ALBERT

Beschreibung: Molecular measurable residual disease (MRD) assessment is not established in approximately 60% of acute myeloid leukemia (AML) patients because of the lack of suitable markers for quantitative real-time polymerase chain reaction. To overcome this limitation, we established an error-corrected next-generation sequencing (NGS) MRD approach that can be applied to any somatic gene mutation. The clinical significance of this approach was evaluated in 116 AML patients undergoing allogeneic hematopoietic cell transplantation (alloHCT) in complete morphologic remission (CR). Targeted resequencing at the time of diagnosis identified a suitable mutation in 93% of the patients, covering 24 different genes. MRD was measured in CR samples from peripheral blood or bone marrow before alloHCT and identified 12 patients with persistence of an ancestral clone (variant allele frequency [VAF] 〉5%). The remaining 96 patients formed the final cohort of which 45% were MRD+ (median VAF, 0.33%; range, 0.016%-4.91%). In competing risk analysis, cumulative incidence of relapse (CIR) was higher in MRD+ than in MRD− patients (hazard ratio [HR], 5.58; P 〈 .001; 5-year CIR, 66% vs 17%), whereas nonrelapse mortality was not significantly different (HR, 0.60; P = .47). In multivariate analysis, MRD positivity was an independent negative predictor of CIR (HR, 5.68; P 〈 .001), in addition to FLT3-ITD and NPM1 mutation status at the time of diagnosis, and of overall survival (HR, 3.0; P = .004), in addition to conditioning regimen and TP53 and KRAS mutation status. In conclusion, NGS-based MRD is widely applicable to AML patients, is highly predictive of relapse and survival, and may help refine transplantation and posttransplantation management in AML patients.

Beschreibung: Abstract 3795 Introduction: Virus-associated hemophagocytic syndrome (VAHS) is a severe complication of various viral infections often resulting in multiple organ failure syndrome and death. Based on previous experience with seasonal (H3N2) and avian (H5N1) infections the 2009 influenza A (H1N1) virus offers the potency of VAHS-development resulting in an aggressive and life-threatening disease. Most patients infected by the novel human influenza A (H1N1) experienced a mild clinical course; however some patients become critically ill with respiratory failure requiring intensive care and ventilator support. Multiple organ failure was one of the leading causes of death suggesting that patients with severe influenza A (H1N1) infection may develop a virus-associated hemophagocytic syndrome (VAHS). Methods: To evaluate frequency, clinical course and outcome of VAHS in critically ill patients a prospective observational study was performed at a single center intensive care unit in Hannover, Germany. Collected data include demographics, comorbid conditions, viral shedding, diagnosis of VAHS, illness progression, treatments and survival. VAHS was suspected when patients developed fever, cytopenia affecting at least two lineages, hepatitis or splenomegaly, hemophagocytosis in bone marrow samples and/or increased serum levels of sIL-2R and ferritin. Diagnosis of VAHS was made according to established HLH-diagnostic criteria. Primary outcome variables were the development of VAHS and VAHS-associated mortality. Results: Between October 5, 2009 and January 4, 2010 twenty five consecutive critically ill patients with RT-PCR confirmed 2009 influenza A (H1N1) infection and respiratory failure were identified. VAHS developed in nine out of 25 (36%) patients. Treatment of VAHS was started in 6 out of the 9 patients; three patients showed terminal disease and were no longer considered candidates for treatment with etoposide and dexamethasone. Despite VAHS-directed therapy, 5 out of the 6 patients died from uncontrolled progress leading to multiorgan failure. Overall, 8 out of the nine patients (89%) with confirmed VAHS died. In contrast, the mortality rate in the remaining 16 patients without VAHS was 25% (p=0.004). Patients were young (median, 45 [IQR, 35–56] years), however 18 (72%) presented one or more risk factors for a severe course of influenza illness. All 25 patients received mechanical ventilation for severe acute respiratory distress syndrome and refractory hypoxemia with duration of mechanical ventilation of median (IQR) 19 (13-26) days. Additionally 17 patients (68%) required extracorporeal membrane oxygenation for median (IQR) 10 (6-19) days. Oseltamivir and zanamivir were used as antiviral treatment in 24 patients (96%) for a median (IQR) of 7 (4-10) days and in 15 patients (60%) for a median (IQR) of 7 (5-12) days, respectively. The median (IQR) duration of viral shedding from disease-onset to the last positive H1N1 RT-PCR was 19 (14-26) days. In patients without VAHS the median (IQR) viral shedding time was 15 (12-22) days as opposed to 21 (14-26) days (p=0.13) in patients with VAHS. Conclusion: The present case series confirms previous post mortem analysis that severe influenza A (H1N1) infection is an important contributor to the development of VAHS in critically ill patients. Development of VAHS was associated with fatal outcome showing rapid clinical deterioration and multi organ failure syndrome and either contributes greatly to, or is itself causative of death in this patient population. Our findings are preliminary but potentially have important implications for future management of patients with influenza A (H1N1) disease as well as other severe virus infections which can induce secondary hemophagocytic syndromes. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Background: Relapse occurs in 30-40% of AML patients undergoing allogeneic hematopoietic stem cell transplantation (alloHSCT). Detecting molecular relapse before clinical relapse offers the opportunity of early interventions (e.g. donor lymphocyte infusions, reduction of immunosuppression etc.). Next-generation sequencing (NGS)-based error-corrected sequencing approaches have shown promising results in AML patients prior to alloHSCT, which identified MRD in 45% of patients and predicted a cumulative incidence of relapse of 66% versus 17% in MRD negative patients at 5 years. However, NGS-based MRD is not well studied in patients after alloHSCT. Aim: To evaluate the prognostic impact of MRD on day 90 and day 180 after alloHSCT in AML patients in morphologic complete remission (CR) using error-corrected NGS applicable to the majority of AML patients. Patients and Methods: We quantified MRD in 138 patients who underwent myeloablative (MA, n=47) or reduced-intensity conditioned (RIC, n=91) alloHSCT for AML on day 90 and 180 after alloHSCT. All patients had at least one mutation at the time of diagnosis that was identified by NGS with a myeloid panel on the Illumina platform. Amplicon-based error-corrected sequencing and bioinformatics analysis was applied to samples on day 90 (n=133) and day 180 (n=125) after alloHSCT as described previously (Thol et al. Blood 2018). In the first approach we analysed 1-2 diagnostic mutations (=limited marker approach). In the second approach an extended marker set with (2-4) markers was used (=extended marker approach). Genomic DNA from peripheral blood (PB) was used for the majority of analyses (PB n= 394; bone marrow n=17). Cumulative incidence of relapse (CIR) and non-relapse mortality (NRM) were evaluated by competing risk analysis. Results: The median follow up time of the cohort was 5.5 years. The mean limit of detection was a variant allele frequency (VAF) of 0.012% using error correction and 0.071 when using forward/reverse read error correction. MRD positivity on day 90 and/or day 180 was detected in 22 out of 138 patients (16%) with the limited marker approach, while MRD was found in 28 patients (20.3%) with the extended marker approach. Using the limited marker approach, the 5-year CIR was 52% for MRD positive and 30% for MRD negative patients (P=0.001), while NRM was similar between both groups (Figure 1A). Overall survival (OS) was shorter in MRD positive patients compared to MRD negative patients (P=.044, Figure 1B). In multivariate analysis using variables significant in univariate analysis (P