Publications by authors named "James S Blachly"

65 Publications

Hairy cell leukemia and COVID-19 adaptation of treatment guidelines.

Leukemia 2021 May 4. Epub 2021 May 4.

Haematology Department, Peter MacCallum Cancer Centre and Royal Melbourne Hospital, University of Melbourne, Melbourne, VIC, Australia.

Standard treatment options in classic HCL (cHCL) result in high response rates and near normal life expectancy. However, the disease itself and the recommended standard treatment are associated with profound and prolonged immunosuppression, increasing susceptibility to infections and the risk for a severe course of COVID-19. The Hairy Cell Leukemia Foundation (HCLF) has recently convened experts and discussed different clinical strategies for the management of these patients. The new recommendations adapt the 2017 consensus for the diagnosis and management with cHCL to the current COVID-19 pandemic. They underline the option of active surveillance in patients with low but stable blood counts, consider the use of targeted and non-immunosuppressive agents as first-line treatment for cHCL, and give recommendations on preventive measures against COVID-19.
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http://dx.doi.org/10.1038/s41375-021-01257-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8093591PMC
May 2021

Phase 2 Study of Ibrutinib in Classic and Variant Hairy Cell Leukemia.

Blood 2021 Mar 22. Epub 2021 Mar 22.

Ohio State University, Colmubus, Ohio, United States.

Hairy cell leukemia is a rare B-cell malignancy where there is a need for novel treatments for patients who do not benefit from purine analogues. Ibrutinib, an oral agent targeting Bruton tyrosine kinase in the B-cell receptor signaling pathway, is highly effective in several malignancies. Its activity in HCL was unknown. Therefore, we conducted a multisite phase 2 study (NCT01841723) of oral ibrutinib in patients with either relapsed classic or variant hairy cell leukemia. The primary outcome measure was the overall response rate at 32 weeks with response at 48 weeks and best response during treatment also assessed. Key secondary objectives were characterization of toxicity and determination of progression-free and overall survival. Thirty-seven patients were enrolled (24 at 420mg, 13 at 840mg). The median duration of follow-up was 3.5 years (range 0-5.9). The overall response rate at 32 weeks was 24% which increased to 36% at 48 weeks. The best overall response rate was 54%. The estimated 36-month progression-free and overall survivals were 73% and 85%, respectively. The most frequent adverse events were diarrhea (59%), fatigue (54%), myalgia (54%), and nausea (51%). Hematologic adverse events were common with anemia (43%), thrombocytopenia (41%), and neutropenia (35%). Ibrutinib can be safely administered to hairy cell leukemia patients with objective responses and results in prolonged disease control. While the initial primary outcome objective of the study was not met, the observation of objective responses in heavily pretreated patients coupled with a favorable progression-free survival suggest that ibrutinib may be beneficial in these patients.
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http://dx.doi.org/10.1182/blood.2020009688DOI Listing
March 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
May 2021

Targeting DNA Damage Repair Functions of Two Histone Deacetylases, HDAC8 and SIRT6, Sensitizes Acute Myeloid Leukemia to NAMPT Inhibition.

Clin Cancer Res 2021 Apr 4;27(8):2352-2366. Epub 2021 Feb 4.

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

Purpose: Nicotinamide phosphoribosyltransferase (NAMPT) inhibitors (NAMPTi) are currently in development, but may be limited as single-agent therapy due to compound-specific toxicity and cancer metabolic plasticity allowing resistance development. To potentially lower the doses of NAMPTis required for therapeutic benefit against acute myeloid leukemia (AML), we performed a genome-wide CRISPRi screen to identify rational disease-specific partners for a novel NAMPTi, KPT-9274.

Experimental Design: Cell lines and primary cells were analyzed for cell viability, self-renewal, and responses at RNA and protein levels with loss-of-function approaches and pharmacologic treatments. efficacy of combination therapy was evaluated with a xenograft model.

Results: We identified two histone deacetylases (HDAC), and , whose knockout conferred synthetic lethality with KPT-9274 in AML. Furthermore, HDAC8-specific inhibitor, PCI-34051, or clinical class I HDAC inhibitor, AR-42, in combination with KPT-9274, synergistically decreased the survival of AML cells in a dose-dependent manner. AR-42/KPT-9274 cotreatment attenuated colony-forming potentials of patient cells while sparing healthy hematopoietic cells. Importantly, combined therapy demonstrated promising efficacy compared with KPT-9274 or AR-42 monotherapy. Mechanistically, genetic inhibition of SIRT6 potentiated the effect of KPT-9274 on PARP-1 suppression by abolishing mono-ADP ribosylation. AR-42/KPT-9274 cotreatment resulted in synergistic attenuation of homologous recombination and nonhomologous end joining pathways in cell lines and leukemia-initiating cells.

Conclusions: Our findings provide evidence that HDAC8 inhibition- or shSIRT6-induced DNA repair deficiencies are potently synergistic with NAMPT targeting, with minimal toxicity toward normal cells, providing a rationale for a novel-novel combination-based treatment for AML.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-3724DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8054771PMC
April 2021

Poor Survival and Differential Impact of Genetic Features of Black Patients with Acute Myeloid Leukemia.

Cancer Discov 2021 Mar 4;11(3):626-637. Epub 2020 Dec 4.

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

Clinical outcome of patients with acute myeloid leukemia (AML) is associated with cytogenetic and molecular factors and patient demographics (e.g., age and race). We compared survival of 25,523 non-Hispanic Black and White adults with AML using Surveillance Epidemiology and End Results (SEER) Program data and performed mutational profiling of 1,339 patients with AML treated on frontline Alliance for Clinical Trials in Oncology (Alliance) protocols. Black patients had shorter survival than White patients, both in SEER and in the setting of Alliance clinical trials. The disparity was especially pronounced in Black patients <60 years, after adjustment for socioeconomic (SEER) and molecular (Alliance) factors. Black race was an independent prognosticator of poor survival. Gene mutation profiles showed fewer and more mutations in younger Black patients. Overall survival of younger Black patients was adversely affected by mutations and -ITD, but, in contrast to White patients, was not improved by mutations. SIGNIFICANCE: We show that young Black patients have not benefited as much as White patients from recent progress in AML treatment in the United States. Our data suggest that both socioeconomic factors and differences in disease biology contribute to the survival disparity and need to be urgently addressed...
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http://dx.doi.org/10.1158/2159-8290.CD-20-1579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7933110PMC
March 2021

TP-0903 is active in models of drug-resistant acute myeloid leukemia.

JCI Insight 2020 12 3;5(23). Epub 2020 Dec 3.

Division of Pharmaceutics and Pharmacology, College of Pharmacy.

Effective treatment for AML is challenging due to the presence of clonal heterogeneity and the evolution of polyclonal drug resistance. Here, we report that TP-0903 has potent activity against protein kinases related to STAT, AKT, and ERK signaling, as well as cell cycle regulators in biochemical and cellular assays. In vitro and in vivo, TP-0903 was active in multiple models of drug-resistant FLT3 mutant AML, including those involving the F691L gatekeeper mutation and bone marrow microenvironment-mediated factors. Furthermore, TP-0903 demonstrated preclinical activity in AML models with FLT3-ITD and common co-occurring mutations in IDH2 and NRAS genes. We also showed that TP-0903 had ex vivo activity in primary AML cells with recurrent mutations including MLL-PTD, ASXL1, SRSF2, and WT1, which are associated with poor prognosis or promote clinical resistance to AML-directed therapies. Our preclinical studies demonstrate that TP-0903 is a multikinase inhibitor with potent activity against multiple drug-resistant models of AML that will have an immediate clinical impact in a heterogeneous disease like AML.
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http://dx.doi.org/10.1172/jci.insight.140169DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7714403PMC
December 2020

Synergistic effect of BCL2 and FLT3 co-inhibition in acute myeloid leukemia.

J Hematol Oncol 2020 10 19;13(1):139. Epub 2020 Oct 19.

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

Acute myeloid leukemia (AML) is a heterogeneous and complex disease, and treatments for this disease have not been curative for the majority of patients. In younger patients, internal tandem duplication of FLT3 (FLT3-ITD) is a common mutation for which two inhibitors (midostaurin and gilteritinib) with varied potency and specificity for FLT3 are clinically approved. However, the high rate of relapse or failed initial response of AML patients suggests that the addition of a second targeted therapy may be necessary to improve efficacy. Using an unbiased large-scale CRISPR screen, we genetically identified BCL2 knockout as having synergistic effects with an approved FLT3 inhibitor. Here, we provide supportive studies that validate the therapeutic potential of the combination of FLT3 inhibitors with venetoclax in vitro and in vivo against multiple models of FLT3-ITD-driven AML. Our unbiased approach provides genetic validation for co-targeting FLT3 and BCL2 and repurposes CRISPR screening data, utilizing the genome-wide scope toward mechanistic understanding.
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http://dx.doi.org/10.1186/s13045-020-00973-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7574303PMC
October 2020

Characterization and mitigation of fragmentation enzyme-induced dual stranded artifacts.

NAR Genom Bioinform 2020 Dec 2;2(4):lqaa070. Epub 2020 Oct 2.

Division of Hematology, Ohio State University, Columbus, OH 43210, USA.

High-throughput short-read sequencing relies on fragmented DNA for optimal sampling of input nucleic acid. Several vendors now offer proprietary enzyme cocktails as a cheaper and more streamlined method of fragmentation when compared to acoustic shearing. We have discovered that these enzymes induce the formation of library molecules containing regions of nearby DNA from opposite strands. Sequencing reads derived from these molecules can lead to artifact-derived variant calls appearing at variant allele frequencies <5%. We present Fragmentation Artifact Detection and Elimination (FADE), software to remove these artifacts from mapped reads and mitigate artifact-related effects on downstream analysis. We find that the artifacts principally affect downstream analyses that are sensitive to a 1-3% artifact bias in the sequencing reads, such as targeted resequencing and rare variant discovery.
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http://dx.doi.org/10.1093/nargab/lqaa070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7531576PMC
December 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

Incidence of venous thrombosis after peg-asparaginase in adolescent and young adults with acute lymphoblastic leukemia.

Int J Hematol Oncol 2020 Sep 4;9(3):IJH28. Epub 2020 Sep 4.

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

Aim: There are limited data describing incidence of symptomatic venous thromboembolism (VTE) in adolescent and young adult (AYA) acute lymphoblastic leukemia (ALL) patients receiving peg-asparaginase.

Materials & Methods: Single-institution retrospective analysis of 44 AYA ALL patients treated with peg-asparaginase. Rates of VTE and proposed risk factors were assessed.

Results: 18 patients (41%) had a symptomatic VTE following peg-asparaginase. The cumulative incidence rate was 25% (95% CI: 13-38%) within 30 days of the initial dose. Personal history of thrombosis was statistically significantly associated with an increased risk of VTE with HR of 2.73 (95% CI: 1.40-5.33, p = 0.003) after adjusting for gender.

Conclusion: These data indicate a high rate of VTE in the AYA ALL population following treatment with peg-asparaginase.
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http://dx.doi.org/10.2217/ijh-2020-0009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7521187PMC
September 2020

The Incidence of Invasive Fungal Infections in Patients With AML Treated With a Hypomethylating Agent.

Clin Lymphoma Myeloma Leuk 2021 Jan 21;21(1):e76-e83. Epub 2020 Aug 21.

Division of Hematology, The Ohio State University Wexner Medical Center, Columbus, OH. Electronic address:

Background: Newly diagnosed patients with acute myeloid leukemia (AML) who receive induction with a hypomethylating agent (HMA) are often neutropenic with an increased risk for invasive fungal infections (IFIs). This study analyzed the incidence and risk factors for IFIs in these patients, evaluated clinical patterns in antifungal prophylaxis, and assessed the diagnostic utility of tests in this setting.

Patients And Methods: We studied 117 newly diagnosed patients with AML treated with HMAs at our center, divided into groups based on concern for IFI (cIFI: all possible, probable, and proven IFIs) versus no concern for IFI. The Fisher exact test compared patients with cIFI versus without, and a multivariable logistic regression model estimated odds for cIFI.

Results: Sixty-seven (57%) patients had cIFI, with 48 possible IFIs, 17 probable, and 2 proven cases. There was no difference in incidence based on home zip code, but the presence of chronic obstructive pulmonary disease was highly associated with cIFI (P = .001), as was male gender (P = .01). Neutropenia at treatment initiation was borderline in significance (P = .08). In diagnostics, 9% of patients had positive serum fungal markers, and 30 patients underwent bronchoscopy, with only 27% of cases yielding positive results. There was a difference in treatment regimens between patients receiving antifungal prophylaxis with mold coverage versus without mold coverage with respect to cIFI (P = .04).

Conclusions: cIFI in patients with AML treated with HMAs remains significant, especially in males and those with chronic obstructive pulmonary disease, who were found to be at higher risk. This may prompt clinicians to consider anti-mold prophylaxis in this setting.
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http://dx.doi.org/10.1016/j.clml.2020.08.013DOI Listing
January 2021

Entospletinib in Combination with Induction Chemotherapy in Previously Untreated Acute Myeloid Leukemia: Response and Predictive Significance of and Expression.

Clin Cancer Res 2020 Nov 20;26(22):5852-5859. Epub 2020 Aug 20.

Winship Cancer Institute of Emory University, Atlanta, Georgia.

Purpose: Spleen tyrosine kinase (SYK) signaling is a proposed target in acute myeloid leukemia (AML). Sensitivity to SYK inhibition has been linked to and overexpression in preclinical studies. This trial evaluated the safety and efficacy of entospletinib, a selective inhibitor of SYK, in combination with chemotherapy in untreated AML.

Patients And Methods: This was an international multicenter phase Ib/II study, entospletinib dose escalation (standard 3+3 design between 200 and 400 mg twice daily) + 7+3 (cytarabine + daunorubicin) in phase Ib and entospletinib dose expansion (400 mg twice daily) + 7+3 in phase II.

Results: Fifty-three patients ( = 12, phase Ib and = 41, phase II) with previously untreated ( = 39) or secondary ( = 14) AML were enrolled (58% male; median age, 60 years) in this study. The composite complete response with entospletinib + 7+3 was 70%. Patients with baseline and expression higher than the median had improved overall survival compared with patients with below median and expression. Common adverse events were cytopenias, febrile neutropenia, and infection. There were no dose-limiting toxicities. Entospletinib-related skin rash and hyperbilirubinemia were also observed.

Conclusions: Entospletinib with intensive chemotherapy was well-tolerated in patients with AML. Improved survival was observed in patients with overexpression, contrasting published data demonstrating poor survival in such patients. A randomized study will be necessary to determine whether entospletinib was a mediator this observation.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-1064DOI Listing
November 2020

A phase I study of lenalidomide plus chemotherapy with idarubicin and cytarabine in patients with relapsed or refractory acute myeloid leukemia and high-risk myelodysplastic syndrome.

Am J Hematol 2020 12 19;95(12):1457-1465. Epub 2020 Sep 19.

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

Patients with relapsed/refractory (R/R) acute myeloid leukemia (AML) have poor outcomes and hematopoietic cell transplantation (HCT) is the only curative treatment. New targeted therapies improved survival in select patients with specific mutations, however management of patients without these molecular alterations is an unmet need. We conducted a phase one study of lenalidomide in combination with cytarabine/idarubicin salvage chemotherapy in patients with R/R AML and high-risk myelodysplastic syndromes. A total of 33 patients were enrolled in the study (30 AML, 3 MDS), and treated at three dose levels with 3 + 3 design. Dose-limiting toxicity (DLT) was seen in eight patients, including four hematologic DLTs. The most commonly observed non-hematologic serious adverse events were febrile neutropenia, rash, sepsis and renal injury. Dose level -1, consisting of 25 mg/d lenalidomide D1-21, 1 g/m cytarabine D5-8, and 8 mg/m idarubicin D5-7 was determined to be the maximum tolerated dose. Note, 15/33 (45%) of patients were able to receive pre-planned 21 days of lenalidomide. Overall, 18 patients achieved complete remission (CR) (n = 14) or CR with incomplete count recovery (CRi) (n = 4) with total CR/CRi rate of 56%. The 1-year and 2-year overall survival (OS) were 24% and 10%, respectively. Among responders, 10/18 underwent allogeneic HCT and had a 1-year OS of 40%. There was no molecular pattern associated with response. These data demonstrate that the combination had clinical activity in R/R AML. This regimen should be further investigated for patients who relapsed after HCT, and as a bridge therapy to HCT. (ClinicalTrials.gov identifier: NCT01132586).
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http://dx.doi.org/10.1002/ajh.25958DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7821016PMC
December 2020

LC-FACSeq is a method for detecting rare clones in leukemia.

JCI Insight 2020 06 18;5(12). Epub 2020 Jun 18.

Division of Hematology, Department of Internal Medicine and Comprehensive Cancer Center.

Detecting, characterizing, and monitoring rare populations of cells can increase testing sensitivity, give insight into disease mechanism, and inform clinical decision making. One area that can benefit from increased resolution is management of cancers in clinical remission but with measurable residual disease (MRD) by multicolor FACS. Detecting and monitoring genomic clonal resistance to treatment in the setting of MRD is technically difficult and resource intensive due to the limited amounts of disease cells. Here, we describe limited-cell FACS sequencing (LC-FACSeq), a reproducible, highly sensitive method of characterizing clonal evolution in rare cells relevant to different types of acute and chronic leukemias. We demonstrate the utility of LC-FACSeq for broad multigene gene panels and its application for monitoring sequential acquisition of mutations conferring therapy resistance and clonal evolution in long-term ibrutinib treatment of patients with chronic lymphocytic leukemia. This technique is generalizable for monitoring of other blood and marrow infiltrating cancers.
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http://dx.doi.org/10.1172/jci.insight.134973DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7406301PMC
June 2020

Cotargeting of XPO1 Enhances the Antileukemic Activity of Midostaurin and Gilteritinib in Acute Myeloid Leukemia.

Cancers (Basel) 2020 Jun 14;12(6). Epub 2020 Jun 14.

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

Acute myeloid leukemia (AML) is a hematopoietic stem-cell-derived leukemia with often successive derived driver mutations. Late onset acquisition of internal tandem duplication in (-ITD) at a high variant allele frequency often contributes to full transformation to a highly proliferative, rapidly progressive disease with poor outcome. The -ITD mutation is targetable with approved FLT3 small molecule inhibitors, including midostaurin and gilteritinib. However, outside of patients receiving allogeneic transplant, most patients fail to respond or relapse, suggesting alternative approaches of therapy will be required. We employed genome-wide pooled CRISPR knockout screening as a method for large-scale identification of targets whose knockout produces a phenotypic effect that enhances the antitumor properties of FLT3 inhibitors. Among the candidate targets we identified the effect of XPO1 knockout to be synergistic with midostaurin treatment. Next, we validated the genetic finding with pharmacologic combination of the slowly reversible XPO1 inhibitor selinexor with midostaurin and gilteritinib in -ITD AML cell lines and primary patient samples. Lastly, we demonstrated improved survival with either combination therapy compared to its monotherapy components in an aggressive AML murine model, supporting further evaluation and rapid clinical translation of this combination strategy.
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http://dx.doi.org/10.3390/cancers12061574DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7352446PMC
June 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

Quantifying Hematopoietic Stem Cell Clonal Diversity by Selecting Informative Amplicon Barcodes.

Sci Rep 2020 02 7;10(1):2153. Epub 2020 Feb 7.

The Ohio State University College of Medicine, Department of Medicine, Division of Hematology, The Ohio State University Comprehensive Cancer Center, Ohio, USA.

Hematopoietic stem cells (HSCs) are functionally and genetically diverse and this diversity decreases with age and disease. Numerous systems have been developed to quantify HSC diversity by genetic barcoding, but no framework has been established to empirically validate barcode sequences. Here we have developed an analytical framework, Selection of informative Amplicon Barcodes from Experimental Replicates (SABER), that identifies barcodes that are unique among a large set of experimental replicates. Amplicon barcodes were sequenced from the blood of 56 adult zebrafish divided into training and validation sets. Informative barcodes were identified and samples with a high fraction of informative barcodes were chosen by bootstrapping. There were 4.2 ± 1.8 barcoded HSC clones per sample in the training set and 3.5 ± 2.1 in the validation set (p = 0.3). SABER reproducibly quantifies functional HSCs and can accommodate a wide range of experimental group sizes. Future large-scale studies aiming to understand the mechanisms of HSC clonal evolution will benefit from this new approach to identifying informative amplicon barcodes.
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http://dx.doi.org/10.1038/s41598-020-59119-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7005852PMC
February 2020

Acalabrutinib plus Obinutuzumab in Treatment-Naïve and Relapsed/Refractory Chronic Lymphocytic Leukemia.

Cancer Discov 2020 03 8;10(3):394-405. Epub 2020 Jan 8.

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

Acalabrutinib is a selective irreversible Bruton tyrosine kinase (BTK) inhibitor that does not affect IL2-associated tyrosine kinase or antibody-dependent cellular cytotoxicity, making it an attractive candidate for combination therapy with anti-CD20 antibodies. We investigated acalabrutinib plus obinutuzumab in a phase Ib/II study (NCT02296918) of patients with treatment-naïve or relapsed/refractory chronic lymphocytic leukemia (CLL). Nineteen treatment-naïve and 26 relapsed/refractory patients were treated with acalabrutinib (100 mg twice daily) until progression and obinutuzumab (cycle 1: 100 mg day 1, 900 mg day 2, 1000 mg days 8 and 15; cycles 2-6: 1,000 mg day 1). Grade 3/4 adverse events occurred in 71% of patients. Overall response rates were 95% (treatment-naïve) and 92% (relapsed/refractory). Thirty-two percent of treatment-naïve and 8% of relapsed/refractory patients achieved complete remission. At 36 months, 94% (treatment-naïve) and 88% (relapsed/refractory) were progression free. Acalabrutinib plus obinutuzumab was well tolerated, producing high and durable responses in treatment-naïve and relapsed/refractory CLL. SIGNIFICANCE: Rituximab plus the less selective BTK inhibitor ibrutinib has not shown benefit in CLL; however, the selective BTK inhibitor acalabrutinib plus the antibody-dependent cellular cytotoxicity-enhanced antibody obinutuzumab yielded durable responses that deepened over time in treatment-naïve and relapsed/refractory CLL, supporting the evaluation of this approach in larger, comparative studies in CLL..
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http://dx.doi.org/10.1158/2159-8290.CD-19-1130DOI Listing
March 2020

Resistance Mechanisms to SYK Inhibition in Acute Myeloid Leukemia.

Cancer Discov 2020 02 26;10(2):214-231. Epub 2019 Nov 26.

Department of Pediatric Oncology, Dana-Farber Cancer Institute and Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts.

Spleen tyrosine kinase (SYK) is a nonmutated therapeutic target in acute myeloid leukemia (AML). Attempts to exploit SYK therapeutically in AML have shown promising results in combination with chemotherapy, likely reflecting induced mechanisms of resistance to single-agent treatment . We conducted a genome-scale open reading frame (ORF) resistance screen and identified activation of the RAS-MAPK-ERK pathway as one major mechanism of resistance to SYK inhibitors. This finding was validated in AML cell lines with innate and acquired resistance to SYK inhibitors. Furthermore, patients with AML with select mutations activating these pathways displayed early resistance to SYK inhibition. To circumvent SYK inhibitor therapy resistance in AML, we demonstrate that a MEK and SYK inhibitor combination is synergistic and . Our data provide justification for use of ORF screening to identify resistance mechanisms to kinase inhibitor therapy in AML lacking distinct mutations and to direct novel combination-based strategies to abrogate these. SIGNIFICANCE: The integration of functional genomic screening with the study of mechanisms of intrinsic and acquired resistance in model systems and human patients identified resistance to SYK inhibitors through MAPK signaling in AML. The dual targeting of SYK and the MAPK pathway offers a combinatorial strategy to overcome this resistance..
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http://dx.doi.org/10.1158/2159-8290.CD-19-0209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7058374PMC
February 2020

Transcriptionally Active Androgen Receptor Splice Variants Promote Hepatocellular Carcinoma Progression.

Cancer Res 2020 02 4;80(3):561-575. Epub 2019 Nov 4.

Division of Pharmaceutics and Pharmaceutical Chemistry, College of Pharmacy, The Ohio State University, Columbus, Ohio.

Owing to the marked sexual dimorphism of hepatocellular carcinoma (HCC), sex hormone receptor signaling has been implicated in numerous aspects of liver cancer pathogenesis. We sought to reconcile the clear contribution of androgen receptor (AR) activity that has been established in preclinical models of HCC with the clinical failure of antagonists in patients with advanced HCC by evaluating potential resistance mechanisms to AR-targeted therapy. The locus was interrogated for resistance-causing genomic modifications using publicly available primary HCC datasets (1,019 samples). Analysis of HCC tumor and cell line RNA-seq data revealed enriched expression of constitutively active, treatment-refractory AR splice variants (AR-SV). HCC cell lines expressed C-terminal-truncated AR-SV; 28 primary HCC samples abundantly expressed AR-SV. Low molecular weight AR species were nuclear localized and constitutively active. Furthermore, AR/AR-SV signaling promoted AR-mediated HCC cell progression and conferred resistance to AR antagonists. Ligand-dependent and -independent AR signaling mediated HCC epithelial-to-mesenchymal transition by regulating the transcription factor SLUG. These data suggest that AR-SV expression in HCC drives HCC progression and resistance to traditional AR antagonists. Novel therapeutic approaches that successfully target AR-SVs may be therapeutically beneficial for HCC. SIGNIFICANCE: Treatment-refractory, constitutively active androgen receptor splice variants promote hepatocellular carcinoma progression by regulating the epithelial-to-mesenchymal transition pathway.
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http://dx.doi.org/10.1158/0008-5472.CAN-19-1117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7002251PMC
February 2020

Selinexor in combination with decitabine in patients with acute myeloid leukemia: results from a phase 1 study.

Leuk Lymphoma 2020 02 23;61(2):387-396. Epub 2019 Sep 23.

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

Current treatment options for older and relapsed or refractory (R/R) acute myeloid leukemia (AML) patients are limited and represent an unmet need. Based on preclinical studies showing strong anti-leukemic effects this phase I dose-escalation study assessed the safety and preliminary clinical activity of the oral exportin-1 inhibitor, selinexor, in combination with the hypomethylating agent, decitabine 20 mg/m, in adults with R/R AML and in older (age ≥ 60) untreated AML patients. There were no protocol-defined dose limiting toxicities. The recommended phase 2 dose of selinexor was 60 mg (∼35 mg/m) given twice-weekly. Notable grade ≥3 toxicities included asymptomatic hyponatremia (68%), febrile neutropenia (44%), sepsis (44%), hypophosphatemia (36%), and pneumonia (28%). In 25 patients, the overall response rate was 40%. Modification of selinexor to a flat dose of 60 mg, twice-weekly for two weeks after decitabine, improved tolerability of the regimen and demonstrated preliminary clinical activity in poor-risk patients with AML.
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http://dx.doi.org/10.1080/10428194.2019.1665664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7552944PMC
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

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

Uncovering the Genomic Landscape in Newly Diagnosed and Relapsed Pediatric Cytogenetically Normal FLT3-ITD AML.

Clin Transl Sci 2019 11 20;12(6):641-647. Epub 2019 Aug 20.

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

Fms-like tyrosine kinase 3 (FLT3) internal tandem duplication (ITD) mutations, common in pediatric acute myeloid leukemia (AML), associate with early relapse and poor prognosis. Past studies have suggested additional cooperative mutations are required for leukemogenesis in FLT3-ITD+ AML. Using RNA sequencing and a next-generation targeted gene panel, we broadly characterize the co-occurring genomic alterations in pediatric cytogenetically normal (CN) FLT3-ITD+ AML to gain a deeper understanding of the clonal patterns and heterogeneity at diagnosis and relapse. We show that chimeric transcripts were present in 21 of 34 (62%) of de novo samples, 2 (6%) of these samples included a rare reoccurring fusion partner BCL11B. At diagnosis, the median number of mutations other than FLT3 per patient was 1 (range 0-3), which involved 8 gene pathways; WT1 and NPM1 mutations were frequently observed (35% and 24%, respectively). Fusion transcripts and high variant allele frequency (VAF) mutants, which included WT1, NPM1, SMARCA2, RAD21, and TYK2, were retained from diagnosis to relapse. We did observe reduction in VAF of simple or single mutation clones, but VAFs were preserved or expanded in more complex clones with multiple mutations. Our data provide the first insight into the genomic complexity of pediatric CN FLT3-ITD+ AML and could help stratify future targeted treatment strategies.
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http://dx.doi.org/10.1111/cts.12669DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6853146PMC
November 2019

Developmental subtypes assessed by DNA methylation-iPLEX forecast the natural history of chronic lymphocytic leukemia.

Blood 2019 08 10;134(8):688-698. Epub 2019 Jul 10.

Division of Hematology, Department of Internal Medicine, and.

Alterations in global DNA methylation patterns are a major hallmark of cancer and represent attractive biomarkers for personalized risk stratification. Chronic lymphocytic leukemia (CLL) risk stratification studies typically focus on time to first treatment (TTFT), time to progression (TTP) after treatment, and overall survival (OS). Whereas TTFT risk stratification remains similar over time, TTP and OS have changed dramatically with the introduction of targeted therapies, such as the Bruton tyrosine kinase inhibitor ibrutinib. We have shown that genome-wide DNA methylation patterns in CLL are strongly associated with phenotypic differentiation and patient outcomes. Here, we developed a novel assay, termed methylation-iPLEX (Me-iPLEX), for high-throughput quantification of targeted panels of single cytosine guanine dinucleotides from multiple independent loci. Me-iPLEX was used to classify CLL samples into 1 of 3 known epigenetic subtypes (epitypes). We examined the impact of epitype in 1286 CLL patients from 4 independent cohorts representing a comprehensive view of CLL disease course and therapies. We found that epitype significantly predicted TTFT and OS among newly diagnosed CLL patients. Additionally, epitype predicted TTP and OS with 2 common CLL therapies: chemoimmunotherapy and ibrutinib. Epitype retained significance after stratifying by biologically related biomarkers, immunoglobulin heavy chain mutational status, and ZAP70 expression, as well as other common prognostic markers. Furthermore, among several biological traits enriched between epitypes, we found highly biased immunogenetic features, including IGLV3-21 usage in the poorly characterized intermediate-programmed CLL epitype. In summary, Me-iPLEX is an elegant method to assess epigenetic signatures, including robust classification of CLL epitypes that independently stratify patient risk at diagnosis and time of treatment.
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http://dx.doi.org/10.1182/blood.2019000490DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6706807PMC
August 2019

Complex karyotype in de novo acute myeloid leukemia: typical and atypical subtypes differ molecularly and clinically.

Leukemia 2019 07 8;33(7):1620-1634. Epub 2019 Feb 8.

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

Complex karyotype (CK) with ≥ 3 abnormalities is detected in 10-12% of patients with acute myeloid leukemia (AML) and associated with poor prognosis. The most common unbalanced abnormalities found in CK result in loss of material from the 5q, 7q, and/or 17p chromosome arms. The presence of 5q, 7q, and/or 17p abnormalities denotes typical CK and their absence denotes atypical CK. Since molecular features of CK-AML are not well characterized, we investigated mutational status of 81 leukemia/cancer-associated genes in 160 clinically well-characterized patients. They included 136 patients with ≥ 3 exclusively unbalanced chromosome abnormalities, 96 of whom had a typical CK and 40 atypical CK, and 24 patients with ≥ 1 balanced abnormality in addition to ≥ 2 unbalanced ones. Patients with atypical CK-AML differed from those with typical CK-AML: they carried TP53 mutations less often (P < 0.001) and more often PHF6 (P = 0.008), FLT3-TKD (P = 0.02), MED12 (P = 0.02), and NPM1 (P = 0.02) mutations. They were younger (P = 0.007), had higher WBC (P = 0.001) and percentages of marrow (P < 0.001) and blood (P = 0.006) blasts, higher complete remission rates (P = 0.02), and longer overall survival (P < 0.001), thus indicating that atypical and typical CK-AMLs constitute distinct disease subtypes. We also identified smaller patient subsets within both typical and atypical CK-AML that differed molecularly and clinically.
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http://dx.doi.org/10.1038/s41375-019-0390-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6609457PMC
July 2019

Selective targeting of NAMPT by KPT-9274 in acute myeloid leukemia.

Blood Adv 2019 02;3(3):242-255

Division of Hematology.

Treatment options for acute myeloid leukemia (AML) remain extremely limited and associated with significant toxicity. Nicotinamide phosphoribosyltransferase (NAMPT) is involved in the generation of NAD and a potential therapeutic target in AML. We evaluated the effect of KPT-9274, a p21-activated kinase 4/NAMPT inhibitor that possesses a unique NAMPT-binding profile based on in silico modeling compared with earlier compounds pursued against this target. KPT-9274 elicited loss of mitochondrial respiration and glycolysis and induced apoptosis in AML subtypes independent of mutations and genomic abnormalities. These actions occurred mainly through the depletion of NAD, whereas genetic knockdown of p21-activated kinase 4 did not induce cytotoxicity in AML cell lines or influence the cytotoxic effect of KPT-9274. KPT-9274 exposure reduced colony formation, increased blast differentiation, and diminished the frequency of leukemia-initiating cells from primary AML samples; KPT-9274 was minimally cytotoxic toward normal hematopoietic or immune cells. In addition, KPT-9274 improved overall survival in vivo in 2 different mouse models of AML and reduced tumor development in a patient-derived xenograft model of AML. Overall, KPT-9274 exhibited broad preclinical activity across a variety of AML subtypes and warrants further investigation as a potential therapeutic agent for AML.
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http://dx.doi.org/10.1182/bloodadvances.2018024182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6373756PMC
February 2019