Abstract: OBJECTIVE: We report the outcome of pregnancies treated with intravenous immunoglobulin (IVIG) for severe red blood cell alloimmunization, evaluating whether IVIG defers the development of severe fetal anaemia and its consequences. BACKGROUND: Although fetal anemia can be treated very successfully with intrauterine transfusion (IUT), procedures before 20 weeks' gestation can be very challenging technically and may be hemodynamically stressful to an extremely premature and already compromised fetus. The procedure-related fetal loss rate is approximately 5.6% for IUTs performed & lt; 20 weeks' gestation, compared to 1.6% overall. IVIG may prevent hemolysis and could therefore be a noninvasive alternative for early transfusions. STUDY DESIGN: We included consecutive pregnancies over a nineteen year period in the Fetal Medicine Unit, Mount Sinai Hospital, University of Toronto, Canada, of alloimmunized women with a history of severe early onset haemolytic disease who received IVIG until intrauterine transfusion could safely be performed. Previous untreated pregnancies were used as controls. IVIG therapy was commenced between 11 and 14 weeks' gestation. Our usual protocol was IVIG 2 g/kg per week every 3 weeks, until the first IUT could be performed. Each 2g/kg dose was administered over 2 days, 1g/kg per day, to reduce the chance of severe headaches. In three pregnancies, IVIG 1g/kg was given weekly. We compared the clinical outcomes (gestation at first IUT, fetal Hb at first FBS, gestation at delivery, perinatal survival) between previous pregnancies without IVIG and the subsequent pregnancy treated with IVIG. In comparing fetal Hb's between two pregnancies, a linear relationship between fetal Hb and gestation was used to correct for variable gestations. The fetal Hb was converted to a standardized fetal Hb value (multiples of the standard deviation [SD]). Statistical analysis was performed on 'Statistical Package for Social Science Version 16.0' (SPSS Inc, Chicago, Illinois). RESULTS: Seventeen women referred to our unit for a previous pregnancy loss secondary to severe RBC alloimmunization received IVIG treatment in 20 subsequent pregnancies; all eventually requiring intrauterine transfusion. For previous early losses despite transfusion, immunoglobulin was associated with a relative increase in fetal hemoglobin between treated and untreated pregnancies of 32.6 g/L (95%CI 15.2-50.0, P=0.003) and improved perinatal survival (8/8 vs 0/6, P=0.001). For previous losses & lt;20 weeks, it enabled first transfusion deferral in subsequent pregnancies to at least 19.9 (mean 23.2) weeks. Of the 17 live-born babies from IVIG-treated pregnancies, three (18%) required an exchange transfusion, eight (47%) a simple "top-up" transfusion, and six (35%) phototherapy. CONCLUSION: Our results show that, among severely sensitized cases with previous early fetal loss despite IUT, use of IVIG in subsequent pregnancies is associated with a significantly higher fetal Hb before first IUT, deferral of first IUT, delivery at a later gestation and increased perinatal survival. The timing of the first FBS/IUT was delayed by 3 weeks in pregnancies treated with IVIG compared to a previous untreated pregnancy. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Abstract: Background/Objectives The optimal management of pregnancy with sickle cell disease (SCD) is controversial (Cohen and O'Brien, Chapter 14, 2012). In particular, the value of prophylactic exchange transfusions in preventing complications from SCD and/or pregnancy itself is unclear. For the past decade at University College London Hospitals (UCLH), UK, we have undertaken prenatal exchange transfusions for most patients with sickle cell syndromes (SCS), HbSS, SC or Sb-thalassaemia. This is due to the poor outcomes we had experienced previously in a proportion of patients in whom prophylactic transfusion was not undertaken. We review the outcome over seven years in all such pregnancies managed at UCLH. Methodology/Patients We undertook a retrospective analysis of pregnancy outcomes of women with SCS delivering between 1 Jan 2011 and 31 Dec 2017. Data were collected from electronic hospital-held records. We recorded 50 pregnancies in 22 women, of which 25 resulted in live births. Most women had HbSS (82%), 14% had HbSC and 4% HbS-bthalassemia. The early pregnancy ( 〈 12 weeks) miscarriage rate of 28% was greater than the UK National Guidelines, 2012, which was 22% (Table 1). The median age (range) of our patients was 32 (19-40) years, which might contribute to the increased miscarriage rate. One woman sustained 6 of the 11 recorded miscarriages. Only 4% of pregnancies occurred in women 〉 40, which is similar to national data for England (3.6%) (Centre for Maternal and Child Enquiries [CMACE], 2011). This age distribution is comparable to that in the population of Oteng-Ntim et al, 2015, who looked at pregnancy outcomes in a UK sickle population between 2010 and 2011. Patients were managed antenatally with exchange transfusions where there were no contraindications with respect to prior allo-immunisation, to reduce sickle related complications and improve pregnancy outcomes. Eight All patients were adequately hydrated antenatally. To gain insight into the effect of this management on maternal complications and fetal outcomes, these were compared with a recently reported cohort of SCS in London who did not receive exchange transfusions (Oteng-Ntim et al, 2015). Results Painful crises requiring admission antenatally were 9% compared with 17.6% reported by Oteng-Ntim et al, 2015. There were no admissions with sickle chest syndrome, either antenatally or postnatally, compared with 6.4% (Oteng-Ntim et al.). Preterm labour before 34 weeks' and 37 weeks' gestation occurred in one case (4.5%) and in 38% of cases, compared to 44.2% reported by Serjeant et al, 2004 and 47.1% reported by Oteng-Ntim et al before 37 weeks. There were no late miscarriages or stillbirths in our cohort. By contrast, in Oteng-Ntim et al, the stillbirth rate was 2.8% which is higher than the 0.92% reported in the general population (CMACE, 2011). The Caesarean section (CS) rate was 64%, which is higher than previously reported (39% in Barfield et al, 2010, 52.9% in Oteng-Ntim et al, 2015) (Table 3). One woman sustained a significant post-partum haemorrhage. The following complication rates were similar to those reported by Oteng-Ntim et al, 2015 (Table 3): postnatal pain: 18% vs 21.6%; pre-eclampsia 8% vs 7.8%; intrauterine growth restriction (IUGR): 22% vs 27.3% (Table 4). Four patients had pre-existing allo-antibodies, of whom three had two or more antibodies. No new allo-immunisations developed during pregnancy. Conclusions Although our cohort numbers are small, they suggest a lower rate of painful crisis/chest syndrome than reported in series of SCS in which prophylactic exchange transfusions were typically not practised. While the rate of IUGR was lower than in historic reports, the difference was less clear than for painful crisis. There were no stillbirths in our series, despite the previously documented high incidence in women with SCS. The high rate of CS reflects local practice by clinicians wishing to avoid prolonged labour in this patient population. No serious adverse reactions to transfusion in pregnancy were seen despite one patient having had a previous hyperhaemolytic episode outside pregnancy. This study shows that prophylactic antenatal exchange transfusion is well tolerated and associated with fewer veno-occlusive crisis complications than historically reported with sickle disorders. Randomised studies are indicated in a larger cohort of patients to determine whether other benefits or risks accrue from this approach. Disclosures Porter: Cerus: Honoraria; Novartis: Consultancy; Agios: Honoraria.

Abstract: Eltrombopag (ELT), a small molecular thrombopoietin (TPO) mimetic approved for children and adults, could offer a therapeutic alternative to selected neonates and young children with chronic thrombocytopenias. ELT has also been proposed as a potential anti-cancer drug due to its anti-proliferative effects in tumor cells, which are mediated by its strong iron chelating properties. This raises the potential concern that rapidly proliferating normal cells, like bone marrow cells in neonates or young infants, could also be susceptible to the anti-proliferative effects of ELT. In this study, we first assessed the responses of neonatal (cord blood, CB) and adult (peripheral blood, PB) megakaryocyte (MK) progenitors to increasing concentrations of ELT, TPO and Romiplostim (ROM) in vitro. Consistent with the previously described pattern of neonatal megakaryopoiesis, CB progenitors generated 10-times more MKs, which were of lower ploidy but had higher CD42 surface expression levels than PB-MKs. Unlike TPO or ROM, ELT exhibited dose-dependent opposing effects on in vitro megakaryopoiesis: Low concentrations (≤6µM) stimulated megakaryopoiesis, but high concentrations (30µM) suppressed MK differentiation and proliferation, as evidenced by a severe reduction in the percentage and absolute number of CD41+ cells after 7 days of culture. The toxic effects of high ELT concentrations were not abrogated by the addition of TPO at concentrations achieved in hyporegenerative thrombocytopenias (3 ng/mL), and were reproduced by the addition of the iron chelators deferoxamine (DFO) or deferiprone (DFP) at concentrations of 100µM to MK cultures, suggesting iron deficiency as a potential mechanism. To further assess the iron chelating effects of ELT in human MKs, we used the calcein assay. In K562 cells, as well as in CB MKs, ELT concentrations 〉 10µM reduced the labile intracellular iron pool (LIP) to lower levels than the iron chelator DFP at a concentration of 200 µM, indicating the strength of ELT's iron chelating properties. These concentrations of ELT also resulted in a severe reduction of mitochondrial transmembrane potential, detected by the JC-1 assay, decreased proliferation (EdU click assay) and apoptosis (Annexin-V binding). During MK differentiation, committed MK progenitors (CD34+/CD41+ cells) were the cell population most sensitive to the antiproliferative and apoptotic effects of high-dose ELT. Next, we evaluated whether the iron status of differentiated MK progenitors would influence their responses to ELT. To test this, we generated iron-depleted and iron-repleted CB-MK progenitors by culturing CD34+ cells for 7 days in the presence of apo-transferrin (iron-free) or holo-transferrin (iron-saturated), followed by a 72 hour culture with different concentrations of ELT. These studies revealed a strong iron status/ELT dose interaction (interaction p 〈 0.0001); In iron-depleted cultures, 30 µM ELT and 100 µM DFO similarly impaired MK expansion and induced MK apoptosis, but these effects were attenuated in iron-repleted cultures. Interestingly, iron-depleted adult PB progenitors did not show reduced MK expansion or increased apoptosis in response to 30 µM ELT, suggesting a lower susceptibility of adult megakaryopoiesis to the iron chelating effects of ELT. Taken together, these findings suggest that iron status is an important variable affecting the response to ELT, particularly in neonates and children. Disclosures Cooper: Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Rigel: Consultancy, Membership on an entity's Board of Directors or advisory committees; Principia: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Porter:Celgene: Consultancy, Honoraria; Protagonism: Honoraria; Agios: Consultancy, Honoraria; La Jolla: Honoraria; Vifor: Honoraria; Silence therapeutics: Honoraria; Bluebird bio: Consultancy, Honoraria. Bussel:Regeneron: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; argenx: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; RallyBio: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Kezar Life Sciences: Consultancy, Membership on an entity's Board of Directors or advisory committees; Physician Education Resource: Speakers Bureau; 3S Bio: Speakers Bureau; Rigel: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees; Tranquil: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Momenta Pharmaceuticals: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Dova Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; UCB: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Sola-Visner:Sysmex America, Inc.: Other: Laboratory equipment on loan, Research Funding.

Abstract: Background: Kell and Rhesus (Rh) maternal red blood cell (RBC) alloimmunization are the most common causes of severe fetal haemolytic disease. Widespread use of anti-D immune globulin has dramatically reduced the incidence of Rh(D) alloimmunization, leaving K alloimmunization responsible for a significant proportion of cases of fetal anemia requiring intrauterine transfusion ( ). K antigens are expressed on fetal erythrocytes at 10-11 weeks and k antigens at 6-7 weeks gestation (Tovey et al, 1986). In alloimmunized women, erythroid specific antibodies traverse the placenta, causing immune destruction of fetal erythroid cells leading to progressive haemolytic anaemia. The mechanism of fetal anaemia in K alloimmunization differs somewhat from that in Rh-D, in that anti-K antibodies cause suppression of fetal erythropoiesis (Gariod et al, 2004; Weiner et al, 1996). Whilst IUT of RBCs has improved fetal and neonatal survival, important information such as the critical anti-K titres to guide appropriate timing and frequency of IUT, is somewhat conflicting. Currently anti-K alloimmunized pregnancies do not have a standardized protocol for titer monitoring throughout pregnancy and, once alloimmunized, patients are usually referred to a regional fetal center for close ultrasound (US) surveillance. Our primary objective is to determine from a retrospective analysis of our population whether there is a critical anti-K titer that should trigger intensive US monitoring or intervention and to investigate the rate of progression of fetal anemia following IUTs. Methods: This is a retrospective single-center study at Mount Sinai Hospital (MSH), Toronto, Canada, of all pregnant patients with anti-K as the primary alloimmunizing antibody, between 1991 and 2018. MSH is the largest fetal medicine center in Canada and the largest referral center for IUTs. Ethical approval was granted by the Research Ethics Board (REB # 12-0113-C). Data were obtained from a database of patient medical records, US reports and the transfusion medicine laboratory, including maternal demographics, pregnancy history, presence of other alloantibodies and hemoglobin concentration before and after all IUTs. Neonatal outcomes included survival, mode of delivery, gestational age (GA) at delivery, birth weight and need for neonatal exchange transfusion, phototherapy or IVIG. Data were analyzed using GraphPad Prism 6 and linear correlations are expressed as a p-value. Results: Thirty-eight women underwent 163 IUTs in 44 pregnancies where K was the predominant antibody. Two patients in whom anti-K was a secondary antibody were excluded. In 5 of these pregnancies, 2 had a total of three alloantibodies and 5 had 2 alloantibodies each. The median maternal age was 31 (29 - 35) years. Four women had a history of intrauterine fetal death (IUFD) and 9 of neonatal haemolytic disease. The median GA at 1st IUT was 24.2 weeks (14.9-34.7), and there was a median of 4 IUTs per patient. There were seven cases of hydrops fetalis. The number of IUTs a patient received throughout pregnancy was correlated directly with the anti-K titer (Figure 1). Every 4-fold dilution resulted in a further increase in the IUT number by 2.2 above the mean of 2.5 at a titer of 1:32 (p=0.0137). Figure 2 illustrates the correlation between the GA at 1st IUT and antibody titer. IUTs were required at earlier GAs if the titers were higher. Following a 1:32 titer, every 2-fold titer increase reduced the mean gestational age at 1st IUT by 2 weeks (p 〈 0.0001). No pregnancy required an intervention with an anti-K titer 〈 1:32. The median fetal Hb at the first IUT was 48 (8-91) g/L with an average daily decrease of hemoglobin of 4.7 g/L/day between the first and second IUT (Figure 3), which is similar to the 4.1 g/L/day previously reported in Rh(D) alloimmunization. The median GA was 37.1 (24-39.7) weeks at delivery with median birth weight centile of 59.5%; 8.9% of neonates required a blood transfusion and 24.4% required phototherapy. There were four IUFDs and one neonatal . Conclusion: Our data support a critical anti-K titer of 1:32 and support a role for anti-K titer monitoring as a predictor of disease severity, counselling women appropriately and establishing a balance between paternal K antigen typing, US middle cerebral artery peak systolic velocity monitoring of the fetus and IUTs. Disclosures Garbowski: Vifor Pharma: Consultancy; Imara: Consultancy.

Abstract: Background: The rhesus (Rh) and Kell blood group systems are the most common of over 50 different antigens capable of causing maternal red blood cell (RBC) alloimmunization and severe fetal hemolytic disease. Anti-K and anti-D are responsible for a significant proportion of fetal anemia requiring intrauterine transfusion (IUT). Whilst IUT of packed RBCs improves neonatal survival and morbidity, clinical prognostic indicators are lacking. Our primary objective was to identify predictors of adverse outcome. Methods: We conducted a retrospective single-center study at Mount Sinai Hospital (MSH), Toronto, Canada. All pregnant patients alloimmunized with anti-K and anti-D as a single antibody, between 1991 and 2018 were included. Data were obtained from patient medical records, ultrasound reports and information from the transfusion medicine laboratory. Data included maternal demographics, antibody titers, pregnancy history, number of IUTs, hemoglobin (HB) concentration at the beginning and end of all IUTs. Neonatal outcomes included survival, mode of delivery, gestational age at delivery, birth weight, HB at birth and need for neonatal transfusion, phototherapy or intravenous immunoglobulins (IVIG). Our primary outcome was the composite outcome of stillbirth or neonatal death (SB/NND). We also constructed a secondary outcome consisting of top-up neonatal transfusion, exchange transfusion, phototherapy, or use of IVIG. Medians and interquartile ranges (IQR) or mean±SD were used as summary statistics and compared by Mann-Whitney or t-test; p & lt;0.05 was statistically significant. Outcome predictors were identified using multivariable logistic or linear regression analysis; accounting for patient level clustering by marginal Generalized Estimating Equation did not significantly affect variables. Data were analyzed using SPSS. Results: 116 women with 128 pregnancies and 425 IUTs with anti-K or anti-D as a single antibody were identified. Median maternal age was 31 years (27.0-35.0) for anti-K and 32 years (23.6-40.6) for anti-D. The gestational age at 1st IUT differed significantly between anti-K and anti-D (24.3 vs 28.7 weeks respectively, p=0.004). Women with anti-K antibodies required more IUTs than women with anti-D (3.84 vs 3.12 IUTs, p=0.036) and HB at 1st IUT was significantly lower in the anti-K group (5.10 vs 7.05 g/dL, p=0.001) (Table 1). Following initiation of IUT, the time from 1st IUT to delivery was 69.6 days in the anti-K group and 54.6 in the anti-D group (p=0.06). The daily decrease of HB between 1st and 2nd IUT (as a marker of disease severity), development of fetal hydrops and severe preterm birth did not differ significantly between the two groups. Mean gestation age at delivery was 35.0 weeks in the anti-K and 36.0 weeks in the anti-D group (p=0.28), with 87.1% and 93.9% survival (p=0.37), respectively. The proportion of neonates requiring phototherapy, IVIG and exchange/top-up transfusion was comparable across the two antibody groups (Table 1). Regression analysis showed that delivery occurred sooner if HB dropped more rapidly between the first two IUTs (p=0.01). Each additional transfusion gained on average 22.5 days in utero (Table 2). In multivariable analysis, gestational age at 1st IUT was the only predictor of a SB/NND outcome (adjusted OR 0.79 [95%CI 0.67-0.93]; p=0.006). With 1st IUT at 23 weeks, the risk of SB/NND was 8%, but only 2.5% at 28 weeks and & lt;0.01% at 36 weeks (Table 3). Gestational age at delivery (adjusted OR 1.11 [95%CI 1.00 to 1.23]; p=0.046) and HB at 1st IUT (adjusted OR 0.84 [95%CI 0.72 to 0.99] ; p=0.038) were the only predictors of a composite adverse blood product requirement/intervention outcome (Table 4). The odds of blood product requirement/intervention postnatally at 1st IUT (median HB at 1st IUT of 6.6g/dL) at 28 weeks were 43% and increased by a further 38% for delivery at 34 weeks. Conclusion: The earlier in gestation that IUTs are implemented, the higher the odds of a SB/NND; however the later the gestation at delivery, the greater the odds of the neonate requiring blood products post-partum. The greater the HB drop between the 1st and 2nd IUT, the shorter the 'time between the first IUT and delivery', which increases the odds of a SB/NND outcome. Disclosures Garbowski: Vifor Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees; Imara: Consultancy. Shehata:Ferring: Honoraria.