Malte von Bonin

Long-Term Prognosis of Acute Myeloid Leukemia According to the New Genetic Risk Classification of the European LeukemiaNet Recommendations: Evaluation of the Proposed Reporting System

PURPOSE The current European LeukemiaNet (ELN) recommendations for acute myeloid leukemia (AML) propose a new risk reporting system, integrating molecular and cytogenetic factors and subdividing the large heterogenous group of intermediate-risk patients into intermediate-I (IR-I) and intermediate-II (IR-II). We assessed the prognostic value of the new risk classification in a large cohort of patients. PATIENTS AND METHODS Complete data for classification were available for 1,557 of 1,862 patients treated in the AML96 trial. Patients were assigned to the proposed genetic groups from the ELN recommendations, and survival analyses were performed using the Kaplan-Meier method and log-rank test for significance testing. RESULTS The median age of all patients was 67 years. With a median follow-up of 8.3 years, significant differences between all risk categories were observed in patients age ≤ 60 years regarding the time to relapse, relapse-free survival, and overall survival (OS). Patients in the IR-II group had a better prognosis than patients in the IR-I group. The median OS times in young patients with favorable risk (FR), IR-I, IR-II, and adverse risk (AR) were 5.3, 1.1, 1.6, and 0.5 years, respectively. Separate analyses in the age group older than 60 years revealed significant differences between FR, AR, and IR as a whole, but not between IR-I and IR-II. CONCLUSION In younger patients with AML, the ELN classification seems to be the best available framework for prognostic estimations to date. Caution is advised concerning its use for prospective treatment allocation before it has been prospectively validated. In elderly patients, alternative prognostic factors are desirable for further risk stratification of IR.

A novel prognostic model in elderly patients with acute myeloid leukemia: results of 909 patients entered into the prospective AML96 trial.

We present an analysis of prognostic factors derived from a trial in patients with acute myeloid leukemia older than 60 years. The AML96 trial included 909 patients with a median age of 67 years (range, 61-87 years). Treatment included cytarabine-based induction therapy followed by 1 consolidation. The median follow-up time for all patients is 68 months (5.7 years). A total of 454 of all 909 patients reached a complete remission (50%). Five-year overall survival (OS) and disease-free survival were 9.7% and 14%, respectively. Multivariate analyses revealed that karyotype, age, NPM1 mutation status, white blood cell count, lactate dehydrogenase, and CD34 expression were of independent prognostic significance for OS. On the basis of the multivariate Cox model, an additive risk score was developed that allowed the subdivision of the largest group of patients with an intermediate-risk karyotype into 2 groups. We are, therefore, able to distinguish 4 prognostic groups: favorable risk, good intermediate risk, adverse intermediate risk, and high risk. The corresponding 3-year OS rates were 39.5%, 30%, 10.6%, and 3.3%, respectively. The risk model allows further stratification of patients with intermediate-risk karyotype into 2 prognostic groups with implications for the therapeutic strategy.

Addition of sorafenib versus placebo to standard therapy in patients aged 60 years or younger with newly diagnosed acute myeloid leukaemia (SORAML): a multicentre, phase 2, randomised controlled trial

BACKGROUND Preclinical data and results from non-randomised trials suggest that the multikinase inhibitor sorafenib might be an effective drug for the treatment of acute myeloid leukaemia. We investigated the efficacy and tolerability of sorafenib versus placebo in addition to standard chemotherapy in patients with acute myeloid leukaemia aged 60 years or younger. METHODS This randomised, double-blind, placebo-controlled, phase 2 trial was done at 25 sites in Germany. We enrolled patients aged 18-60 years with newly diagnosed, previously untreated acute myeloid leukaemia who had a WHO clinical performance score 0-2, adequate renal and liver function, no cardiac comorbidities, and no recent trauma or operation. Patients were randomly assigned (1:1) to receive two cycles of induction therapy with daunorubicin (60 mg/m(2) on days 3-5) plus cytarabine (100 mg/m(2) on days 1-7), followed by three cycles of high-dose cytarabine consolidation therapy (3 g/m(2) twice daily on days 1, 3, and 5) plus either sorafenib (400 mg twice daily) or placebo on days 10-19 of induction cycles 1 and 2, from day 8 of each consolidation, and as maintenance for 12 months. Allogeneic stem-cell transplantation was scheduled for all intermediate-risk patients with a sibling donor and for all high-risk patients with a matched donor in first remission. Computer-generated randomisation was done in blocks. The primary endpoint was event-free survival, with an event defined as either primary treatment failure or relapse or death, assessed in all randomised patients who received at least one dose of study treatment. We report the final analysis. This trial is registered with ClinicalTrials.gov, number NCT00893373, and the EU Clinical Trials Register (2008-004968-40). FINDINGS Between March 27, 2009, and Nov 28, 2011, 276 patients were enrolled and randomised, of whom nine did not receive study medication. 267 patients were included in the primary analysis (placebo, n=133; sorafenib, n=134). With a median follow-up of 36 months (IQR 35·5-38·1), median event-free survival was 9 months (95% CI 4-15) in the placebo group versus 21 months (9-32) in the sorafenib group, corresponding to a 3-year event-free survival of 22% (95% CI 13-32) in the placebo group versus 40% (29-51) in the sorafenib group (hazard ratio [HR] 0·64, 95% CI; 0·45-0·91; p=0·013). The most common grade 3-4 adverse events in both groups were fever (71 [53%] in the placebo group vs 73 [54%] in the sorafenib group), infections (55 [41%] vs 46 [34%]), pneumonia (21 [16%] vs 20 [14%]), and pain (13 [10%] vs 15 [11%]). Grade 3 or worse adverse events that were significantly more common in the sorafenib group than the placebo group were fever (relative risk [RR] 1·54, 95% CI 1·04-2·28), diarrhoea (RR 7·89, 2·94-25·2), bleeding (RR 3·75, 1·5-10·0), cardiac events (RR 3·46, 1·15-11·8), hand-foot-skin reaction (only in sorafenib group), and rash (RR 4·06, 1·25-15·7). INTERPRETATION In patients with acute myeloid leukaemia aged 60 years or younger, the addition of sorafenib to standard chemotherapy has antileukaemic efficacy but also increased toxicity. Our findings suggest that kinase inhibitors could be a useful addition to curative treatment for acute myeloid leukaemia. Overall survival after long-term follow-up and strategies to reduce toxicity are needed to determine the future role of sorafenib in treatment of this disease. FUNDING Bayer HealthCare.

Tightly anchored tissue-mimetic matrices as instructive stem cell microenvironments

A major obstacle in defining the exact role of extracellular matrix (ECM) in stem cell niches is the lack of suitable in vitro methods that recapitulate complex ECM microenvironments. Here we describe a methodology that permits reliable anchorage of native cell–secreted ECM to culture carriers. We validated our approach by fabricating two types of human bone marrow–specific ECM substrates that were robust enough to support human mesenchymal stem cells (MSCs) and hematopoietic stem and progenitor cells in vitro. We characterized the molecular composition, structural features and nanomechanical properties of the MSC-derived ECM preparations and demonstrated their ability to support expansion and differentiation of bone marrow stem cells. Our methodology enables the deciphering and modulation of native-like multicomponent ECMs of tissue-resident stem cells and will therefore prepare the ground for a more rational design of engineered stem cell niches.

Adoptive transfer of allogeneic regulatory T cells into patients with chronic graft-versus-host disease

BACKGROUND AIMS Mouse models indicate that adoptive transfer of regulatory T cells (Treg) may suppress graft-versus-host-disease (GvHD) while preserving graft-versus-leukemia reactions. We aimed to develop a protocol for the efficient isolation and in vitro expansion of donor-derived Treg and to establish the proof-of-concept for the clinical application of ex vivo-generated Treg preparations in five patients with otherwise treatment-refractory chronic GvHD (cGvHD). METHODS Allogeneic Treg were isolated from unstimulated leukapheresis products of the corresponding human leukocyte antigen-matched donors by use of clinical-grade magnetic-activated bead sorting. To increase the amount and purity, Treg were cultivated for 7-12 days and infused after a median time of 35 months after allogeneic hematopoietic cell transplantation. RESULTS Final products contained Treg with a median purity of 84.1% CD4(+)CD25(high)CD127(low)FOXP3(+)of CD45(+) cells and a mean quantity of 2.4 × 10(6) Treg per kg body wt. All isolated cell products showed in vitro suppressive activity. On transfusion, two of five patients showed a clinical response with improvement of cGvHD symptoms. The other three patients showed stable cGvHD symptoms for up to 21 months. In four of five patients, increased counts of Treg were detectable on Treg transfusion, immunosuppressive treatment could be reduced and suppression of CD69 activation marker expression on T-effector cells was observed. However, one patient had development of malignant melanoma and another patient had Bowen skin cancer 4 months and 11 months after Treg transfusion, respectively. CONCLUSIONS We demonstrate a feasible and reproducible approach of isolating functional Treg in high quantity and purity for clinical application and show opportunities and risks of adoptive Treg transfer into patients with cGvHD.

Influence of Enzyme-Inducing Antiepileptic Drugs on Trough Level of Imatinib in Glioblastoma Patients

Background: Imatinib mesylate is used in combination with hydroxyurea (HU) in ongoing clinical phase II studies in recurrent glioblastoma multiforme (GBM). CYP3A4 enzyme-inducing antiepileptic drugs (EIAEDs) like carbamazepine, phenytoin, and oxcarbazepine - as well as non-EIAEDs like valproic acid, levetiracetam, and lamotrigine - are frequently used in patients with GBM. Since CYP3A4 is the major isozyme involved in the metabolism of imatinib, we investigated the influence of EIAEDs on imatinib pharmacokinetics (pk). Methods: GBM patients received 600 mg imatinib p.o./o.d. in combination with 1.0 g HU p.o./o.d..together with either EIAEDs, non-EIAEDs, or no antiepileptic drug (non-AEDs) comedication. Trough plasma levels of imatinib and its active main metabolite N-desmethyl-imatinib (CGP74588) were determined biweekly in these patients, total 543 samples being collected from 224 patients (up to 6 times / patient). All three groups were compared to each other and with historical pharmacokinetic data obtained from patients with chronic myeloid leukemia (CML). Results: Mean imatinib trough levels in patients not receiving AEDs ( 1404 ng/ml, CV 64%) and on non-EIAEDs (1374 ng/ml, CV 46%) were comparable with mean imatinib trough levels of the historical control group of CML patients (1400 ng/ml, CV 50%). Mean trough levels of imatinib were reduced up to 2.9-fold (477 ng/ml, CV 70%) in patients treated with EIAEDs. Only slight, but although significant differences were observed in the mean trough level of the metabolite CGP74588 between EIAED-, non-EIAED and no-AED patients, 240 ng/ml (CV 57%) , 351 ng/ml (CV 34%) and 356 ng/ml (CV 52%), respectively. The corresponding mean level for CML patients was 300 ng/ml (CV 50%). Conclusion: Significant decreases of imatinib and CGP74588 trough levels were observed for patients receiving EIAEDs. The EIAED-induced reduction in trough imatinib levels can be avoided by switching to non-EIAEDs comedication or compensated by administering higher imatinib doses. In addition these data demonstrate that there is no significant difference in the pharmacokinetics of imatinib between patients with glioblastoma and CML.

MRI-Based Liver Iron Content Predicts for Nonrelapse Mortality in MDS and AML Patients Undergoing Allogeneic Stem Cell Transplantation

Purpose: Retrospective, surrogate marker–based studies have found inconsistent associations between systemic iron overload (SIO) and adverse outcome in patients undergoing allogeneic stem cell transplantation (allo-SCT). As a consequence, the impact of SIO in this context remains under debate. The aim of this study was to test whether the objective pretransplant quantification of liver-iron content (LIC) by magnetic resonance imaging (MRI) could circumvent these limitations and conclusively define the prognostic relevance of SIO. Experimental Design: The correlation between pretransplant LIC and surrogate parameters as well as the impact of SIO on posttransplant outcome was assessed within an observational study of patients (n = 88) with either myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) undergoing allo-SCT. Results: Ferritin levels of 1,000 ng/mL or more provided only poor specificity (31.8%) for predicting elevated LIC (≥125 μmol/g) and even higher thresholds (≥2,500 ng/mL) lacked an association with nonrelapse mortality (NRM). In contrast, LIC 125 μmol/g or more was a significant risk factor for NRM in uni- and multivariate analysis (HR = 2.98; P = 0.016). Multivariate Cox-regression further showed that LIC 125 μmol/g or more was associated with a decreased overall survival (HR = 2.24, P = 0.038), whereas ferritin or transfusion burden were not. Conclusions: SIO reflected by LIC is an independent negative prognostic factor for posttransplant outcome in patients with AML and MDS undergoing allo-SCT. Therefore, MRI-based LIC, and not interference-prone serum markers such as ferritin, should be preferred for pretransplant risk stratification and patient selection in future clinical trials. Clin Cancer Res; 18(23); 6460–8. ©2012 AACR.

Graft-versus-Host disease Prophylaxis with Everolimus and Tacrolimus Is Associated with a High Incidence of Sinusoidal Obstruction Syndrome and Microangiopathy: Results of the EVTAC Trial

A calcineurin inhibitor combined with methotrexate is the standard prophylaxis for graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT). Everolimus, a derivative of sirolimus, seems to mediate antileukemia effects. We report on a combination of everolimus and tacrolimus in 24 patients (median age, 62 years) with either myelodysplastic syndrome (MDS; n = 17) or acute myeloid leukemia (AML; n = 7) undergoing intensive conditioning followed by HSCT from related (n = 4) or unrelated (n = 20) donors. All patients engrafted, and only 1 patient experienced grade IV mucositis. Nine patients (37%) developed acute grade II-IV GVHD, and 11 of 17 evaluable patients (64%) developed chronic extensive GVHD. Transplantation-associated microangiopathy (TMA) occurred in 7 patients (29%), with 2 cases of acute renal failure. The study was terminated prematurely because an additional 6 patients (25%) developed sinusoidal obstruction syndrome (SOS), which was fatal in 2 cases. With a median follow-up of 26 months, the 2-year overall survival rate was 47%. Although this new combination appears to be effective as a prophylactic regimen for acute GVHD, the incidence of TMA and SOS is considerably higher than seen with other regimens.

Allogeneic Stem-Cell Transplantation in Patients With NPM1-Mutated Acute Myeloid Leukemia: Results From a Prospective Donor Versus No-Donor Analysis of Patients After Upfront HLA Typing Within the SAL-AML 2003 Trial

PURPOSE The presence of a mutated nucleophosmin-1 gene (NPM1(mut)) in acute myeloid leukemia (AML) is associated with a favorable prognosis. To assess the predictive value with regard to allogeneic stem-cell transplantation (SCT), we compared the clinical course of patients with NPM1(mut) AML eligible for allogeneic SCT in a donor versus no-donor analysis. PATIENTS AND METHODS Of 1,179 patients with AML (age 18 to 60 years) treated in the Study Alliance Leukemia AML 2003 trial, we identified all NPM1(mut) patients with an intermediate-risk karyotype. According to the trial protocol, patients were intended to receive an allogeneic SCT if an HLA-identical sibling donor was available. Patients with no available donor received consolidation or autologous SCT. We compared relapse-free survival (RFS) and overall survival (OS) depending on the availability of a suitable donor. RESULTS Of 304 eligible patients, 77 patients had a sibling donor and 227 had no available matched family donor. The 3-year RFS rates in the donor and no-donor groups were 71% and 47%, respectively (P = .005); OS rates were 70% and 60%, respectively (P = .114). In patients with normal karyotype and no FLT3 internal tandem duplication (n = 148), the 3-year RFS rates in the donor and no-donor groups were 83% and 53%, respectively (P = .004); and the 3-year OS rates were 81% and 75%, respectively (P = .300). CONCLUSION Allogeneic SCT led to a significantly prolonged RFS in patients with NPM1(mut) AML. The absence of a statistically significant difference in OS is most likely a result of the fact that NPM1(mut) patients who experienced relapse responded well to salvage treatment. Allogeneic SCT in first remission has potent antileukemic efficacy and is a valuable treatment option in patients with NPM1(mut) AML with a sibling donor.

Unexpected recombinations in single chain bispecific anti-CD3-anti-CD33 antibodies can be avoided by a novel linker module.

CD33 is an attractive immunotarget on the surface of tumor cells from patients with acute myeloid leukemia (AML). In a first attempt for immunotargeting of AML blasts we constructed two bispecific antibodies in the single chain bispecific diabody (scBsDb) format by fusing the variable domains of monoclonal antibodies directed against CD3 and CD33. Unfortunately, protein expression of both scBsDbs resulted in varying mixtures of fragmented and full length proteins. As the non-functional fragments competed with the functional full length antibodies we tried to understand the reason for the fragmentation. We found that the anti-CD3 and anti-CD33 antibody genes show striking sequence homologies: during B cell development the same V(h) J558 heavy and V(l) kk4 light chain genes were selected. Moreover, the closely related D genes DSP2 (9 and 11) were combined with the same JH4 gene. And finally, during VJ recombination of the light chain the same JK5 element was selected. These homologies between the two monoclonal antibodies were the reason for recombinations in the cell lines generated for expression of the scBsDbs. Finally, we solved this problem by (i) rearranging the order of the heavy and light chains of the anti-CD3 and anti-CD33 domains, and (ii) a replacement of one of the commonly used glycine serine linkers with a novel linker domain. The resulting bispecific antibody in a single chain bispecific tandem format (scBsTaFv) was stable and capable of redirecting T cells to CD33-positive tumor cells including AML blasts of patients.