Publications by authors named "Clara D Bloomfield"

240 Publications

Immunosuppression and outcomes in adult patients with de novo acute myeloid leukemia with normal karyotypes.

Proc Natl Acad Sci U S A 2021 12;118(49)

Division of Oncology, Department of Medicine, Washington University School of Medicine in St. Louis, St. Louis, MO 63110;

Acute myeloid leukemia (AML) patients rarely have long first remissions (LFRs; >5 y) after standard-of-care chemotherapy, unless classified as favorable risk at presentation. Identification of the mechanisms responsible for long vs. more typical, standard remissions may help to define prognostic determinants for chemotherapy responses. Using exome sequencing, RNA-sequencing, and functional immunologic studies, we characterized 28 normal karyotype (NK)-AML patients with >5 y first remissions after chemotherapy (LFRs) and compared them to a well-matched group of 31 NK-AML patients who relapsed within 2 y (standard first remissions [SFRs]). Our combined analyses indicated that genetic-risk profiling at presentation (as defined by European LeukemiaNet [ELN] 2017 criteria) was not sufficient to explain the outcomes of many SFR cases. Single-cell RNA-sequencing studies of 15 AML samples showed that SFR AML cells differentially expressed many genes associated with immune suppression. The bone marrow of SFR cases had significantly fewer CD4 Th1 cells; these T cells expressed an exhaustion signature and were resistant to activation by T cell receptor stimulation in the presence of autologous AML cells. T cell activation could be restored by removing the AML cells or blocking the inhibitory major histocompatibility complex class II receptor, LAG3. Most LFR cases did not display these features, suggesting that their AML cells were not as immunosuppressive. These findings were confirmed and extended in an independent set of 50 AML cases representing all ELN 2017 risk groups. AML cell-mediated suppression of CD4 T cell activation at presentation is strongly associated with unfavorable outcomes in AML patients treated with standard chemotherapy.
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http://dx.doi.org/10.1073/pnas.2116427118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8673586PMC
December 2021

The age of the bone marrow microenvironment influences B-cell acute lymphoblastic leukemia progression via CXCR5-CXCL13.

Blood 2021 11;138(19):1870-1884

Georg-Speyer-Haus, Institute for Tumor Biology and Experimental Therapy, Frankfurt am Main, Germany.

B-cell acute lymphoblastic leukemia (B-ALL) occurs most commonly in children, whereas chronic myeloid leukemia is more frequent in adults. The myeloid bias of hematopoiesis in elderly individuals has been considered causative, but the age of the bone marrow microenvironment (BMM) may be contributory. Using various murine models of B-ALL in young vs old mice, we recapitulated B-ALL preponderance in children vs adults. We showed differential effects of young vs old BM macrophages on B-ALL cell function. Molecular profiling using RNA- and ATAC-sequencing revealed pronounced differences in young vs old BMM-derived macrophages and enrichment for gene sets associated with inflammation. In concordance with the role of C-X-C motif chemokine (CXCL) 13 for disease-associated B-cell chemoattraction, we found CXCL13 to be highly expressed in young macrophages on a translational compared with a transcriptional level. Inhibition of CXCL13 in BM macrophages impaired leukemia cell migration and decreased the proliferation of cocultured B-ALL cells, whereas recombinant CXCL13 increased pAKT and B-ALL cell expansion. Pretreatment of B-ALL-initiating cells with CXCL13 accelerated B-ALL progression. Deficiency of Cxcr5, the receptor for CXCL13, on B-ALL-initiating cells prolonged murine survival, whereas high expression of CXCR5 in pediatric B-ALL may predict central nervous system relapse. CXCL13 staining was increased in bone sections from pediatric compared with adult patients with B-ALL. Taken together, our study shows that the age of the BMM and, in particular, BM macrophages influence the leukemia phenotype. The CXCR5-CXCL13 axis may act as prognostic marker and an attractive novel target for the treatment of B-ALL.
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http://dx.doi.org/10.1182/blood.2021011557DOI Listing
November 2021

Molecular landscape and prognostic impact of FLT3-ITD insertion site in acute myeloid leukemia: RATIFY study results.

Leukemia 2022 Jan 28;36(1):90-99. Epub 2021 Jul 28.

Department of Internal Medicine III, University Hospital of Ulm, Ulm, Germany.

In acute myeloid leukemia (AML) internal tandem duplications of the FLT3 gene (FLT3-ITD) are associated with poor prognosis. Retrospectively, we investigated the prognostic and predictive impact of FLT3-ITD insertion site (IS) in 452 patients randomized within the RATIFY trial, which evaluated midostaurin additionally to intensive chemotherapy. Next-generation sequencing identified 908 ITDs, with 643 IS in the juxtamembrane domain (JMD) and 265 IS in the tyrosine kinase domain-1 (TKD1). According to IS, patients were categorized as JMDsole (n = 251, 55%), JMD and TKD1 (JMD/TKD1; n = 117, 26%), and TKD1sole (n = 84, 19%). While clinical variables did not differ among the 3 groups, NPM1 mutation was correlated with JMDsole (P = 0.028). Overall survival (OS) differed significantly, with estimated 4-year OS probabilities of 0.44, 0.50, and 0.30 for JMDsole, JMD/TKD1, and TKD1sole, respectively (P = 0.032). Multivariate (cause-specific) Cox models for OS and cumulative incidence of relapse using allogeneic hematopoietic cell transplantation (HCT) in first complete remission as a time-dependent variable identified TKD1sole as unfavorable and HCT as favorable factors. In addition, Midostaurin exerted a significant benefit only for JMDsole. Our results confirm the distinct molecular heterogeneity of FLT3-ITD and the negative prognostic impact of TKD1 IS in AML that was not overcome by midostaurin.
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http://dx.doi.org/10.1038/s41375-021-01323-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8727286PMC
January 2022

Clinical and molecular relevance of genetic variants in the non-coding transcriptome of patients with cytogenetically normal acute myeloid leukemia.

Haematologica 2021 Jul 15. Epub 2021 Jul 15.

The Ohio State University, Comprehensive Cancer Center, Columbus.

Expression levels of long non-coding RNAs (lncRNAs) have been shown to associate with clinical outcome of patients with cytogenetically normal acute myeloid leukemia (CN-AML). However, the frequency and clinical significance of genetic variants in the nucleotide sequences of lncRNAs in AML patients is unknown. Herein, we analyzed total RNA sequencing data of 377 younger adults (aged.
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http://dx.doi.org/10.3324/haematol.2021.266643DOI Listing
July 2021

Phase 3 randomized trial of chemotherapy with or without oblimersen in older AML patients: CALGB 10201 (Alliance).

Blood Adv 2021 07;5(13):2775-2787

University of Chicago Comprehensive Cancer Center, Chicago, IL.

Overexpression of B-cell leukemia/lymphoma 2 (BCL2) renders acute myeloid leukemia (AML) cells resistant to chemotherapy and has been associated with unfavorable outcomes. Oblimersen (G3139) is a phosphorothioate 18-mer antisense oligonucleotide directed against the first 6 BCL2 codons. In a phase 1 study of AML patients treated with G3139, cytarabine, and daunorubicin induction with cytarabine consolidation, no antisense-related toxicity was reported, and BCL2 downregulation occurred in patients achieving complete remission. In this phase 3 trial, untreated older AML patients were randomized to cytarabine (100 mg/m2 per day on days 4-10) and daunorubicin (60 mg/m2 per day on days 4-6) followed by cytarabine consolidation (2000 mg/m2 per day on days 4-8) with (arm A) or without (arm B) G3139 (7 mg/m2 per day on days 1-10 [induction] or days 1-8 [consolidation]). A total of 506 patients were enrolled. No differences in toxicity were observed between arms. Estimated overall survival (OS) at 1 year was 43% for arm A and 40% for arm B (1-sided log rank P = .13), with no differences in disease-free (DFS; P = .26) or event-free survival (P = .80). Subgroup analyses showed patients age <70 years in arm A had improved OS by 1 month vs those in arm B (P = .04), and patients with secondary AML in arm A had better DFS vs those in arm B (P = .04). We conclude that addition of G3139 to chemotherapy failed to improve outcomes of older AML patients. However, more effective means of inhibiting BCL2 are showing promising results in combination with chemotherapy in AML. This trial was registered at www.clinicaltrials.gov as #NCT00085124.
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http://dx.doi.org/10.1182/bloodadvances.2021004233DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8288671PMC
July 2021

Precision oncology in AML: validation of the prognostic value of the knowledge bank approach and suggestions for improvement.

J Hematol Oncol 2021 07 6;14(1):107. Epub 2021 Jul 6.

The Ohio State University Comprehensive Cancer Center, 460 West 12th Avenue, Columbus, OH, 43210-1228, USA.

Recently, a novel knowledge bank (KB) approach to predict outcomes of individual patients with acute myeloid leukemia (AML) was developed using unbiased machine learning. To validate its prognostic value, we analyzed 1612 adults with de novo AML treated on Cancer and Leukemia Group B front-line trials who had pretreatment clinical, cytogenetics, and mutation data on 81 leukemia/cancer-associated genes available. We used receiver operating characteristic (ROC) curves and the area under the curve (AUC) to evaluate the predictive values of the KB algorithm and other risk classifications. The KB algorithm predicted 3-year overall survival (OS) probability in the entire patient cohort (AUC = 0.799), and both younger (< 60 years) (AUC = 0.747) and older patients (AUC = 0.770). The KB algorithm predicted non-remission death (AUC = 0.860) well but was less accurate in predicting relapse death (AUC = 0.695) and death in first complete remission (AUC = 0.603). The KB algorithm's 3-year OS predictive value was higher than that of the 2017 European LeukemiaNet (ELN) classification (AUC = 0.707, p < 0.001) and 2010 ELN classification (AUC = 0.721, p < 0.001) but did not differ significantly from that of the 17-gene stemness score (AUC = 0.732, p = 0.10). Analysis of additional cytogenetic and molecular markers not included in the KB algorithm revealed that taking into account atypical complex karyotype, infrequent recurrent balanced chromosome rearrangements and mutational status of the SAMHD1, AXL and NOTCH1 genes may improve the KB algorithm. We conclude that the KB algorithm has a high predictive value that is higher than those of the 2017 and 2010 ELN classifications. Inclusion of additional genetic features might refine the KB algorithm.
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http://dx.doi.org/10.1186/s13045-021-01118-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8261916PMC
July 2021

A precision medicine classification for treatment of acute myeloid leukemia in older patients.

J Hematol Oncol 2021 06 23;14(1):96. Epub 2021 Jun 23.

The Ohio State University Comprehensive Cancer Center, 320 West 10th Avenue, Starling Loving Hall B302, Columbus, OH, 43210, USA.

Background: Older patients (≥ 60 years) with acute myeloid leukemia (AML) often have multiple, sequentially acquired, somatic mutations that drive leukemogenesis and are associated with poor outcome. Beat AML is a Leukemia and Lymphoma Society-sponsored, multicenter umbrella study that algorithmically segregates AML patients based upon cytogenetic and dominant molecular abnormalities (variant allele frequencies (VAF) ≥ 0.2) into different cohorts to select for targeted therapies. During the conception of the Beat AML design, a historical dataset was needed to help in the design of the genomic algorithm for patient assignment and serve as the basis for the statistical design of individual genomic treatment substudies for the Beat AML study.

Methods: We classified 563 newly diagnosed older AML patients treated with standard intensive chemotherapy on trials conducted by Cancer and Leukemia Group B based on the same genomic algorithm and assessed clinical outcomes.

Results: Our classification identified core-binding factor and NPM1-mutated/FLT3-ITD-negative groups as having the best outcomes, with 30-day early death (ED) rates of 0 and 20%, respectively, and median overall survival (OS) of > 1 year and 3-year OS rates of ≥ 20%. All other genomic groups had ED rates of 17-42%, median OS ≤ 1 year and 3-year OS rates of ≤ 15%.

Conclusions: By classifying patients through this genomic algorithm, outcomes were poor and not unexpected from a non-algorithmic, non-dominant VAF approach. The exception is 30-day ED rate typically is not available for intensive induction for individual genomic groups and therefore difficult to compare outcomes with targeted therapeutics. This Alliance data supported the use of this algorithm for patient assignment at the initiation of the Beat AML study. This outcome data was also used for statistical design for Beat AML substudies for individual genomic groups to determine goals for improvement from intensive induction and hopefully lead to more rapid approval of new therapies. Trial registration ClinicalTrials.gov Identifiers: NCT00048958 (CALGB 8461), NCT00900224 (CALGB 20202), NCT00003190 (CALGB 9720), NCT00085124 (CALGB 10201), NCT00742625 (CALGB 10502), NCT01420926 (CALGB 11002), NCT00039377 (CALGB 10801), and NCT01253070 (CALGB 11001).
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http://dx.doi.org/10.1186/s13045-021-01110-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8220739PMC
June 2021

Genomic analysis of cellular hierarchy in acute myeloid leukemia using ultrasensitive LC-FACSeq.

Leukemia 2021 12 21;35(12):3406-3420. Epub 2021 May 21.

Division of Hematology, The Ohio State University Comprehensive Cancer Center, Columbus, OH, USA.

Hematopoiesis is hierarchical, and it has been postulated that acute myeloid leukemia (AML) is organized similarly with leukemia stem cells (LSCs) residing at the apex. Limited cells acquired by fluorescence activated cell sorting in tandem with targeted amplicon-based sequencing (LC-FACSeq) enables identification of mutations in small subpopulations of cells, such as LSCs. Leveraging this, we studied clonal compositions of immunophenotypically-defined compartments in AML through genomic and functional analyses at diagnosis, remission and relapse in 88 AML patients. Mutations involving DNA methylation pathways, transcription factors and spliceosomal machinery did not differ across compartments, while signaling pathway mutations were less frequent in putative LSCs. We also provide insights into TP53-mutated AML by demonstrating stepwise acquisition of mutations beginning from the preleukemic hematopoietic stem cell stage. In 10 analyzed cases, acquisition of additional mutations and del(17p) led to genetic and functional heterogeneity within the LSC pool with subclones harboring varying degrees of clonogenic potential. Finally, we use LC-FACSeq to track clonal evolution in serial samples, which can also be a powerful tool to direct targeted therapy against measurable residual disease. Therefore, studying clinically significant small subpopulations of cells can improve our understanding of AML biology and offers advantages over bulk sequencing to monitor the evolution of disease.
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http://dx.doi.org/10.1038/s41375-021-01295-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8606012PMC
December 2021

Secondary cytogenetic abnormalities in core-binding factor AML harboring inv(16) vs t(8;21).

Blood Adv 2021 05;5(10):2481-2489

Adult Bone Marrow Transplant Service, Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY.

Patients with core-binding factor (CBF) acute myeloid leukemia (AML), caused by either t(8;21)(q22;q22) or inv(16)(p13q22)/t(16;16)(p13;q22), have higher complete remission rates and longer survival than patients with other subtypes of AML. However, ∼40% of patients relapse, and the literature suggests that patients with inv(16) fare differently from those with t(8;21). We retrospectively analyzed 537 patients with CBF-AML, focusing on additional cytogenetic aberrations to examine their impact on clinical outcomes. Trisomies of chromosomes 8, 21, or 22 were significantly more common in patients with inv(16)/t(16;16): 16% vs 7%, 6% vs 0%, and 17% vs 0%, respectively. In contrast, del(9q) and loss of a sex chromosome were more frequent in patients with t(8;21): 15% vs 0.4% for del(9q), 37% vs 0% for loss of X in females, and 44% vs 5% for loss of Y in males. Hyperdiploidy was more frequent in patients with inv(16) (25% vs 9%, whereas hypodiploidy was more frequent in patients with t(8;21) (37% vs 3%. In multivariable analyses (adjusted for age, white blood counts at diagnosis, and KIT mutation status), trisomy 8 was associated with improved overall survival (OS) in inv(16), whereas the presence of other chromosomal abnormalities (not trisomy 8) was associated with decreased OS. In patients with t(8;21), hypodiploidy was associated with improved disease-free survival; hyperdiploidy and del(9q) were associated with improved OS. KIT mutation (either positive or not tested, compared with negative) conferred poor prognoses in univariate analysis only in patients with t(8;21).
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http://dx.doi.org/10.1182/bloodadvances.2020003605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8152510PMC
May 2021

DNA methylation epitypes highlight underlying developmental and disease pathways in acute myeloid leukemia.

Genome Res 2021 May 11;31(5):747-761. Epub 2021 Mar 11.

Division of Hematology, Department of Internal Medicine, The Ohio State University, Columbus, Ohio 43210, USA.

Acute myeloid leukemia (AML) is a molecularly complex disease characterized by heterogeneous tumor genetic profiles and involving numerous pathogenic mechanisms and pathways. Integration of molecular data types across multiple patient cohorts may advance current genetic approaches for improved subclassification and understanding of the biology of the disease. Here, we analyzed genome-wide DNA methylation in 649 AML patients using Illumina arrays and identified a configuration of 13 subtypes (termed "epitypes") using unbiased clustering. Integration of genetic data revealed that most epitypes were associated with a certain recurrent mutation (or combination) in a majority of patients, yet other epitypes were largely independent. Epitypes showed developmental blockage at discrete stages of myeloid differentiation, revealing epitypes that retain arrested hematopoietic stem-cell-like phenotypes. Detailed analyses of DNA methylation patterns identified unique patterns of aberrant hyper- and hypomethylation among epitypes, with variable involvement of transcription factors influencing promoter, enhancer, and repressed regions. Patients in epitypes with stem-cell-like methylation features showed inferior overall survival along with up-regulated stem cell gene expression signatures. We further identified a DNA methylation signature involving STAT motifs associated with -ITD mutations. Finally, DNA methylation signatures were stable at relapse for the large majority of patients, and rare epitype switching accompanied loss of the dominant epitype mutations and reversion to stem-cell-like methylation patterns. These results show that DNA methylation-based classification integrates important molecular features of AML to reveal the diverse pathogenic and biological aspects of the disease.
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http://dx.doi.org/10.1101/gr.269233.120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092005PMC
May 2021

Gene expression signature predicts relapse in adult patients with cytogenetically normal acute myeloid leukemia.

Blood Adv 2021 03;5(5):1474-1482

The Ohio State University Comprehensive Cancer Center, Columbus OH.

Although ∼80% of adult patients with cytogenetically normal acute myeloid leukemia (CN-AML) achieve a complete remission (CR), more than half of them relapse. Better identification of patients who are likely to relapse can help to inform clinical decisions. We performed RNA sequencing on pretreatment samples from 268 adults with de novo CN-AML who were younger than 60 years of age and achieved a CR after induction treatment with standard "7+3" chemotherapy. After filtering for genes whose expressions were associated with gene mutations known to impact outcome (ie, CEBPA, NPM1, and FLT3-internal tandem duplication [FLT3-ITD]), we identified a 10-gene signature that was strongly predictive of patient relapse (area under the receiver operating characteristics curve [AUC], 0.81). The signature consisted of 7 coding genes (GAS6, PSD3, PLCB4, DEXI, JMY, NRP1, C10orf55) and 3 long noncoding RNAs. In multivariable analysis, the 10-gene signature was strongly associated with relapse (P < .001), after adjustment for the FLT3-ITD, CEBPA, and NPM1 mutational status. Validation of the expression signature in an independent patient set from The Cancer Genome Atlas showed the signature's strong predictive value, with AUC = 0.78. Implementation of the 10-gene signature into clinical prognostic stratification could be useful for identifying patients who are likely to relapse.
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http://dx.doi.org/10.1182/bloodadvances.2020003727DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7948288PMC
March 2021

Midostaurin reduces relapse in FLT3-mutant acute myeloid leukemia: the Alliance CALGB 10603/RATIFY trial.

Leukemia 2021 09 2;35(9):2539-2551. Epub 2021 Mar 2.

Novartis Pharma AG, Basel, Switzerland.

The prospective randomized, placebo-controlled CALGB 10603/RATIFY trial (Alliance) demonstrated a statistically significant overall survival benefit from the addition of midostaurin to standard frontline chemotherapy in a genotypically-defined subgroup of 717 patients with FLT3-mutant acute myeloid leukemia (AML). The risk of death was reduced by 22% on the midostaurin-containing arm. In this post hoc analysis, we analyzed the cumulative incidence of relapse (CIR) on this study and also evaluated the impact of 12 4-week cycles of maintenance therapy. CIR analyses treated relapses and AML deaths as events, deaths from other causes as competing risks, and survivors in remission were censored. CIR was improved on the midostaurin arm (HR = 0.71 (95% CI, 0.54-0.93); p = 0.01), both overall and within European LeukemiaNet 2017 risk classification subsets when post-transplant events were considered in the analysis as events. However, when transplantation was considered as a competing risk, there was overall no significant difference between the risks of relapse on the two randomized arms. Patients still in remission after consolidation with high-dose cytarabine entered the maintenance phase, continuing with either midostaurin or placebo. Analyses were inconclusive in quantifying the impact of the maintenance phase on the overall outcome. In summary, midostaurin reduces the CIR.
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http://dx.doi.org/10.1038/s41375-021-01179-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8591906PMC
September 2021

Clonal evolution of acute myeloid leukemia with FLT3-ITD mutation under treatment with midostaurin.

Blood 2021 06;137(22):3093-3104

Berlin Institute of Health at Charité-Universitätsmedizin Berlin, Germany.

In the international randomized phase 3 RATIFY (Randomized AML Trial In FLT3 in patients less than 60 Years old) trial, the multikinase inhibitor midostaurin significantly improved overall and event-free survival in patients 18 to 59 years of age with FLT3-mutated acute myeloid leukemia (AML). However, only 59% of patients in the midostaurin arm achieved protocol-specified complete remission (CR), and almost half of patients achieving CR relapsed. To explore underlying mechanisms of resistance, we studied patterns of clonal evolution in patients with FLT3-internal tandem duplications (ITD)-positive AML who were entered in the RATIFY or German-Austrian Acute Myeloid Leukemia Study Group 16-10 trial and received treatment with midostaurin. To this end, paired samples from 54 patients obtained at time of diagnosis and at time of either relapsed or refractory disease were analyzed using conventional Genescan-based testing for FLT3-ITD and whole exome sequencing. At the time of disease resistance or progression, almost half of the patients (46%) became FLT3-ITD negative but acquired mutations in signaling pathways (eg, MAPK), thereby providing a new proliferative advantage. In cases with FLT3-ITD persistence, the selection of resistant ITD clones was found in 11% as potential drivers of disease. In 32% of cases, no FLT3-ITD mutational change was observed, suggesting either resistance mechanisms bypassing FLT3 inhibition or loss of midostaurin inhibitory activity because of inadequate drug levels. In summary, our study provides novel insights into the clonal evolution and resistance mechanisms of FLT3-ITD-mutated AML under treatment with midostaurin in combination with intensive chemotherapy.
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http://dx.doi.org/10.1182/blood.2020007626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8233666PMC
June 2021

Midostaurin in patients with acute myeloid leukemia and FLT3-TKD mutations: a subanalysis from the RATIFY trial.

Blood Adv 2020 10;4(19):4945-4954

Medizinische Klinik und Poliklinik I, Universitätsklinikum Carl Gustav Carus der TU Dresden, Dresden, Germany; and.

The results from the RATIFY trial (ClinicalTrials.gov: NCT00651261; CALGB 10603) showed that midostaurin combined with standard chemotherapy significantly improved outcomes in patients with FMS-like tyrosine kinase 3 (FLT3)-mutated acute myeloid leukemia (AML), compared with placebo. In this post hoc subgroup analysis from the trial, we evaluated the impact of midostaurin in 163 patients with FLT3-tyrosine kinase domain (TKD) mutations. At a median follow-up of 60.7 months (95% CI, 55.0-70.8), the 5-year event-free survival (EFS) rate was significantly higher in patients treated with midostaurin than in those treated with placebo (45.2% vs 30.1%; P = .044). A trend toward improved disease-free survival was also observed with midostaurin (67.3% vs 53.4%; P = .089), whereas overall survival (OS) was similar in the 2 groups. Patients with AML and NPM1mut/FLT3-TKDmut or core binding factor (CBF)-rearranged/FLT3-TKDmut genotypes had significantly prolonged OS with or without censoring at hematopoietic cell transplantation (HCT), compared with NPM1WT/CBF-negative AMLs. The multivariable model for OS and EFS adjusted for allogeneic HCT in first complete remission as a time-dependent covariable, revealed NPM1 mutations and CBF rearrangements as significant favorable factors. These data show that NPM1 mutations or CBF rearrangements identify favorable prognostic groups in patients with FLT3-TKD AMLs, independent of other factors, also in the context of midostaurin treatment.
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http://dx.doi.org/10.1182/bloodadvances.2020002904DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7556122PMC
October 2020

Mutational landscape and clinical outcome of patients with de novo acute myeloid leukemia and rearrangements involving 11q23/.

Proc Natl Acad Sci U S A 2020 10 5;117(42):26340-26346. Epub 2020 Oct 5.

The Ohio State University Comprehensive Cancer Center, Columbus, OH 43210.

Balanced rearrangements involving the gene, located at 11q23, are among the most frequent chromosome aberrations in acute myeloid leukemia (AML). Because of numerous fusion partners, the mutational landscape and prognostic impact of specific 11q23/ rearrangements are not fully understood. We analyzed clinical features of 172 adults with AML and recurrent 11q23/ rearrangements, 141 of whom had outcome data available. We compared outcomes of these patients with outcomes of 1,097 patients without an 11q23/ rearrangement categorized according to the 2017 European LeukemiaNet (ELN) classification. Using targeted next-generation sequencing, we investigated the mutational status of 81 leukemia/cancer-associated genes in 96 patients with 11q23/ rearrangements with material for molecular studies available. Patients with 11q23/ rearrangements had a low number of additional gene mutations (median, 1; range 0 to 6), which involved the RAS pathway (, , and ) in 32% of patients. mutations occurred more often in patients with t(6;11)(q27;q23)/- compared with patients with the other 11q23/ subsets. Specific gene mutations were too infrequent in patients with specific 11q23/ rearrangements to assess their associations with outcomes. We demonstrate that younger (age <60 y) patients with t(9;11)(p22;q23)/- had better outcomes than patients with other 11q23/ rearrangements and those without 11q23/ rearrangements classified in the 2017 ELN intermediate-risk group. Conversely, outcomes of older patients (age ≥60 y) with t(9;11)(p22;q23) were poor and comparable to those of the ELN adverse-risk group patients. Our study shows that patients with an 11q23/ rearrangement have distinct mutational patterns and outcomes depending on the fusion partner.
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http://dx.doi.org/10.1073/pnas.2014732117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7584992PMC
October 2020

Additional gene mutations may refine the 2017 European LeukemiaNet classification in adult patients with de novo acute myeloid leukemia aged <60 years.

Leukemia 2020 12 27;34(12):3215-3227. Epub 2020 May 27.

The Ohio State University Comprehensive Cancer Center, Columbus, OH, USA.

The European LeukemiaNet (ELN) recommendations for diagnosis and management of acute myeloid leukemia (AML) have become an important tool to assess patients' prognosis and guide treatment. We tested the prognostic impact of the 2017 ELN classification in a large cohort of 863 AML patients aged <60 years similarly treated on Cancer and Leukemia Group B/Alliance for Clinical Trials in Oncology studies. Based on multivariable models within each ELN genetic-risk group, we identified additional gene mutations that may refine the 2017 ELN risk classification. BCOR- or SETBP1-mutated favorable-risk patients with non-core-binding factor AML and IDH-mutated adverse-risk patients had intermediate-risk outcomes. Outcomes of NPM1/WT1 co-mutated patients and those of ZRSR2-mutated patients resembled outcome of adverse-risk patients. Moreover, FLT3-ITD allelic ratio conferred adverse rather than intermediate-risk irrespective of the NPM1 mutation status, and DNMT3A mutations associated with very poor survival. Application of these refinements reclassified 9% of current favorable-risk patients and 53% of current intermediate-risk patients to the adverse-risk group, with similar poor survival as current adverse-risk patients. Furthermore, 4% of current favorable-risk patients and 9% of adverse-risk patients were reclassified to the intermediate-risk group.
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http://dx.doi.org/10.1038/s41375-020-0872-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7882079PMC
December 2020

Combination of dasatinib with chemotherapy in previously untreated core binding factor acute myeloid leukemia: CALGB 10801.

Blood Adv 2020 02;4(4):696-705

Section of Hematology/Oncology, Department of Medicine, University of Chicago, Chicago, IL.

Acute myeloid leukemia (AML) with either t(8;21)(q22;q22) or inv(16)(p13q22)/t(16;16)(p13;q22) is referred to as core binding factor (CBF) AML. Although categorized as favorable risk, long-term survival for these patients is only ∼50% to 60%. Mutated (mut) or overexpressed KIT, a gene encoding a receptor tyrosine kinase, has been found almost exclusively in CBF AML and may increase the risk of disease relapse. We tested the safety and clinical activity of dasatinib, a multi-kinase inhibitor, in combination with chemotherapy. Sixty-one adult patients with AML and CBF fusion transcripts (RUNX1/RUNX1T1 or CBFB/MYH11) were enrolled on Cancer and Leukemia Group B (CALGB) 10801. Patients received cytarabine/daunorubicin induction on days 1 to 7 and oral dasatinib 100 mg/d on days 8 to 21. Upon achieving complete remission, patients received consolidation with high-dose cytarabine followed by dasatinib 100 mg/d on days 6 to 26 for 4 courses, followed by dasatinib 100 mg/d for 12 months. Fifteen (25%) patients were older (aged ≥60 years); 67% were CBFB/MYH11-positive, and 19% harbored KITmut. There were no unexpected or dose-limiting toxicities. Fifty-five (90%) patients achieved complete remission. With a median follow-up of 45 months, only 16% have relapsed. The 3-year disease-free survival and overall survival rates were 75% and 77% (79% and 85% for younger patients [aged <60 years], and 60% and 51% for older patients). Patients with KITmut had comparable outcome to those with wild-type KIT (3-year rates: disease-free survival, 67% vs 75%; overall survival, 73% vs 76%), thereby raising the question of whether dasatinib may overcome the negative impact of these genetic lesions. CALGB 10801 was registered at www.clinicaltrials.gov as #NCT01238211.
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http://dx.doi.org/10.1182/bloodadvances.2019000492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7042984PMC
February 2020

Clinical and functional significance of circular RNAs in cytogenetically normal AML.

Blood Adv 2020 01;4(2):239-251

The Ohio State University Comprehensive Cancer Center, Columbus, OH.

Circular RNAs (circRNAs) are noncoding RNA molecules that display a perturbed arrangement of exons, called backsplicing. To examine the prognostic and biologic significance of circRNA expression in cytogenetically normal acute myeloid leukemia (CN-AML), we conducted whole-transcriptome profiling in 365 younger adults (age 18-60 years) with CN-AML. We applied a novel pipeline, called Massive Scan for circRNA, to identify and quantify circRNA expression. We validated the high sensitivity and specificity of our pipeline by performing RNase R treatment and RNA sequencing in samples of AML patients and cell lines. Unsupervised clustering analyses identified 3 distinct circRNA expression-based clusters with different frequencies of clinical and molecular features. After dividing our cohort into training and validation data sets, we identified 4 circRNAs (circCFLAR, circKLHL8, circSMC1A, and circFCHO2) that were prognostic in both data sets; high expression of each prognostic circRNA was associated with longer disease-free, overall, and event-free survival. In multivariable analyses, high circKLHL8 and high circFCHO2 expression were independently associated with better clinical outcome of CN-AML patients, after adjusting for other covariates. To examine the biologic relevance of circRNA expression, we performed knockdown screening experiments in a subset of prognostic and gene mutation-related candidate circRNAs. We identified circFBXW7, but not its linear messenger RNA, as a regulator of the proliferative capacity of AML blasts. In summary, our findings underscore the molecular associations, prognostic significance, and functional relevance of circRNA expression in CN-AML.
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http://dx.doi.org/10.1182/bloodadvances.2019000568DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6988408PMC
January 2020

Impact of NPM1/FLT3-ITD genotypes defined by the 2017 European LeukemiaNet in patients with acute myeloid leukemia.

Blood 2020 01;135(5):371-380

Department of Internal Medicine I, University Hospital Carl Gustav Carus, Dresden, Germany.

Patients with acute myeloid leukemia (AML) harboring FLT3 internal tandem duplications (ITDs) have poor outcomes, in particular AML with a high (≥0.5) mutant/wild-type allelic ratio (AR). The 2017 European LeukemiaNet (ELN) recommendations defined 4 distinct FLT3-ITD genotypes based on the ITD AR and the NPM1 mutational status. In this retrospective exploratory study, we investigated the prognostic and predictive impact of the NPM1/FLT3-ITD genotypes categorized according to the 2017 ELN risk groups in patients randomized within the RATIFY trial, which evaluated the addition of midostaurin to standard chemotherapy. The 4 NPM1/FLT3-ITD genotypes differed significantly with regard to clinical and concurrent genetic features. Complete ELN risk categorization could be done in 318 of 549 trial patients with FLT3-ITD AML. Significant factors for response after 1 or 2 induction cycles were ELN risk group and white blood cell (WBC) counts; treatment with midostaurin had no influence. Overall survival (OS) differed significantly among ELN risk groups, with estimated 5-year OS probabilities of 0.63, 0.43, and 0.33 for favorable-, intermediate-, and adverse-risk groups, respectively (P < .001). A multivariate Cox model for OS using allogeneic hematopoietic cell transplantation (HCT) in first complete remission as a time-dependent variable revealed treatment with midostaurin, allogeneic HCT, ELN favorable-risk group, and lower WBC counts as significant favorable factors. In this model, there was a consistent beneficial effect of midostaurin across ELN risk groups.
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http://dx.doi.org/10.1182/blood.2019002697DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6993016PMC
January 2020

The long non-coding RNA HOXB-AS3 regulates ribosomal RNA transcription in NPM1-mutated acute myeloid leukemia.

Nat Commun 2019 11 25;10(1):5351. Epub 2019 Nov 25.

Center for RNA Medicine, Department of Clinical Medicine, Aalborg University, Copenhagen, Denmark.

Long non-coding RNAs (lncRNAs) are important regulatory molecules that are implicated in cellular physiology and pathology. In this work, we dissect the functional role of the HOXB-AS3 lncRNA in patients with NPM1-mutated (NPM1mut) acute myeloid leukemia (AML). We show that HOXB-AS3 regulates the proliferative capacity of NPM1mut AML blasts in vitro and in vivo. HOXB-AS3 is shown to interact with the ErbB3-binding protein 1 (EBP1) and guide EBP1 to the ribosomal DNA locus. Via this mechanism, HOXB-AS3 regulates ribosomal RNA transcription and de novo protein synthesis. We propose that in the context of NPM1 mutations, HOXB-AS3 overexpression acts as a compensatory mechanism, which allows adequate protein production in leukemic blasts.
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http://dx.doi.org/10.1038/s41467-019-13259-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6877618PMC
November 2019

EGFL7 Antagonizes NOTCH Signaling and Represents a Novel Therapeutic Target in Acute Myeloid Leukemia.

Clin Cancer Res 2020 02 31;26(3):669-678. Epub 2019 Oct 31.

The Ohio State University Comprehensive Cancer Center, Columbus, Ohio.

Purpose: EGF-like domain 7 (EGFL7) is a secreted protein and recently has been shown to play an important role in acute myeloid leukemia (AML); however, the underlying mechanism by which EGFL7 promotes leukemogenesis is largely unknown.

Experimental Design: Using an antibody interaction array, we measured the ability of EGFL7 to bind directly approximately 400 proteins expressed by primary AML blasts. Primary patient samples were stimulated with recombinant EGFL7 (rEGFL7) or anti-EGFL7 blocking antibody to assess alterations in downstream signaling and the ability to effect blast differentiation and survival. We treated three independent AML models with anti-EGFL7 or IgG1 control to determine whether anti-EGFL7 could prolong survival .

Results: We found EGFL7 significantly binds several signaling proteins important for normal and malignant hematopoiesis including NOTCH. Stimulation of AML blasts with rEGFL7 reduced NOTCH intracellular domain and NOTCH target gene expression while treatment with an anti-EGFL7 blocking antibody resulted in reactivation of NOTCH signaling, increased differentiation, and apoptosis. Competitive ligand-binding assays showed rEGFL7 inhibits DELTA-like (DLL) 4-mediated NOTCH activation while anti-EGFL7 combined with DLL4 significantly increased NOTCH activation and induced apoptosis. Using three different AML mouse models, we demonstrated that treatment with anti-EGFL7 alone results in increased survival.

Conclusions: Our data demonstrate that EGFL7 contributes to NOTCH silencing in AML by antagonizing canonical NOTCH ligand binding. Reactivation of NOTCH signaling using anti-EGFL7 results in prolonged survival of leukemic mice, supporting the use of EGFL7 as a novel therapeutic target in AML.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-2479DOI Listing
February 2020

Clinical and molecular characterization of patients with acute myeloid leukemia and sole trisomies of chromosomes 4, 8, 11, 13 or 21.

Leukemia 2020 02 28;34(2):358-368. Epub 2019 Aug 28.

Division of Hematology, Department of Internal Medicine, The Ohio State University, Columbus, OH, USA.

Sole trisomies of chromosomes 4, 8, 11, 13 and 21 account for 89-95% of all sole trisomies in adult AML patients. We analyzed clinical and molecular characteristics of 138 de novo AML patients with sole +4, +8, +11, +13 or +21, and compared them with AML patients with those trisomies occurring in addition to other chromosome abnormalities (non-sole trisomy) and with cytogenetically normal AML (CN-AML) patients. Mutations in methylation-related genes were most commonly observed within each sole trisomy group (+4, 55%; +8, 58%; +11, 71%; +13, 71%; +21, 75% of patients). Patients with sole trisomies, excluding +4, also had frequent mutations in spliceosome genes (+8, 43%; +11, 65%; +13, 65%; +21, 45% of patients). In contrast, +4 patients frequently had mutations in transcription factor genes (44%) and NPM1 (36%). While 48% of patients with sole trisomies harbored mutations in a spliceosome gene, spliceosome mutations were observed in only 24% of non-sole trisomy (n = 131, P < 0.001) and 19% of CN-AML patients (n = 716, P < 0.001). Our data suggest that mutations affecting methylation-related genes are a molecular hallmark of sole trisomies. Mutations in spliceosome genes were also commonly observed in many sole trisomy patients and represent a novel finding in this cytogenetic subgroup.
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http://dx.doi.org/10.1038/s41375-019-0560-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6995758PMC
February 2020

Mutations associated with a 17-gene leukemia stem cell score and the score's prognostic relevance in the context of the European LeukemiaNet classification of acute myeloid leukemia.

Haematologica 2020 03 14;105(3):721-729. Epub 2019 Aug 14.

The Ohio State University Comprehensive Cancer Center, Columbus, OH

Leukemia stem cells (LSC) are more resistant to standard chemotherapy and their persistence during remission can cause relapse, which is still one of the major clinical challenges in the treatment of acute myeloid leukemia (AML). A better understanding of the mutational patterns and the prognostic impact of molecular markers associated with stemness could lead to better clinical management and improve patients' outcomes. We applied a previously described 17-gene expression score comprising genes differently expressed between LSC and leukemic bulk blasts, for 934 adult patients with AML, and studied associations of the 17-gene LSC score with clinical data and mutation status of 81 genes recurrently mutated in cancer and leukemia. We found that patients with a high 17-gene score were older and had more mutations. The 17-gene score was found to have a prognostic impact in both younger (aged <60 years) and older (aged ≥60 years) patients with AML. We also analyzed the 17-gene LSC score in the context of the 2017 European LeukemiaNet genetic-risk classification and found that for younger patients the score refined the classification, and identified patients currently classified in the European LeukemiaNet Favorable-risk category who had a worse outcome.
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http://dx.doi.org/10.3324/haematol.2019.225003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7049376PMC
March 2020

Genetic Characterization and Prognostic Relevance of Acquired Uniparental Disomies in Cytogenetically Normal Acute Myeloid Leukemia.

Clin Cancer Res 2019 11 2;25(21):6524-6531. Epub 2019 Aug 2.

The Ohio State University Comprehensive Cancer Center, Columbus, Ohio.

Purpose: Uniparental disomy (UPD) is a way cancer cells duplicate a mutated gene, causing loss of heterozygosity (LOH). Patients with cytogenetically normal acute myeloid leukemia (CN-AML) do not have microscopically detectable chromosome abnormalities, but can harbor UPDs. We examined the prognostic significance of UPDs and frequency of LOH in patients with CN-AML. We examined the frequency and prognostic significance of UPDs in a set of 425 adult patients with CN-AML who were previously sequenced for 81 genes typically mutated in cancer. Associations of UPDs with outcome were analyzed in the 315 patients with CN-AML younger than 60 years.

Results: We detected 127 UPDs in 109 patients. Most UPDs were large and typically encompassed all or most of the affected chromosome arm. The most common UPDs occurred on chromosome arms 13q (7.5% of patients), 6p (2.8%), and 11p (2.8%). Many UPDs significantly cooccurred with mutations in genes they encompassed, including 13q UPD with -internal tandem duplication (-ITD; < 0.001), and 11p UPD with mutations ( = 0.02). Among patients younger than 60 years, UPD of 11p was associated with longer overall survival (OS) and 13q UPD with shorter disease-free survival (DFS) and OS. In multivariable models that accounted for known prognostic markers, including -ITD and mutations, UPD of 13q maintained association with shorter DFS, and UPD of 11p maintained association with longer OS.

Conclusions: LOH mediated by UPD is a recurrent feature of CN-AML. Detection of UPDs of 13q and 11p might be useful for genetic risk stratification of patients with CN-AML.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-0725DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6825549PMC
November 2019

Prognostic and Biologic Relevance of Clinically Applicable Long Noncoding RNA Profiling in Older Patients with Cytogenetically Normal Acute Myeloid Leukemia.

Mol Cancer Ther 2019 08 4;18(8):1451-1459. Epub 2019 Jun 4.

The Ohio State University Comprehensive Cancer Center, Columbus, Ohio.

We have previously shown that expression levels of 48 long noncoding RNAs (lncRNA) can generate a prognostic lncRNA score that independently associates with outcome of older patients with cytogenetically normal acute myeloid leukemia (CN-AML). However, the techniques used to identify and measure prognostic lncRNAs (i.e., RNA sequencing and microarrays) are not tailored for clinical testing. Herein, we report on an assay (based on the nCounter platform) that is designed to produce targeted measurements of prognostic lncRNAs in a clinically applicable manner. We analyzed a new cohort of 76 older patients with CN-AML and found that the nCounter assay yielded reproducible measurements and that the lncRNA score retained its prognostic value; patients with high lncRNA scores had lower complete remission (CR) rates ( = 0.009; 58% vs. 87%), shorter disease-free ( = 0.05; 3-year rates: 0% vs. 21%), overall (OS; = 0.02, 3-year rates: 10% vs. 29%), and event-free survival (EFS; = 0.002, 3-year rates: 0% vs. 18%) than patients with low lncRNA scores. In multivariable analyses, the lncRNA score independently associated with CR rates ( = 0.02), OS ( = 0.02), and EFS ( = 0.02). To gain biological insights, we examined our initial cohort of 71 older patients with CN-AML, previously analyzed with RNA sequencing. Genes involved in immune response and B-cell receptor signaling were enriched in patients with high lncRNA scores. We conclude that clinically applicable lncRNA profiling is feasible and potentially useful for risk stratification of older patients with CN-AML. Furthermore, we identify potentially targetable molecular pathways that are active in the high-risk patients with high lncRNA scores.
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http://dx.doi.org/10.1158/1535-7163.MCT-18-1125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6677601PMC
August 2019

Allogeneic hematopoietic cell transplantation compared to chemotherapy consolidation in older acute myeloid leukemia (AML) patients 60-75 years in first complete remission (CR1): an alliance (A151509), SWOG, ECOG-ACRIN, and CIBMTR study.

Leukemia 2019 11 9;33(11):2599-2609. Epub 2019 May 9.

City of Hope, Duarte, CA, USA.

The preferred post-remission therapy for older patients with acute myeloid leukemia (AML) in first complete remission (CR1) remains uncertain. In this retrospective, multicenter study, we compared the outcomes for older AML patients (age 60-77 years) receiving allogeneic hematopoietic cell transplantation (alloHCT) (n = 431) with those treated on prospective National Clinical Trials Network induction and nontransplantation chemotherapy (CT) consolidation trials (n = 211). AlloHCT patients were younger (median age: 64.2 versus 67.9 years, p < 0.001), but more frequently had high-risk AML (high WBC, secondary AML, and unfavorable cytogenetics). Overall survival (OS) was worse in alloHCT during the first 9 months after CR1 (HR = 1.52, p = 0.02), but was significantly better thereafter (HR = 0.53, p < 0.0001) relative to CT. Treatment-related mortality (TRM) following HCT was worse in the first 9 months (HR = 2.8, 95% CI: 1.5-5.2, p = 0.0009), while post-HCT relapse was significantly less frequent beyond 9 months (HR = 0.42, 95% CI: 0.29-0.61, p < 0.0001). Despite higher early TRM, alloHCT recipients had superior long-term OS [29% (24-34%) versus CT 13.8% (9-21%) at 5 years]. Although this is a retrospective analysis with potential biases, it indicates that alloHCT led to heightened early risks from TRM, yet reduced relapse and superior long-term survival relative to CT in older AML patients in CR1.
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http://dx.doi.org/10.1038/s41375-019-0477-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6842042PMC
November 2019
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