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BACKGROUND The successful initiation of enteral nutrition is frequently hampered by various complications occurring in patients treated in the intensive care unit (ICU). Successful placement of a nasojejunal tube by CORTRAK enteral access system (CEAS) has been reported to be a simple bedside tool for placing the postpyloric (PP) feeding tube. METHODS We evaluated the efficacy and side effects using CEAS to establish EN in patients with critical illness, thrombocytopenia, and/or anticoagulation. RESULTS Fifty-six mechanically ventilated patients were analyzed. Twenty-four of them underwent prior hematopoietic stem cell transplantation (SCT). Sixteen patients received extracorporeal membrane oxygenation treatment because of acute respiratory distress syndrome. The median platelet count at PP placement was 26 g/L (range, 4-106 g/L); 16 patients received therapeutic anticoagulation (activated partial thromboplastin time, 50-70 s). CEAS-assisted placement of a PP nasojejunal tube was performed successfully in all patients. The most frequent adverse event was epistaxis in 27 patients (48.2%), which was mostly mild (Common Terminology Criteria for Adverse Events grade 1, n = 21 [77.8%], and grade 2, n = 6). A significant association between a low platelet count and bleeding complications was observed (P

Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) using T-cell-replete grafts and post-transplantation cyclophosphamide (PTCY) provides a popular curative approach for older patients (pts) with high-risk (HR) MDS/AML. The sequential therapeutic concept herein, is used to optimize disease control and gain time, especially in patients suffering from active disease at time of haplo-HSCT. Yet, sequential conditioning regimens prior to haplo-HSCT are often associated with a considerable risk of severe adverse events especially in older comorbid patients. Previous studies in the HLA-matched setting have demonstrated feasibility and safety of treosulfan-based reduced intensity conditioning (RIC) by stable engraftment and low non-relapse mortality (NRM). However, data for treosulfan-based conditioning in the unmanipulated HLA-haplo-HSCT setting in HR AML/MDS pts are rare, especially in the context of sequential conditioning. Here we report on a matched-pair analysis of 26 patients treated with either a treosulfan- or melphalan-based sequential conditioning for haplo-HSCT using PTCY as GvHD prophylaxis in HR MDS/AML. We retrospectively analyzed the outcome and toxicity profile of 26 patients undergoing sequential haplo-HSCT at our center between January 2009 and June 2019. Thirteen patients with HR AML/MDS and >54 years old who underwent sequential haplo-HSCT using treosulfan (3x10g/m2) for RIC were considered for potential matching with recipients (n=30) of a sequential melphalan-based RIC regimen. Matching criteria comprised (1) disease activity (blast yes or no), (2) disease status (relapse, refractory, high-risk cytogenetics), (3) HCT-CI and (4) age (+/- 5 years). Post-grafting immunosuppression consisted of cyclophosphamide, tacrolimus and MMF. Thirteen patients undergoing treosulfan-haplo-HSCT were successfully pair-matched with thirteen recipients of melphalan-based haplo-HSCT, respectively ((1) p=1.0; (2) p=1.0; (3) p=1.0; (4) p=0,76). Median age of the entire cohort was 63 years (54-71). Each group consisted of two MDS patients and eleven AML patients. All recipients treated with treosulfan showed neutrophil engraftment with a median of 20 days while only 69% of the melphalan treated patients engrafted (median=19 days, p= 0.9). In the melphalan group one graft rejection occurred, three patients died in early aplasia. Acute GvHD °II-IV occurred in 23% and 44% of the patients treated with treosulfan or melphalan, respectively. Severe (°III-IV) non-hematologic regimen-related toxicities were seen in 2/13 pts of the treosulfan and 7/13 pts of the melphalan group, predominately affecting the GI-tract in both. NRM at day +100 was 0% and 31% (p=0.06) for the treosulfan and melphalan group, respectively. Thereby, infections made up for most NRM events. Conversely, CI of relapse at one year was 23% vs 0 % (p=0.004) for treosulfan vs melphalan. OS at 1-year (treosulfan group: 69% vs melphalan gropup: 62%) and PFS at 1-year (treosulfan group: 69% vs melphalan group: 62%) did not differ significantly between the groups (p=0.72) Sequential haplo-HSCT using treosulfan and PTCY in older advanced MDS/AML patients is safe, resulting in lower NRM at the expense of higher relapse incidence compared to the melphalan-based sequential conditioning approach. Treosulfan-based RIC in haplo-HSCT though might be an alternative in older pts with low leukemic burden. However for disease control its intensity should be reconsidered and all available post-grafting mantainance strategies applied. Disclosures Fraccaroli: Medac: Other: Travel Grant. Buecklein:Celgene: Research Funding; Amgen: Consultancy; Pfizer: Consultancy; Gilead: Consultancy, Research Funding; Novartis: Research Funding.

Transplant-associated microangiopathy (TAM) is a life-threatening complication after allogeneic transplantation, related to endothelial toxicity caused by chemoradiotherapy, infections, immunosuppressive drugs and GvHD. Clinical manifestations include severe thrombocytopenia, microangiopathic hemolysis and organ dysfunction (e.g. renal, neurological) in the absence of disseminated intravascular coagulation. The reported incidence of TAM after allogeneic hematopoietic stem cell transplantation (HSCT) varies between 0,5 to 70%. Data for TAM occurrence and outcome in the context of HLA-haploidentical transplantation (haplo-HSCT) are rare. Here, we report our experience on TAM incidence, disease manifestation, treatment and outcome in T-cell-replete haplo-HSCT using PTCY as GvHD prophylaxis. We retrospectively analyzed the treatment course of 148 adult patients undergoing haplo-HSCT with regard to the incidence of TAM at our center from January 2012 to April 2019. The diagnosis of TAM required the fulfillment of the criteria as defined by Ruutu et al., Haematologica 2007. Median age of the entire cohort was 54 years (23-74). The majority of patients was transplanted for myeloid disease (60%). Postgrafting immunosuppression consisted of PTCY, CNI and MMF in all patients. Median follow-up time upon survivors was 3 years (4 months - 7 years). Twenty of 148 patients developed TAM, with a cumulative incidence of 10% at day 100. Median interval from haplo-HSCT to TAM onset was 63 days (23-295 days), with seven patients showing TAM after day +100. Median platelet count at diagnosis was 20 x 109/L (range: 6-116), predominantly de novo or worsening thrombocytopenia. Median LDH level was 437 IU/L (305-1445 IU/L). Serum haptoglobin levels were decreased in 85% of the TAM patients. 12/20 patients had active infection at the time of TAM diagnosis, most commonly viral (10/12). All but three patients presented with concurrent active acute GvHD requiring systemic steroid treatment in 16 patients Renal function abnormalities were diagnosed in 55% of the patients affected by TAM, with 4 patients requiring hemodialysis. Nine patients developed grade III hypertension and three patients showed neurologic dysfunction. Upon diagnosis of TAM, CNI dose was reduced (7 patients) or temporarily discontinued (7 patients). No switch to another CNI was performed. FFPs were transfused in nine patients, three patients underwent plasma exchange therapy without any effect. In four patients rituximab was applied. With a CI of 18% vs 2% (p0.002) using a PBSC graft was a risk factor for TAM development when compared to BM. Neither origin of disease (myeloid/lymphoid) nor sex of patient/donor or conditioning intensity (RIC/MAC) influenced the incidence significantly. Estimated one-year and three-year overall survival did not differ between the TAM and the non-TAM cohort (61% vs 61% and 46% vs 47%, p=0.9). Only one patient died because of TAM. With an incidence of 10% at day +100 at our center TAM is a serious but not rare complication after haplo-HSCT using PTCY, CNI and MMF for GvHD prophylaxis. However, in our cohort disease course was moderate with low TAM-associated mortality. The use of a PBSC graft was the only factor influencing TAM incidence. Disclosures Hiddemann: F. Hoffmann-La Roche: Research Funding.

Sequential conditioning regimens, comprising cytoreductive chemotherapy shortly applied prior to reduced intensity conditioning are successfully used for high-risk (HR) AML/MDS in matched related and unrelated donor hematopoietic stem cell transplantation. However, few data are available for sequential conditioning in the context of HLA-haploidentical transplantation (haplo-HSCT), especially in the elderly. To investigate the relative merits of sequential haplo-HSCT in the elderly we retrospectively analyzed the outcome of thirty-five patients (pts) with advanced AML/MDS (>=50 years old). Thirty-three pts suffering from HR AML, defined by refractory, relapsed or secondary leukemia, or complete remission with adverse-risk genetics according to ELN criteria and two pts with HR MDS according to IPSS-R, who underwent T-cell-replete haplo-HSCT at our institution between January 2009 and November 2016 were included. Disease was active in 29 pts while 6 had achieved CR. Pre-transplantation risk factors were scored using the hematopoietic cell transplantation-specific comorbidity index (HCT-CI) which was ≥3 in 13 pts (median HCT-CI:2, range 0-8). A sequential therapeutic concept using either FLAMSA (n=26) or clofarabine (n=9) for cytoreduction was used prior to RIC in all pts. Bone marrow (54%) and peripheral blood stem cells (46%) were both used as graft source. Post-grafting immunosuppression consisted of high-dose cyclophosphamide, tacrolimus and MMF in all pts. Median age was 60 years (50-70). One graft rejection occurred. Three pts died early in aplasia. Neutrophil and platelet engraftment was achieved in 95% and 77% of evaluable pts, respectively at a median of 16,5 (13-89) and 31,5 (11-103) days.Acute GvHD grade I-III occurred in 25/32 of the pts (grade III n=2); no patient developed grade IV aGvHD. Chronic GvHD was observed in 13/29 pts and was most frequently assessed as mild (n=6) or moderate (n=5) while 2 pts developed severe cGvHD. No GvHD related death was observed. CI of NRM at day 100, 1-year and 3-years was 11%, 23% and 23%, respectively. CI of relapse at 1- and 3-years was 15% and 27%, with a median time to relapse of 152 days (20-413). At a median follow up of 27 months (4-74), estimated one- and three-year overall survival (OS) was 62% and 52% respectively. One- and three-year leukemia-free survival (LFS) was 59% and 52%. Our results suggest that using a sequential therapeutic concept in PTCY-based haplo-HSCT is safe and properly tolerated while it provides a favorable disease control when treating elderly HR MDS/AML pts. Thus, sequential haplo-HSCT seems to be a valuable alternative in pts who lack a conventional donor or are in urgent need for prompt transplantation. Disclosures Tischer: Jazz Pharmaceuticals: Other: Jazz Advisory Board.

Fms-like tyrosine kinase 3 (FLT3) is a receptor tyrosine kinase constitutively expressed by acute myeloid leukaemia (AML) blasts. In addition, 25% of AML patients harbour a FLT3-ITD mutation, associated with inferior outcome due to increased relapse rate. Relapse might be propagated by interactions between AML blasts and the bone marrow microenvironment. Besides cellular elements of the microenvironment (e.g. mesenchymal stromal cells), bone marrow hypoxia has emerged as an additional crucial component. Hence, effects of hypoxia on FLT3 expression and biology could provide novel insight into AML biology. Here we show that 25% of AML patients down-regulate FLT3 expression on blasts in response to in vitro hypoxia (1% O2), which was independent of its mutational state. While virtually no AML cell lines regulate FLT3 in response to hypoxia, the down-regulation could be observed in Ba/F3 cells stably transfected with different FLT3 mutants. Hypoxia-mediated down-regulation was specific for FLT3, reversible and proteasome-dependent; with FLT3 half-life being significantly shorter at hypoxia. Also, PI-3K inhibition could partially abrogate down-regulation of FLT3. Hypoxia-mediated down-regulation of FLT3 conferred resistance against cytarabine in vitro. In conclusion, FLT3 expression in AML is dependent on the oxygen partial pressure, but response to hypoxia differs.

Reduced oxygen partial pressure (pO2, hypoxia) is an important component of the bone marrow microenvironment and the hematopoietic stem cell niche. It is unclear whether this applies to the leukemic stem cell as well and if differences in pO2 between the normal hematopoetic and the leukemic stem cell niche exits. Here, we demonstrate that while there is no detectable difference in the hypoxic level of bone marrow infiltrated by acute myeloid leukemia (AML) and healthy bone marrow, physiological hypoxia of 1% O2 itself leads to cell cycle arrest of AML blasts (both cell lines and primary AML samples) in the G0/G1 phase with upregulation of p27 and consecutive decrease of cells in the S phase. Hence, susceptibility of AML blasts toward cytarabine as S phase dependent drug is significantly decreased as shown by decreased cytotoxicity in vitro. In addition, cells exposed to hypoxia activate PI3K/Akt and increase expression of anti-apoptotic XIAP. Inhibition of PI3K can restore cytarabine sensitivity of AML blasts at hypoxic conditions. In conclusion, hypoxia mediated effects encountered in the bone marrow might contribute to chemoresistance of AML blasts.

Immune therapy for acute myeloid leukaemia (AML) has been largely disappointing. One possible explanation might lie in the microenvironment of the bone marrow, comprising cellular (e.g. mesenchymal stromal cells, MSC) and non-cellular components (e.g. hypoxia). The purpose of this study was to investigate the effects of these components in the immune response against AML in vitro. In vitro exposure of lymphocytes to hypoxia resulted in an increased expression of CD69 as an activation marker in NK cells only, with subsequently enhanced cell lysis of K-562 cell line by NK cells but not in lysis of primary blast. However, co-culture of AML cells with MSC significantly protected leukemic blasts from NK cell mediated lysis, mainly in a specific manner requiring cell-to-cell contact with supportive MSC. These data imply a relevant but unequivocal role of hypoxia and MSC the immune response against AML blasts.

Background & Aims Hydrophobic bile salts such as glycochenodeoxycholate (GCDC) accumulate in cholestatic liver disease and induce hepatocellular apoptosis, promoting profibrotic signalling. The tissue microenvironment is an integral player in cellular pathophysiology, but it is not routinely incorporated into laboratory studies. Tissue oxygen partial pressure (pO2) may be an underestimated component of the microenvironment: in the liver, a pO2 of 30–45 mmHg (approximately 6% O2) is physiological, because of predominant portal blood supply. It was the aim of this project to investigate the impact of physiological hypoxia (i.e. 6% O2) on hepatocellular function, namely, bile salt-induced apoptosis. Methods Human hepatoma cells (HepG2-Ntcp) and primary rat hepatocytes were cultured at standard laboratory (hyperoxic) conditions (21% O2) and at physiological hypoxia (6% O2) in parallel for 1–8 days to study hepatocellular apoptosis and activation of signalling pathways. Standard laboratory analyses were applied for bile salt uptake, caspase-3/-7 activity, western blotting and gene-array analysis. Results Culturing at physiological hypoxia protected both human and rat hepatocytes against GCDC-induced apoptosis: caspase-3/-7 activation was diminished by 3.1 ± 0.5-fold in human HepG2-Ntcp and completely abolished in primary rat hepatocytes. Bile salt uptake was unaffected. Induction of hypoxia-inducible factor-1α indicated adaption to physiological hypoxia. The MEK/ERK cascade was activated and anti-apoptotic mediators were induced: N-Myc down-regulated gene, gelsolin and carbonic anhydrase IX were upregulated 12.4-, 6.5- and 5.2-fold respectively. Conclusions We conclude from these data that (i) physiological hypoxia protects hepatocytes from bile salt-induced apoptosis, (ii) tissue pO2 is a crucial, underestimated component of the microenvironment and should (iii) be considered when studying hepatocellular physiology in vitro.