In this study, we compared immunoglobulin heavy-chain-gene-based minimal residual disease (MRD) detection by real-time quantitative PCR (RQ-PCR) and next-generation sequencing (NGS) to assess whether ...NGS could overcome some limitations of RQ-PCR and further increase sensitivity, specificity, accuracy and reproducibility. In total, 378 samples from 55 patients with acute lymphoblastic leukemia (ALL), mantle cell lymphoma (MCL) or multiple myeloma (MM) were investigated for clonotype identification, clonotype identity and comparability of MRD results. Forty-five clonotypes were identified by RQ-PCR and 49 by NGS. Clonotypes identified by both tools were identical or >97% homologous in 96% of cases. Both tools were able to routinely reach a sensitivity level of 1 × E-05. A good correlation of MRD results was observed (R=0.791, P<0.001), with excellent concordance in 79.6% of cases. Few discordant cases were observed across all disease subtypes. NGS showed at least the same level of sensitivity as allele-specific oligonucleotides-PCR, without the need for patient-specific reagents. We conclude that NGS is an effective tool for MRD monitoring in ALL, MCL and MM. Prospective comparative analysis of unselected cases is required to validate the clinical impact of NGS-based MRD assessment.
Minimal residual disease (MRD) after allogeneic stem cell transplantation (SCT) for Ph+ acute lymphoblastic leukemia (ALL) is predictive of relapse. Imatinib administration subsequent to SCT may ...prevent relapse, but the role of scheduling and its impact on outcome are not known. In a prospective, randomized multicenter trial, we compared the tolerability and efficacy of post-transplant imatinib administered either prophylactically (arm A; n=26) or following detection of MRD (arm B; n=29). Prophylactic imatinib significantly reduced the incidence of molecular recurrence after SCT compared with MRD-triggered imatinib (40% vs 69%; P=0.046). Median duration of PCR negativity was 26.5 and 6.8 months, respectively (P=0.065). Five-year survival in both interventional groups was high (80 and 74.5%), despite premature discontinuation of imatinib in the majority of patients because of poor tolerability. Relapse probability was significantly higher in patients who became MRD positive (P=0.017). In conclusion, post-transplant imatinib results in a low relapse rate, durable remissions and excellent long-term outcome in patients with BCR-ABL1-positive ALL irrespective of whether it is given prophylactically or MRD-triggered. Reappearance of BCR-ABL1 transcripts early after SCT or at higher levels identifies a small subset of patients who do not benefit sufficiently from imatinib, and in whom alternative approaches should be explored.
Assessment of minimal residual disease (MRD) has acquired a prominent position in European treatment protocols for patients with acute lymphoblastic leukemia (ALL), on the basis of its high ...prognostic value for predicting outcome and the possibilities for implementation of MRD diagnostics in treatment stratification. Therefore, there is an increasing need for standardization of methodologies and harmonization of terminology. For this purpose, a panel of representatives of all major European study groups on childhood and adult ALL and of international experts on PCR- and flow cytometry-based MRD assessment was built in the context of the Second International Symposium on MRD assessment in Kiel, Germany, 18-20 September 2008. The panel summarized the current state of MRD diagnostics in ALL and developed recommendations on the minimal technical requirements that should be fulfilled before implementation of MRD diagnostics into clinical trials. Finally, a common terminology for a standard description of MRD response and monitoring was established defining the terms 'complete MRD response', 'MRD persistence' and 'MRD reappearance'. The proposed MRD terminology may allow a refined and standardized assessment of response to treatment in adult and childhood ALL, and provides a sound basis for the comparison of MRD results between different treatment protocols.
We compared outcomes from a single-arm study of blinatumomab in adult patients with B-precursor Ph-negative relapsed/refractory acute lymphoblastic leukemia (R/R ALL) with a historical data set from ...Europe and the United States. Estimates of complete remission (CR) and overall survival (OS) were weighted by the frequency distribution of prognostic factors in the blinatumomab trial. Outcomes were also compared between the trial and historical data using propensity score methods. The historical cohort included 694 patients with CR data and 1112 patients with OS data compared with 189 patients with CR and survival data in the blinatumomab trial. The weighted analysis revealed a CR rate of 24% (95% CI: 20-27%) and a median OS of 3.3 months (95% CI: 2.8-3.6) in the historical cohort compared with a CR/CRh rate of 43% (95% CI: 36-50%) and a median OS of 6.1 months (95% CI: 4.2-7.5) in the blinatumomab trial. Propensity score analysis estimated increased odds of CR/CRh (OR=2.68, 95% CI: 1.67-4.31) and improved OS (HR=0.536, 95% CI: 0.394-0.730) with blinatumomab. The analysis demonstrates the application of different study designs and statistical methods to compare novel therapies for R/R ALL with historical data.
GATA3 is pivotal for the development of T lymphocytes. While its effects in later stages of T cell differentiation are well recognized, the role of GATA3 in the generation of early T cell precursors ...(ETP) has only recently been explored. As aberrant GATA3 mRNA expression has been linked to cancerogenesis, we investigated the role of GATA3 in early T cell precursor acute lymphoblastic leukemia (ETP-ALL).
We analyzed GATA3 mRNA expression by RT-PCR (n = 182) in adult patients with T-ALL. Of these, we identified 70 of 182 patients with ETP-ALL by immunophenotyping. DNA methylation was assessed genome wide (Illumina Infinium® HumanMethylation450 BeadChip platform) in 12 patients and GATA3-specifically by pyrosequencing in 70 patients with ETP-ALL. The mutational landscape of ETP-ALL with respect to GATA3 expression was investigated in 18 patients and validated by Sanger sequencing in 65 patients with ETP-ALL. Gene expression profiles (Affymetrix Human genome U133 Plus 2.0) of an independent cohort of adult T-ALL (n = 83) were used to identify ETP-ALL and investigate GATA3
and GATA3
expressing T-ALL patients. In addition, the ETP-ALL cell line PER-117 was investigated for cytotoxicity, apoptosis, GATA3 mRNA expression, DNA methylation, and global gene expression before and after treatment with decitabine.
In our cohort of 70 ETP-ALL patients, 33 % (23/70) lacked GATA3 expression and were thus defined as GATA3
. DNA methylation analysis revealed a high degree of GATA3 CpG island methylation in GATA3
compared with GATA3
ETP-ALL patients (mean 46 vs. 21 %, p < 0.0001). Genome-wide expression profiling of GATA3
ETP-ALL exhibited enrichment of myeloid/lymphoid progenitor (MLP) and granulocyte/monocyte progenitor (GMP) genes, while T cell-specific signatures were downregulated compared to GATA3
ETP-ALL. Among others, FLT3 expression was upregulated and mutational analyses demonstrated a high rate (79 %) of FLT3 mutations. Hypomethylating agents induced reversal of GATA3 silencing, and gene expression profiling revealed downregulation of hematopoietic stem cell genes and upregulation of T cell differentiation.
We propose GATA3
ETP-ALL as a novel stem cell-like leukemia with implications for the use of myeloid-derived therapies.
A subgroup of pediatric acute T-lymphoblastic leukemia (T-ALL) was characterized by a gene expression profile comparable to that of early T-cell precursors (ETPs) with a highly unfavorable outcome. ...We have investigated clinical and molecular characteristics of the ETP-ALL subgroup in adult T-ALL. As ETP-ALL represents a subgroup of early T-ALL we particularly focused on this cohort and identified 178 adult patients enrolled in the German Acute Lymphoblastic Leukemia Multicenter studies (05/93-07/03). Of these, 32% (57/178) were classified as ETP-ALL based on their characteristic immunophenotype. The outcome of adults with ETP-ALL was poor with an overall survival of only 35% at 10 years, comparable to the inferior outcome of early T-ALL with 38%. The molecular characterization of adult ETP-ALL revealed distinct alterations with overexpression of stem cell-related genes (BAALC, IGFBP7, MN1, WT1). Interestingly, we found a low rate of NOTCH1 mutations and no FBXW7 mutations in adult ETP-ALL. In contrast, FLT3 mutations, rare in the overall cohort of T-ALL, were very frequent and nearly exclusively found in ETP-ALL characterized by a specific immunophenotype. These molecular characteristics provide biologic insights and implications with respect to innovative treatment strategies (for example, tyrosine kinase inhibitors) for this high-risk subgroup of adult ETP-ALL.
TBI-based preparative regimens are considered as standard conditioning therapy for allogeneic stem cell transplantation (AHSC) in patients with ALL. We investigated toxicity and efficacy of a ...non-TBI-based regimen consisting of treosulfan, etoposide and cyclophosphamide for ALL within a prospective study. Major inclusion criteria were CR and non-eligibility for TBI. Fifty patients with a median age of 46.5 years (range, 18-64) were included. Donors were HLA-identical sibling (n=8), matched (n=42) or mismatched (n=10) unrelated. The toxicity was moderate, resulting in a cumulative incidence of non-relapse mortality (NRM) at 1 year of 8% (90% confidence interval: 2-15%). Acute GvHD grade II-IV and grade III/IV was noted in 53% and 14%, respectively. Chronic GvHD at one year was seen in 41%. After a median follow-up of 24 months the cumulative incidence of relapse was 36% (90% confidence interval: 24-48) and 51% (90% confidence interval: 37-65) at 1 and 2 years, respectively. The estimated 2-year disease-free and overall survivals were 36 and 48%, respectively. Treosulfan, etoposide and cyclophosphamide followed by AHSC has a favorable toxicity profile with low NRM and therefore represents a potential alternative regimen for ALL in 1. CR (NCT00682305).
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