ALBERT

Description: Introduction: Hemophagocytic lymphohistiocytosis (HLH) is a severe hyperinflammatory syndrome caused by excessive immune activation leading to a life-threatening pro-inflammatory cytokine storm. HLH is not an entity of its own, but a clinical syndrome triggered by various conditions like infections, malignancy or autoimmune disorders. HLH pathogenesis is complex and still not fully understood. Contributing factors include immunosuppression (chemotherapy, long-term immunosuppressive therapy), cytokine release from tumor cells, imbalance between infected and immune effector cells as well as genetic predisposition. Despite improved HLH-specific therapy (adapted components from the pediatric HLH-1994 protocol), prognosis is still poor among adult HLH patients. Due to the lack of data on adult HLH in Germany, a national multicenter registry (http://www.hlh-registry.org/) was initiated. Methods: Patients (pts) with proven or suspected HLH were registered by 35 institutions across Germany from August 2010 to July 2016. Both HLH-2004 diagnostic criteria and the HScore (www.saintantoine.aphp.fr/score/) were used to confirm HLH diagnosis. To characterize adult HLH patients, data referring to underlying disease, treatment, outcome, clinical manifestations and laboratory findings were recorded. Where available, patient samples were tested for mutations of the perforin gene PRF1 by standard Sanger sequencing. Results: A total of 125 pts (48 female) were enrolled in our registry, of whom 105 pts either fulfilled diagnostic criteria according to HLH-2004 diagnostic guidelines (n=96) or met at least 4 out of 8 criteria and reached HLH-probability of over 90 % in the HScore (n=9). Among these 105 pts, n=38 (36 %) were female and n=67 (64 %) male. Median age at diagnosis was 49 years (range 17 - 81). Trigger diseases were in line with the literature, with infections (n=34) and malignancy (n=40) being most frequent (Table 1). Patients show a wide spectrum of underlying conditions, i.e. allogeneic stem cell transplantation (alloSCT) or HLH-mimicking diseases due to cytokine release in response to blinatumomab therapy. Late onset hereditary HLH was found in 3 pts (XLP-1 and -2 respectively in EBV-coinfected pts, one pt with perforin mutation - see below). 22 of 105 patients (21 %) were tested for PRF1 mutations. A compound heterozygous PRF1 A91V/Q405X mutation was identified in one pt presenting with NK/T-cell lymphoma. Heterozygous PRF1 A91V mutations were found in 2 pts with B-cell lymphoma and HLH following alloSCT respectively, in one case with available buccal swab DNA to prove germline origin of the mutation. Table 2 summarizes clinical and laboratory findings in the cohort. A median ferritin value of 32,000 µg/L underlines the importance to evaluate pts with highly elevated ferritin with respect to potential HLH diagnosis. Apart from fever and splenomegaly, clinical presentation frequently comprised hepatomegaly, liver failure, hyperbilirubinemia, renal failure, lung involvement like ARDS, or bleeding. Treatment included steroids in the vast majority of pts (n=89), often combined with i.v. immunoglobulins (n=47) and etoposide (n=49). After a median follow up time of 164 days, 48/97 pts (49.5 %) were alive, 8 pts were lost to follow up. Survival analysis revealed median overall survival of 454 days (Figure 1a). Comparing malignancy-associated HLH and HLH after alloSCT with infection- and autoimmune-associated HLH or HLH due to unknown triggers, survival was significantly poorer in the malignancy/alloSCT group (Figure 1b). Conclusions: HLH trigger conditions in adult patients in Germany are in accordance with published case series. Outcome in adult HLH is still poor, with malignancy-associated and HLH after alloSCT showing the worst prognosis. Diagnostic vigilance and early treatment is a prerequisite for improving outcome of adult HLH. In particular, high ferritin values should raise suspicion of HLH. Disclosures Hochhaus: Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding.

Description: Acute lymphoblastic leukemia (ALL) diagnosed in the first month of life (congenital ALL) is very rare. Although congenital ALL is often assumed to be fatal, no studies have been published on outcome except for case reports. The present study reports the outcome of 30 patients with congenital ALL treated with the uniform Interfant-99 protocol, a hybrid regimen combining ALL treatment with elements designed for treatment of acute myeloid leukemia. Congenital ALL was characterized by a higher white blood cell count and a strong trend for higher incidence of MLL rearrangements and CD10-negative B-lineage ALL compared with older infants. Induction failure rate was 13% and not significantly different from that in older infants (7%, P = .14), but relapse rate was significantly higher in congenital ALL patients (2-year cumulative incidence [SE] was 60.0 [9.3] vs 34.2 [2.3], P 〈 .001). Two-year event-free survival and survival of congenital ALL patients treated with this protocol was 20% (SE 9.1%). Early death in complete remission and treatment delays resulting from toxicity were not different. The survival of 17% after last follow-up, combined with a toxicity profile comparable with that in older infants, justifies treating congenital ALL with curative intent. This trial was registered at www.clinicaltrials.gov as no. NCT 00015873, and at www.controlled-trials.com as no. ISRCTN24251487.

Description: Key Points Hermansky-Pudlak syndrome type 2 confers a moderate risk for hemophagocytic lymphohistiocytosis.

Description: Primary hemophagocytic lymphohistiocytosis (HLH) is a severe inflammatory condition that is caused by an abnormal accumulation of activated macrophages. The common NK- and cytotoxic T-cell dysfunction in HLH has been related to genetic defects of vesicle transport and apoptosis. Here, we identify a novel form of hereditary HLH that is related to and possibly caused by a homozygous missense mutation (G139V) of the active centre of heme oxygenase-1 (HO-1). The clinical picture of this male patient is characterized by persistent, severe microcytic hemolytic anemia without hyperbilirubinemia since birth. In the second year of life, the clinical syndrome of HLH developed with persistent fever, marked hepatosplenomegaly, thrombocytopenia, hyperferritinemia, hypertriglyceridemia, and elevated levels of soluble CD25. Liver histology showed marked lympho- and erythrophagocytosis, which confirmed the diagnosis of HLH. The clinical syndrome responded well to immunosuppressive treatment with etoposide, dexamethasone and cyclosporine A. PRF1, UNC13-D and STX11 mutations were excluded. Functional analysis of NK-activity showed only slightly decreased killing activity with normal responses of NK-cells to cytokine stimulation. Persistent endothelial damage was indicated by a pronounced elevation of vWF and D-dimer concentrations. The persistently very low bilirubin levels despite marked hemolysis suggested a defect of hemoglobin degradation. HO-1 is the rate limiting enzyme of this pathway resulting in bilirubin synthesis. Normally, HO-1 is constitutively expressed at a low level but strongly induced by heme, and hypoxia. Functional analysis of HO-1 protein in the patient’s mononuclear blood cells showed a high constitutive expression that did not respond further to heme treatment. Sequence analysis revealed a homozygous GGT (Gly) to GTT (Val) point mutation of HO-1 codon 139. This mutation of the enzyme’s active center is known from in-vitro analyses to change enzyme activity from an oxygenase into a peroxidase. Consistent with this change in activity, quantitative analysis of reactive oxygen species (ROS) metabolites in the urine of this boy showed high amounts of bilirubin oxidative metabolites, 8-hydroxy-2′-deoxyguanosine and acrolein-lysine. In this report we thus identify a novel disease entity which is characterized by an activating mutation of HO-1 which results in a defect of hemoglobin degradation, bilirubin synthesis and in excessive oxidative stress and inflammation.

Description: Introduction: Familial Hemophagocytic Lymphohistiocytosis (FHLH) is an autosomal recessive disease affecting young children. It manifests itself as a severe hyperinflammatory syndrome with activated macrophages and T-lymphocytes. Mutations in Perforin (PRF1) are responsible for FHL-2 in about 15 to 50 % of the patients. Defective granule exocytosis due to mutations in UNC13D have recently been described as the genetic defect underlying FHL-3. Both types are phenotypically not distinguishable from one another. Patients and Methods: Genetic analysis was performed in either the 2 or the 32 exons of PRF1 and UNC-13D, respectively, in 61 patients with hemophagocytic lymphohistiocytosis (HLH) of different geographic origin (28 from Germany, 24 from Turkey and 9 from other countries). Results: We identified mutations in 28 samples investigated (18 in PRF1, 10 in UNC-13D). Besides several known mutations novel deletions, missense and nonsense mutations were detected in both genes scattering the whole coding regions. The Trp374stop mutation (11/13 patients with a PRF1 mutation from Turkey) was the only single mutation repeatedly observed in several patients with a common geographic origin. In HLH patients from Germany, 7/28 patients showed homozygous or compound heterozygous mutations in either PRF1 (3) or UNC13D (4). In 24 HLH patients from Turkey 13 patients with a mutation in PRF1 and 4 patients with a mutation in UNC13D were identified. In addition, four out of 9 patients with a different geographic origin showed mutations in one of this genes. In 10 patients only one heterozygous mutation in UNC13D was detected so far. In these cases, control samples or regulatory regions of the gene have to be analysed to ascertain the relevance of the underlying mutation. Defect Turkey German Other All UNC13D 4 4 2 10 PRF1 13 3 2 18 Unknown 7 21 5 33 Discussion: Overall, UNC-13D and PRF1 mutations were detectable in 45% of the cases. Our results indicate that FHL-2 and FHL-3 account for more than 60% of the HLH cases of Turkish origin with only a small number involving UNC-13D. In contrast, gene defects were identified in only 25% of the patients with a German ancestry. This work was supported by the Foerdergemeinschaft Kinderkrebszentrum Hamburg e. V.

Description: Abstract 2110 Introduction: Iron chelation is the life-saving therapy in patients with chronic transfusion therapy. Treatment with deferoxamine, deferiprone or deferasirox has dramatically improved the life expectancy, but still myocardial siderosis and hepatic siderosis is cause of morbidity and mortality in regularly transfused patients with ß-thalassemia major (TM) or Diamond-Blackfan Anemia (DBA). Deferasirox (DSX) a once-daily oral iron chelator has demonstrated efficacy in reducing hepatic iron and body iron burden, as well as cardiac iron. But in patients with severe cardiac siderosis (T2* ≤ 10ms) a combination therapy with deferiprone (Ferriprox®) and deferoxamine (Desferal®) is the recommended therapy. However, some patients will not benefit from this treatment due to unacceptable toxicity, poor response or noncompliance. Method: We tested a twice-daily deferasirox (Exjade®) -dose with special respect to its efficacy on reducing cardiac iron overload. A group of six patients with severe secondary siderosis was studied, TM (n=5) and DBA (n=1), (5 females, age 8–37 years, mean age 27.7 years). In all patients the liver iron concentration was measured repeatedly by SQUID biosusceptometry or by magnetic resonance imaging (MRI) using the MRI-R2 technique (St. Pierre et al, 2005). In 4 patients with severe cardiac siderosis (T2* ≤ 10ms) we also followed the cardiac iron concentration by MRI using a single breath-hold, multi-echo T2* method. Patients received a daily DSX dose of 19 mg/kg/d – 45 mg/kg/d, with a mean dose of 32 mg/kg/d. Results: The mean initial liver iron concentration of 2.7 mg/g-liver (0.96 – 5.5mg/g) decreased to 1.5 mg/g-liver (0.6 – 3.9 mg/g). The mean monthly liver iron clearance was 6.8%/month (1.7 – 16.8%/month) in a treatment interval of 4 – 26 months (mean: 9.8 months), the patients demonstrated a significant liver iron reduction of 44.4%. The mean serum ferritin was reduced from 3048 μg/l to 1786 μg/l. The mean monthly cardiac iron clearance was 3.1%/month (1.2 – 4.7%/month) and the mean T2* value improved from 9.5 ms to 14.3 ms (+50%). We showed a substantial improvement in patients with severe cardiac siderosis with a T2* improvement of 50 % after a mean treatment period of 12 months with a mean DSX dose of 32 mg/kg/d. In comparison, an improvement of 23.8% was found in 6 patients with T2* 〈 10 ms, after a treatment period of 18 months with a once daily DSX mean dose of 38 mg/kg/d (Pathare et al, 2010). Other authors reported an improvement of 10.8% in 47 patients (T2* 〈 10 ms, treatment period 12 months) with a once daily DSX mean dose of 32 mg/kg/d (Pennell et al, 2009). No severe side effects were seen in our patients and only minor increases in creatinine values, which were reversible with dose reduction. Conclusion: Deferasirox divided in twice daily doses is a safe and effective therapy for patients with severe cardiac iron overload (T2* 〈 10ms) or hepatic iron overload, who do not well tolerate a combination therapy with deferiprone and deferoxamine. Disclosures: Off Label Use: Deferasirox (Exade)is given instead of a once daily dose, in a twice daily divided dose. The daily dose of Deferasirox is in recommended range.

Description: Abstract 5168 Chelation treatment of iron overload from chronic blood (RBC) transfusion is still a challenge to both, patients and medical caretakers. Different treatment regimes have been recommended so far, especially for chronically transfused patients with low or even normal liver iron concentration. We report the results from 16 regularly transfused patients with thalassemia major (TM) who were on iron chelation treatment under normal to mild liver iron concentration (LIC). All patients received deferoxamine (DFO) treatment before they changed to deferasirox (DSX) treatment. 16 TM patients (mean age 13.6 y) were treated with DSX (median dose 18 mg/kg/d, range: 7 – 33 mg/kg/d) for 6 to 71 months. Liver iron measurements by biomagnetic susceptometry (BLS) and/or MRI-R2 as well as cardiac MRI-R2* were performed in intervals of 6 to 12 months. The median LIC was 782 μ g/g-liver wet weight (range: 460 μ g – 1122 μ g). Median RBC transfusion rate was 8500 ml/y, equivalent to about 2 erythrocyte concentrates per 3 weeks or a daily iron influx of 16.2 mg/d. For each measurement interval, the ratio of daily iron influx and DSX dose rate was calculated. This represents the equilibrium molar efficacy for iron balance. In all 16 TM patients no severe side effects were observed and creatinine was in the normal range of 〈 0.9 mg/d throughout the treatment with DSX. From baseline DFO treatment interval to the endpoint of DSX treatment, liver iron decreased by 124 – 4689 μ g/g-liver (conversion factor of 6 for mg/g-dry-wgt), while serum ferritin decreased by -596 to 8283 μ g/l. For all measurement intervals, molar chelation efficacies between 18 % and 56 % were calculated at equilibrium with a median efficacy of 31 % (interquartile range = 16 %). This agrees with molar efficacies of DSX reported earlier, but for relatively higher LIC and chelation doses (Blood 2005; 106(11):#2690 and Blood 2007; 110(11):#2776). The cardiac R2* (median R2* = 38 s-1) was either below the normal threshold of 50 s-1 (T2* 〉 20 ms) or decreased by about 24 %/y under DSX treatment. In these few patients at low LIC, this was even higher than recently reported. Conclusion: Even in patients with normal to mild LIC iron chelation treatment with DSX is safe, does not result in increased creatinine levels or severe side effects and is as efficient as in patients with higher LIC. Disclosures: No relevant conflicts of interest to declare.