD 7% giving an overall response rate (〉PR) of 52%. Dexamethasone was introduced for PD for 10 patients and prolonged SD for 1 patient. 5/10 of these patients had 〉MR response to the addition of dexamethasone. With a median follow-up of 28 months the median PFS was 13.7 months and median OS was 28 months. Patients who had a PR response or better received POM for a median of 17.5 months and had improved PFS (median 19.8 months) and OS (median 42 months). 8/44 patients subsequently received lenalidomide. 7/8 of these patients have now developed PD at a median 26 months from commencing lenalidomide and 74 months from starting POM. Conclusions POM is a very well tolerated drug with excellent long term responses observed in this Phase 1/2 setting predominantly as monotherapy. Phase 2 studies are ongoing and results of these are awaited with interest. Disclosures: Streetly: Celgene: Honoraria. Schey:Celgene: Honoraria.

Description: Introduction Up to 50% of patients with acute GVHD (aGVHD) have a suboptimal response to first line therapy with corticosteroids. Extracorporeal photopheresis (ECP) is one of the recommended second line options. Overall response rates of 60-77% have been reported with ECP as second line therapy for aGVHD. Factors associated with lower response and survival were Grade 〉II at onset and number of organs involved. In a large retrospective study of ECP as second line therapy, 19.5% of patients developed chronic GVHD (cGVHD). Previous studies have included small numbers of patients with aGVHD post T cell depleted transplants or Donor Lymphocyte Infusion (DLI). Methods We conducted a retrospective analysis of outcomes in 40 patients treated with ECP for steroid refractory or steroid dependent aGVHD at 2 UK transplant centres. The aim was to analyse overall survival and long-term outcomes. All patients received 2-3 treatments per week for 8 weeks after which schedules differed according to centre, cessation or gradual taper. Results Between 2006 and 2014, 44 patients had received ECP as second line therapy, 4 were excluded from analysis as they had received less than 4 treatments. The median age was 51years (21-67years). Table 1 illustrates details of conditioning and GVHD prophylaxis. Thirty percent of patients were post DLI: 5 (12.5%) for disease relapse and 7 (17.5%) for mixed chimerism. Transplant conditioning included T cell depletion in 24 (60%): alemtuzumab n=17, ATG n=7. The median duration of steroids pre-starting ECP was 18 days (range 5-56). Using Modified Glucksberg crtieria at the onset of ECP, 5 patients (12.5%) had Grade II, 30 (75%) Grade III (including 6 with stage 4 gut) and 5 (12.5%) Grade IV aGVHD. Nine patients (22.5%) had 1 organ involvement and 31 (77.5%) had 2 or more organs involved. The dominant organ was gut in 21 (52.5%), liver in 10 (25%)and skin in 9 (22.5%). Maximal response rates were CR 57.5%, PR 27.5% and NR 15%. A number of patients received additional immunosuppressive therapy, predominantly anti-TNF antibodies: 14 patients (35%) (etanercept n=11, infliximab n=3). Using Kaplan-Meier analysis, censoring at last follow up, the overall survival from start of ECP at 5 years was 45.1% (95% CI 31.85-63.04%) Figure 1. The median survival was 2 years (27days-7.8yrs). In the DLI group, the 5 year overall survival was 59.8%. In a subgroup log rank Mantel-Cox analysis no significant difference was observed in the 5 year overall survival according to dominant organ involved: 53% skin, 40.6% gut and 50% liver (p=0.77) Figure 2. The cumulative incidence of death was 57.5% (23patients). The leading cause of death was infection 13/23 deaths (56.5%) followed by 5 deaths due to GVHD, 4 due to relapse, 1 from unrelated causes. The incidence of chronic GVHD was 55%. Treatment for cGVHD varied with 8 patients continuing ECP 〉12 weeks on a tapering schedule, 2 patients restarting ECP for cGVHD and the others receiving a variety of therapies including MMF, tacrolimus, Rituximab and imatinib. At last follow up, 17 patients were alive, 9 were off immunosuppression (2 of whom had not developed cGVHD) and 8 were still on immunosuppression. The performance status in the 17 surviving patients was ECOG 0= 8, 1=8 and 2=1. Discussion Our study shows an encouraging 5 year overall survival of 45.1% in a cohort with predominantly severe Grade III aGVHD. The 5 year survival was higher in the DLI group at 59.8%. Fifty three percent had stage 3-4 gut involvement, which is a recognised poor risk feature. Infection was the leading cause of death, which is of particular relevance in the T deplete setting. Whilst 55% developed cGVHD, over a long follow up period (median of 4years), the cGVHD resolved and performance status was preserved in most of the surviving patients. The incidence of cGVHD was higher than in other studies but the patients had higher grade of aGVHD at onset, were older and had multi-organ involvement. These findings need to be confirmed in prospective studies. Table 1. Pretransplant Characteristics No. of patients (%) Conditioning Regimes Myeloablative 10 (25%) Non-myeloablative 18 (45%) Total post DLI 12 (30%) T cell depletion 24 (60%) Alemtuzumab 17 (42.5%) ATG 7 (17.5%) GVHD prophylaxis CsA 20 (50%) CsA/MTX 9 (22.5%) CsA/MMF Tacro/MMF 8 (20%) 3 (7.5%) Donor Source Matched sibling Matched unrelated donor 10/10 9/10 8/10 Double Cord 9 (22.5%) 21 (52.5%) 3 (7.5%) 1 (2.5%) 6 (15%) Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Radia: Therakos: Other: ASH travel expense, Speakers Bureau. Off Label Use: Extracorporeal Photopheresis (ECP) is not licensed for use in GVHD but is recommended in international guidelines as a therapeutic option.. McLornan:Novartis: Research Funding, Speakers Bureau. Russell:Therakos: Other: shares.

Description: Abstract 3040 The Polyomavirus hominis 1 BK virus (BKV) is a non-encapsulated DNA virus, which infects up to 90% of the world's population, and may reactivate at times of severe immunosuppression, including post haematopoietic stem cell transplantation (HSCT). The significance of BK virus reactivation post Haematopoietic Stem Cell Transplant (HSCT) remains unclear. We collected retrospective data on viruria, viraemia, haemorrhagic cystitis (HC) and acute/chronic graft versus host disease (a/cGVHD) in patients at our institution over the period 2006 to 2011. We compared with a multivariate matched control group of 38, who did not reactivate BK. The groups were matched for age, sex, donor source and conditioning regimen including use of Alemtuzumab. Global BK reactivation incidence was 32% (38/118) of allogeneic HSCT during this period. 73% (28/38) of those who reactivated received volunteer unrelated donor (VUD) grafts, and 50% (18/38) received Alemtuzumab. Patients were sub-divided into those with high grade viraemia (HGV, VL 〉104 copies/ml), 47% (18/38) or low grade viraemia (LGV, VL

Description: Abstract 626 The physical contact between MM cells and the BM microenvironment can lead to cell adhesion mediated drug resistance. Although some components of adhesive structures in MM cells have been identified very little is known about their exact organisation and dynamics. We have observed formation of proteoglycan CD138 and F-actin containing membrane extensions in MM cells in in vitro cultures of BM aspirates as well as in sections of BM trephines of MM patients. MM membrane extensions elongated on the surface of BM stromal cells or interconnected MM cells forming seemingly cellular networks. Morphologically, MM membrane extensions appear similar to other structures in haematopoietic cells such as nanotubes formed by T-cells involved in cell communication and podia in CD34+ cells and leukemia cell lines whose function remains unknown. Dexamethasone (a drug commonly used clinically against MM) increased the percentage of MM cells displaying membrane extensions and the length of these structures. This correlated with enhanced expression of surface CXCR4, increased adhesion of MM cells on fibroblastic stromal cells and protection of MM cells against Dexamethasone-mediated apoptosis. Treatment with Bortezomib at clinically achievable doses decreased CXCR4 levels in MM cells and did not induce podia formation/extension and correlated with inhibition of MM cell adhesion and induction of MM cell apoptosis in a dose dependent manner. Blocking CXCR4 signalling with Plerixafor (AMD3100) inhibited the rate of formation and length of MM membrane extensions and correlated with sensitisation of MM cells to Dexamethasone. Similar inhibitory results were obtained using Dexamethasone in combination with the c-Abl/Src kinase inhibitor Dasatinib. Neither Dasatinib nor Plerixafor at doses achievable in patients directly induced apoptosis in MM or BM stromal cells. We conclude that membrane extensions/podia formed by MM cells are involved in the interaction with BM stromal cells, require CXCR4 and Src and/or c-Abl activity and could be involved in resistance to treatment with Dexamethasone. Disclosures: No relevant conflicts of interest to declare.

Description: Introduction The currently accepted standard of care for patients with relapse/refractory NHL is dose escalation and consolidation with Stem Cell Transplantation. The ability to predict those patients who may benefit or not from this approach would be beneficial. Whole-body Positron Emission Tomography using 18F-fluorodeoxyglucose (FDG) is now recognised to have predictive ability in those patients at high risk of relapse and shortened overall survival in De novo High grade lymphoma. In the salvage setting it is unknown whether the appearance of PET scan after 1 cycle of salvage treatment may predict similarly for outcome post transplant and whether early scanning is indicated before cycle 2 salvage chemotherapy. Aim: The aim of this study is to assess whether the PET-CT after one cycle or two cycles of dose escalation was predictive of outcome in patients with relapse/refractory NHL. Methods: We collected the data prospectively of all patients with relapsed or refractory aggressive NHL or HD treated in our institution over a 2 year period who were considered suitable for salvage therapy followed by Stem Cell Transplantation. We identified 14 male and 10 female patients with a mean age of 44 years (range 24–66). 12 patients had disease refractory to first line treatment and 12 patients had relapsed disease with a median interval from completion of treatment to relapse of 20 months (range 2–60). All patients were initially treated with DHAP as salvage therapy.22/24 patients proceeded to a stem cell transplant. 16/22 had an autologous transplant conditioned with BEAM and 6/22 had allogeneic transplants.FDG PET- CT scans were performed (all positive) prior to salvage chemotherapy and then assessed after 1 cycle of salvage chemotherapy and then after the second cycle using a 5 point visual scoring system as follows: CMR - no uptake at disease sites, MRU1 – visually uptake below level of mediastinum, MRU2 – visually between mediastinum and liver Stable/persistent metabolically active disease and Progressive disease (either new lesions or increased uptake in the same sites. Results FDG-PET Data was collected from 24 patients (14 male and 10 female) and read by two independent observers. The patients had the distribution shown in table: Visual response criteria Appearance PET 1 : Patient Numbers Appearance PET 2 : Patient Numbers CMR 1 6 MRU1 0 2 MRU2 1 3 Stable/persistent 16 5 Progressive 6 8 The median progression free survival of the patients with progression after one cycle of chemotherapy was 1.5 months with 3/6 deaths. Progression on PET1 was seen only in refractory cases pre salvage and was associated with a significant risk of relapse (p=0.014). The median progression free survival of the patients with stable disease after one cycle was 9.5 months. After two cycles of chemotherapy the median progression free survival of patients with CMR/MRU1/MRU2 was 9 months, compared with stable disease of 8 months and progressive disease 6 months. Conclusions: Patients with progressive disease on PET after one cycle of chemotherapy have a poor outcome with 50% early deaths and would be candidates for targeted/experimental therapies. Longer follow up will be required to assess the significance of stable or persistent disease on PET after one cycle and two cycles of salvage chemotherapy.

Description: Abstract 4584 Haploidentical transplantation is an option for patients who do not have a timely identifiable sibling or volunteer unrelated donor (VUD). Benefits of this stem cell source include donor availability, highly motivated donors and the ability to select the best donor from several relatives taking: age/fitness, cytomegalovirus (CMV) status, ABO group and natural killer cell alloreactivity into account. Historically the high level of human leukocyte antigen (HLA) disparity led to increased graft failure and high rates of acute and chronic graft versus host disease (GVHD). Luznik et al (Blood 2001) demonstrated that the use of post stem cell return cyclophosphamide in RIC haploidentical transplantation (using bone marrow as a stem cell source) reduced acute and chronic GVHD to acceptable levels, but at the expense of higher relapse rates in their cohort. We postulated that the use of PBSC's with their inherently higher T cell complement would reduce relapse rates compared to bone marrow, whilst post cell return cyclophosphamide would reduce acute and chronic GVHD. We present 5 patients treated at our centre using a RIC T cell replete haploidentical transplant protocol utilising PBSC's and post cell return cyclophosphamide. The patients, (median age 51; range 44–58), were treated for: relapsed follicular non Hodgkin's lymphoma (NHL), secondary acute myeloid leukaemia, Mycosis Fungoides and Adult T-Cell Leukaemia/Lymphoma (ATLL). Four patients had received 1st line chemotherapy only and remained chemotherapy sensitive, 3 of whom were in complete remission, one in a partial response. None had undergone a previous transplant. The NHL patient was chemotherapy insensitive following 4 previous lines of chemotherapy, a splenectomy and 2 rejected sibling allografts. Three patients were a major ABO mismatch, the remaining 2 fully matched. Four patients were CMV +/+ and 1 mismatched. HLA disparity ranged from 2–5 alleles (2 and 3 patients respectively). Median CD34+ cell dose returned was 6.98×106 cells/kg (range 4.81–8.00), with a median CD3+ cell dose of 2.36×108 cells/kg (range 1.19–2.97). The conditioning regime used was that of Luznik et al's (Blood 2001) phase I trial: Fludarabine 30mg/m2 day -6 to -2, cyclophosphamide 14.5mg/kg day -6 to -5, total body irradiation 2 Gray day -1, post stem cell cyclophosphamide 50mg/kg day +3 to +4, tacrolimus 1mg IV day +5 onwards, mycophenolate mofetil 15mg/kg TDS day +5 to +35. Outcomes: Four of 5 (80%) patients were fully donor chimeric by day 28 however graft failure with autologous reconstitution due to previously undetected HLA antibodies occurred in 1 patient. This patient reconstituted autologous neutrophils and platelets at 15 and 26 days respectively. Median time to neutrophil and platelet engraftment was 16.5 days (range 14–17) and 12 days (range 11–14) respectively. All 5 patients reactivated CMV (the latest at day 112). With pre-emptive treatment however none developed CMV disease. The incidence of acute GVHD grade II – IV and grade III - IV by day 100 was 40% and 20% respectively. Limited chronic GVHD was seen in 3 patients. 2 were assessed as grade I-II and 1 patient grade III. All cases of acute and chronic GVHD were steroid responsive. In both ATLL patients a sustained suppression of human T-lymphotropic virus (HTLV) viral loads was observed post transplant. One patient subsequently died of sepsis at day 113, the patient who had rejected their graft went on to relapse. The remaining 3 patients continue in CR, performance status 0, currently at day 245, 280 and 438. This data shows that RIC T cell replete haploidentical transplantation using PBSC's is well tolerated and enables both early engraftment and full donor chimerism. The rates of acute and chronic GVHD (40 and 60%) are comparable to sibling and fully matched unrelated donors. All of which has resulted in 60% of patients remaining in CR, including both ATLL patients who have gone on to fully suppress their HTLV viral loads. Disclosures: No relevant conflicts of interest to declare.

Description: Background Multiple Sclerosis (MS) is a chronic, immuno-mediated disease of Central Nervous System (CNS), mostly affecting young adults and frequently resulting in a progressive, irreversible disability despite the administration of approved Disease Modifying Treatments (DMTs). Autologous HSCT was shown to induce a high rate of sustained, treatment-free remissions in cases of aggressive MS, seldom associated to a partial reversal of disability. Toxicity of Conditioning Regimen is still a major concern. We retrospectively analyzed the outcome of 926 MS patients reported to the EBMT Registry who underwent autologous HSCT following the two most frequent CRs for this indication in the last 20 years. Patients and Methods Patient data were extracted from both the EBMT database and a disease-specific database developed by the EBMT Autoimmune Diseases Working Party (ADWP). Patients were selected for having received either BEAM + ATG (BEAM) or HD-Cyclophosphamide + ATG (CYC) as conditioning regimen. Hematological toxicity was assessed through Neutrophil (PMN) engraftment and 100-days (early) mortality (eTRM). MS forms at HSCT were reported as Relapsing-Remitting (RR), Secondary Progressive (SP), Primary Progressive (PP) and Progressive-Relapsing (PR). The impact of variables related to both patients (age, gender, year of HSCT, EDSS at HSCT) and disease characteristics (MS form, interval diagnosis-HSCT) at HSCT in the two groups were also evaluated. Results The utilization of conditioning regimens along the observed time period (1998-2018) was variable, with an increase of the HSCT activity in general after 2010 (230 vs 697 procedures) and a prevalence of BEAM before 2010 (205 BEAM vs 25 CYC) and of CYC thereafter (205 BEAM vs 492 CYC, p=0.001). Also, RR forms of MS prevailed over Progressive forms after 2010 (p=0.001) which is reflected in the different distribution across the two regimens, with RR significantly more frequently treated with CYC-based regimen (p

Description: Abstract 1809 Direct contact of multiple myeloma (MM) cells with the bone marrow (BM) stroma promotes cell survival leading to cell adhesion mediated drug resistance (CAM-DR). Dexamethasone is a conventional anti-MM drug that effectively induces MM cell death at presentation and in relapsed patients but the signalling pathways involved in its mechanisms of action are not completely understood. Resistance of MM to Dexamethasone may result from genetic changes in MM cells or through the contact of MM cells with the BM microenvironment. It has been shown that CAM-DR blocks the effect of Dexamethasone by the binding of MM cells to the BM stroma. Our data indicate a key role of the binding of MM CXCR4 receptor on stromal cell derived SDF1-a in CAM-DR against Dexamethasone. Dexamethasone induced upregulation of CXCR4 in the MM1.S cell line (sensitive to Dexamethasone) whilst, as expected, having no effect on the MM1.R cell line (resistant to Dexamethasone by expression of a mutated form of the glucocorticoid receptor). Bortezomib induced down regulation of CXCR4 in both MM1.S and MM1.R cell lines in a dose dependent manner that correlated with decreased adhesion on BM stromal cells and increased sensitisation of MM cells in the presence of the BM stroma. The Wiskott Aldrich Syndrome Protein (WASP) is an adaptor protein that regulates actin polymerization and organization of cell adhesion molecules of the integrin family in haematopoietic cells. WASP is involved in the signalling pathway downstream of CXCR4 in various leukocytes and we hypothesised that blocking this pathway would prevent MM cell adhesion on stromal cells and sensitise them to treatment with Dexamethasone. We found that downregulation of WASP in the Dexamethasone-sensitive cell line MM1.S using shRNA reduced the adhesion to the BM stroma. However, it also rendered MM1.S cells resistant to treatment with Dexamethasone but not with Bortezomib, Doxorubicin or Melphalan independently of the presence of BM stromal cells. Similarly, downregulation of WASP in MM1.R cells did not affect their sensitivity to Bortezomib, Doxorubicin or Melphalan. Dexamethasone has been shown to induce changes in actin dynamics or in levels or activity of actin and/or adhesion related proteins. Western blot analysis showed both Dexamethasone and Bortezomib to cause modulation of WASP expression, inducing partial downregulation and upregulation, respectively. Downregulation of WASP expression in MM1.S cells using shRNA or treatment with Dexamethasone or Bortezomib did not affect the expression of other proteins involved in WASP-mediated F-actin remodelling or cell adhesion such as WIP, Arp 2/3 or vinculin. We conclude that WASP is involved in the signalling pathways that promote cell-adhesion of MM cells on BM stroma but it is also a vital component of the signalling pathway of Dexamethasone's mechanism of action. These data indicate that while blocking WASP signalling could theoretically have potential therapeutic benefits to prevent CAM-DR to Dexamethasone, it would inhibit the action of Dexamethasone rendering cells resistant to treatment with this drug. Our study suggest a possible paradox of a dual pro-apoptotic and pro-survival effect of certain therapies targeting pathways involved in the interaction of MM cells with the microenvironment. Detailed studies of the intricate connexion between the intracellular pathways involved in the process of adhesion and drug induced cell death through modulation of actin dynamics may lead to improved therapeutic strategies to overcome drug resistance against Dexamethasone and perhaps other drugs. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 4532 Myeloproliferative Neoplasm (MPN)-Leukaemic Transformation (LT) uniformly carries a dismal prognosis. Effective therapy for such patients are currently lacking with no established evidence base to guide Allogeneic Haematopoietic Stem Cell Transplant (AHSCT) regimen. We report the outcome of a cohort of patients undergoing AHSCT at our 2 institutions over a 6-year period (2006–2012). 24 patients underwent AHSCT following diagnosis of MPN transformed to an accelerated phase (5–9% blasts on bone marrow (BM), n=9 & 10–19% BM blasts, n=2) or blastic phase (〉20% blasts on peripheral blood or BM, n=13). Disease subtypes were: Polcythaemia Vera (PV, n=4), Essential Thrombocythaemia (ET, n=6), Primary Myelofibrosis (PMF, n=8), Myelodysplastic/Myeloproliferative neoplasm-Unclassified (MDS/MPN-U, n=6). Median age at diagnosis was 50 years (range 29–67) and median time to transformation was 50 months (range 0–271). Cytogenetics were abnormal at transformation in 11 patients (46%), with 6 (25%) demonstrating abnormalities of chromosomes 5, 7 or complex karyotype, and 5 displaying trisomy 8, whilst 1 had isolated chromosome 17p deletion. 13 patients harboured the JAK2V617F mutation. Patients received a median of 3 (range, 1–5) lines of therapy for chronic and acute phase prior to AHSCT, of which 20 patients received intensive AML-type induction therapy. Disease status at time of AHSCT was complete remission (CR) in 13 cases, partial response (PR) in 7, and 4 patients had persisting AML. Conditioning regimes were Reduced Intensity with T-depletion (Alemtuzumab or Anti-Thymocyte globulin) in 23/24 cases (Fludarabine/Busulfan-based n=12, FLAMSA (Fludarabine, Ara-C and Amsacrine, followed by TBI/Cyclophosphamide or Busulfan) n=9, Fludarabine Cyclophosphamide TBI haploidentical protocol n=1, Fludarabine/Melphalan n=1). Median CD34 dose was 6.48 × 10∧6 cells/kg (range 1.17 {BMH} −10.71). Stem cell source was Peripheral Blood in all but one case, from unrelated (n=17) or related (n=7) donors. Median time to both neutrophil and platelet engraftment was 13 days (range 9–25 and 7–68 respectively); 2 patients including the haploidentical transplant had Primary Graft Failure (8%). The incidence of severe (grade 3&4) acute GVHD was 3/24 (12.5%) and 10 patients developed NIH-defined chronic GVHD (8 moderate, 1 severe). Day 100 Non-relapse mortality was 12.5%. Patients underwent sequential chimerism monitoring. Median OS for the entire cohort was only 10 months with a median progression free survival (PFS) of only 6.5 months. 5 patients received therapeutic Donor Lymphocyte Infusion (tDLI) for relapse at a median dose of 1×10∧6 CD3+/kg. 2 patients received DLI alone for chronic phase relapse, of whom 1 achieved remission. 3 patients received chemotherapy + DLI and 1 achieved 2ndCR. At last follow-up, 11/24 patients were alive with median surviving patient follow-up of 25 months. The percentage of BM blasts at progression from chronic phase had a highly significant impact upon outcome post AHSCT, median OS 23 vs. 10 months for 5–9% BM blasts compared to 310% BM blasts (p=0.011) & PFS 11 vs 6 months respectively (p=0.033, Fig 1). This effect was replicated when considering disease response immediately prior to AHSCT, with a median OS of 28 months for those in CR, compared to 10 months for those with excess blasts (p=0.017) and median PFS 11 vs 6 months, p=0.019 (Fig 2). Disease duration, subtype, Jak2 status and age at allograft did not significantly affect survival. Of note for the 3 surviving patients with follow-up over 6 months, all received FLAMSA-RIC conditioning (n=9). 5 patients who received FLAMSA TBI; 2 died of treatment-related complications, and 2 with residual disease at time of AHSCT relapsed early. Of 4 patients who have received a hybrid FLAMSA-Busulfan regimen, 2 remain alive in CR and 2 achieved a relapse free period of 12 months. Interestingly, PFS for FLAMSA-Bu patients appears significantly improved compared to conventional RIC regimens (median PFS 12 vs. 6 months, p=0.035) on univariate analysis, although conclusions are limited by cohort size. Further work into optimising transplantation regimens for accelerated and blastic phase MPN is warranted. Early use of FLAMSA-Busulfan hybrid protocol, before transformation to overt blastic phase, in conjunction with early weaning of immunosuppressive therapy and prophylactic DLI may improve the proportion of long-term survivors. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures: Harrison: Sanofi Aventis: Honoraria; Shire: Honoraria, Research Funding; YM Bioscience: Consultancy, Honoraria; Novartis: Honoraria, Research Funding, Speakers Bureau.

Description: Abstract 3979 Adhesion of MM cells to the BM microenvironment can lead to cell adhesion mediated drug resistance (CAM-DR). We have previously shown that treatment with Dexamethasone (a drug commonly used against MM alone or in combination with new therapeutic agents) induces in MM cells increased assembly of CD138, and F-actin containing membrane extensions named podia. Using this 2D co-culture model we found that podia from MM cells from patients or MM cell lines elongate on the surface of BM stromal cells and also interconnect distant MM cells correlating with CAM-DR. Formation of podia was observed after at least 6 days of co-culture when many soluble factors and extracellular matrix proteins may have been secreted and remodelled. We also showed that preventing SDF1-alpha binding to the CXCR4 receptor in MM cells inhibits podia formation. In order to investigate the possible role of ECM proteins and their ligands in podia assembly we have now tested the formation of podia in 3D co-cultures of MM and stromal cells embedded in gels composed of polymerised extracellular matrix proteins found in the BM including collagen I and collagen IV. We now found that MM cells incubated overnight in 3D matrix lattices assembled podia that elongated along fibres of polymerised extracellular matrix that put them in contact with stromal cells or other distant MM cells. We also found that culture of MM cells in 3D matrices in the absence of stromal cells did not result in podia formation. These results suggest that stromal cells promote podia formation in contact with the extracellular matrix. It has been shown by Neri et al1that beta7 integrins are involved in MM cell adhesion to BM cells and extracellular matrix during recruitment to the BM and they are involved in CAM-DR. We found that integrin beta7 KD MM cells failed to form podia to the same extent as the parental cell lines in 2D and 3D cultures. Podia assembled by beta7 KD cells were shorter and highly irregular in shape in comparison to long and straight podia formed by parental MM cells. Using immunostaining, we found that podia in beta7 KD cells lacked internal F-actin as well as vimentin filaments that we identified in podia of parental MM cells. However, total levels of these cytoskeletal proteins remained unchanged as detected by western blot. Dexamethasone-induced increased formation of podia was also dependent on the expression of beta7 integrins by MM cells and the presence of BM stromal cells in 2D and 3D culture conditions. We conclude that podia promote the formation of cellular networks of distant MM cells as well as the contact with BM stromal cells in 3D and this process is involved in CAM-DR against Dexamethasone. Podia assemble in contact with stromal cells and along extracellular matrix cables and require polymerisation of actin and vimentin cytoskeletal filaments downstream of beta7 integrins. Disclosures: No relevant conflicts of interest to declare.