pevonedistat and Leukemia--Myeloid--Acute

Conventional chemotherapy with cytarabine and anthracycline (often referred to as "7+3") has been used for many years in the treatment of acute myeloid leukemia (AML). Despite meaningful advances in areas of supportive care and transplantation, little progress has been made in developing new chemotherapy options. In 2018, The Food and Drug Administration (FDA) of the US approved several novel agents for AML treatment as follows: ivosidenib, an inhibitor of isocitrate dehydrogenase-1; venetoclax, a potent inhibitor of bcl2; and glasdegib, an inhibitor of hedgehog signaling pathway. Moreover, clinical trials of alvocidib (flavopiridol), an inhibitor of the CDK9, pevonedistat, an inhibitor of NEDD8, and APR-246, a reactivator of mutant p53, are in progress. These agents will either be incorporated into the conventional 7+3 regimen or combined with hypomethylating agents to improve the outcome of AML therapy, and the results will guide the next stage of precision medicine in the treatment of AML.

Topics: Benzimidazoles; Bridged Bicyclo Compounds, Heterocyclic; Cyclopentanes; Drug Approval; Flavonoids; Glycine; Humans; Leukemia, Myeloid, Acute; Molecular Targeted Therapy; Phenylurea Compounds; Piperidines; Pyridines; Pyrimidines; Sulfonamides; United States; United States Food and Drug Administration

Topics: Aged; Azacitidine; Cyclopentanes; Humans; Leukemia, Myeloid, Acute; Pyrimidines; Tumor Suppressor Protein p53

Topics: Antimetabolites, Antineoplastic; Azacitidine; Cyclopentanes; Enzyme Inhibitors; Humans; Leukemia, Myeloid, Acute; Leukemia, Myelomonocytic, Chronic; Myelodysplastic Syndromes; Pyrimidines

The investigational NEDD8-activating enzyme inhibitor pevonedistat is being evaluated in combination with azacitidine versus single-agent azacitidine in patients with higher-risk myelodysplastic syndrome (higher-risk MDS), higher-risk chronic myelomonocytic leukemia (higher-risk CMML), or low-blast acute myeloid leukemia (AML) in a Phase 3 trial PANTHER. To support Asia-inclusive global development, we applied multiregional clinical trial (MRCT) principles of the International Conference on Harmonisation E17 guidelines by evaluating similarity in drug-related and disease-related intrinsic and extrinsic factors. A PubMed literature review (January 2000-November 2019) supported similarity in epidemiology of higher-risk MDS, AML, and CMML in Western and East Asian populations. Furthermore, the treatment of MDS/AML was similar in both East Asian and Western regions, with the same dose of azacitidine being the standard of care. Median overall survival in MDS following azacitidine treatment was generally comparable across regions, and the types and frequencies of molecular alterations in AML and MDS were comparable. Dose-escalation studies established the same maximum tolerated dose of pevonedistat in combination with azacitidine in Western and East Asian populations. Pevonedistat clearance was similar across races. Taken together, conservation of drug-related and disease-related intrinsic and extrinsic factors supported design of an Asia-inclusive Phase 3 trial and a pooled East Asian region. A sample size of ~ 30 East Asian patients (of ~ 450 randomized) was estimated as needed to demonstrate consistency in efficacy relative to the global population. This analysis is presented as an exemplar to illustrate application of clinical pharmacology and translational science principles in designing Asia-inclusive MRCTs. Study Highlights WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC? Azacitidine is the standard of care for myelodysplastic syndromes/low-blast acute myeloid leukemia (AML) across Western and East Asian patients. The first-in-class small-molecule inhibitor of NEDD8-activating enzyme, pevonedistat, has been investigated as a single agent in multiple studies of hematologic and nonhematologic malignancies and in combination with azacitidine in elderly patients with untreated AML. WHAT QUESTION DID THIS STUDY ADDRESS? By applying clinical pharmacology and translational science and International Conference on Harmonisation E17 principles, this study designed an E

Topics: Antineoplastic Combined Chemotherapy Protocols; Asia; Azacitidine; Cyclopentanes; Drugs, Investigational; Global Burden of Disease; Humans; Incidence; International Cooperation; Leukemia, Myeloid, Acute; Leukemia, Myelomonocytic, Chronic; Maximum Tolerated Dose; Myelodysplastic Syndromes; Pharmacology, Clinical; Pyrimidines; Translational Research, Biomedical; Ubiquitin-Activating Enzymes; United States

Topics: Aged, 80 and over; Aniline Compounds; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Bridged Bicyclo Compounds, Heterocyclic; Cyclopentanes; Disease-Free Survival; Female; Glycine; Humans; Leukemia, Myeloid, Acute; Male; Pyrazines; Pyridines; Pyrimidines; Retrospective Studies; Sulfonamides; Survival Rate

Topics: Adult; Aged; Aged, 80 and over; Cyclopentanes; Enzyme Inhibitors; Female; Humans; Leukemia, Myeloid, Acute; Male; Maximum Tolerated Dose; Middle Aged; Myelodysplastic Syndromes; NEDD8 Protein; Pyrimidines; Young Adult

This trial was conducted to determine the dose-limiting toxicities (DLTs) and maximum tolerated dose (MTD) of the first in class NEDD8-activating enzyme (NAE) inhibitor, pevonedistat, and to investigate pevonedistat pharmacokinetics and pharmacodynamics in patients with acute myeloid leukaemia (AML) and myelodysplastic syndromes (MDS). Pevonedistat was administered via a 60-min intravenous infusion on days 1, 3 and 5 (schedule A, n = 27), or days 1, 4, 8 and 11 (schedule B, n = 26) every 21-days. Dose escalation proceeded using a standard '3 + 3' design. Responses were assessed according to published guidelines. The MTD for schedules A and B were 59 and 83 mg/m(2) , respectively. On schedule A, hepatotoxicity was dose limiting. Multi-organ failure (MOF) was dose limiting on schedule B. The overall complete (CR) and partial (PR) response rate in patients treated at or below the MTD was 17% (4/23, 2 CRs, 2 PRs) for schedule A and 10% (2/19, 2 PRs) for schedule B. Pevonedistat plasma concentrations peaked after infusion followed by elimination in a biphasic pattern. Pharmacodynamic studies of biological correlates of NAE inhibition demonstrated target-specific activity of pevonedistat. In conclusion, administration of the first-in-class agent, pevonedistat, was feasible in patients with MDS and AML and modest clinical activity was observed.

Topics: Adult; Aged; Aged, 80 and over; Chemical and Drug Induced Liver Injury; Cyclopentanes; Enzyme Inhibitors; Female; Humans; Leukemia, Myeloid, Acute; Male; Maximum Tolerated Dose; Middle Aged; Multiple Organ Failure; Myelodysplastic Syndromes; Pyrimidines; Ubiquitin-Activating Enzymes

Targeted inhibitors of JAK2 (eg ruxolitinib) often provide symptomatic relief for myeloproliferative neoplasm (MPN) patients, but the malignant clone persists and remains susceptible to disease transformation. These observations suggest that targeting alternative dysregulated signaling pathways may provide therapeutic benefit. Previous studies identified NFκB pathway hyperactivation in myelofibrosis (MF) and secondary acute myeloid leukemia (sAML) that was insensitive to JAK2 inhibition. Here, we provide evidence that NFκB pathway inhibition via pevonedistat targets malignant cells in MPN patient samples as well as in MPN and patient-derived xenograft mouse models that are nonredundant with ruxolitinib. Colony forming assays revealed preferential inhibition of MF colony growth compared with normal colony formation. In mass cytometry studies, pevonedistat blunted canonical TNFα responses in MF and sAML patient CD34+ cells. Pevonedistat also inhibited hyperproduction of inflammatory cytokines more effectively than ruxolitinib. Upon pevonedistat treatment alone or in combination with ruxolitinib, MPN mouse models exhibited reduced disease burden and improved survival. These studies demonstrating efficacy of pevonedistat in MPN cells in vitro as well as in vivo provide a rationale for therapeutic inhibition of NFκB signaling for MF treatment. Based on these findings, a Phase 1 clinical trial combining pevonedistat with ruxolitinib has been initiated.

Topics: Animals; Cyclopentanes; Humans; Leukemia, Myeloid, Acute; Mice; Myeloproliferative Disorders; Primary Myelofibrosis; Pyrimidines

Dysregulation of apoptotic machinery is one mechanism by which acute myeloid leukemia (AML) acquires a clonal survival advantage. B-cell lymphoma protein-2 (BCL2) overexpression is a common feature in hematologic malignancies. The selective BCL2 inhibitor, venetoclax (VEN) is used in combination with azacitidine (AZA), a DNAmethyltransferase inhibitor (DNMTi), to treat patients with AML. Despite promising response rates to VEN/AZA, resistance to the agent is common. One identified mechanism of resistance is the upregulation of myeloid cell leukemia-1 protein (MCL1). Pevonedistat (PEV), a novel agent that inhibits NEDD8-activating enzyme, and AZA both upregulate NOXA (PMAIP1), a BCL2 family protein that competes with effector molecules at the BH3 binding site of MCL1. We demonstrate that PEV/AZA combination induces NOXA to a greater degree than either PEV or AZA alone, which enhances VEN-mediated apoptosis. Herein, using AML cell lines and primary AML patient samples ex vivo, including in cells with genetic alterations linked to treatment resistance, we demonstrate robust activity of the PEV/VEN/AZA triplet. These findings were corroborated in preclinical systemic engrafted models of AML. Collectively, these results provide rational for combining PEV/VEN/AZA as a novel therapeutic approach in overcoming AML resistance in current therapies.

Topics: Azacitidine; Bridged Bicyclo Compounds, Heterocyclic; Cyclopentanes; Humans; Leukemia, Myeloid, Acute; Pyrimidines; Sulfonamides

Treatment of acute myeloid leukemia (AML) remains challenging and novel targets and synergistic therapies still need to be discovered. We performed a high-throughput RNAi screen in three different AML cell lines and primary human leukemic blasts to identify genes that synergize with common antileukemic therapies. We used a pooled shRNA library that covered 5043 different genes and combined transfection with exposure to either azacytidine or cytarabine analog to the concept of synthetic lethality. Suppression of the chemokine CXCL12 ranked highly among the candidates of the cytarabine group. Azacytidine in combination with suppression of genes within the neddylation pathway led to synergistic results. NEDD8 and RBX1 inhibition by the small molecule inhibitor pevonedistat inhibited leukemia cell growth. These findings establish an in vitro synergism between NEDD8 inhibition and azacytidine in AML. Taken together, neddylation constitutes a suitable target pathway for azacytidine combination strategies.

Topics: Azacitidine; Chemokine CXCL12; Combined Modality Therapy; Cyclopentanes; HL-60 Cells; Humans; Leukemia, Myeloid, Acute; NEDD8 Protein; Pre-B-Cell Leukemia Transcription Factor 1; Pyrimidines; RNA Interference; Signal Transduction

Topics: Azacitidine; Cyclopentanes; Enzyme Inhibitors; Humans; Leukemia, Myeloid, Acute; NEDD8 Protein; Pyrimidines

Current understanding suggests that malignant stem and progenitor cells must be reduced or eliminated for prolonged remissions in myeloid neoplasms such as acute myelogenous leukemia (AML) or myelodysplastic syndrome (MDS). Multicolor flow cytometry has been widely used to distinguish stem and myeloid progenitor cells from other populations in normal and malignant bone marrow. In this study, we present a method for assessing drug sensitivity in MDS and AML patient hematopoietic stem and myeloid progenitor cell populations ex vivo using the investigational Nedd8-activating enzyme inhibitor MLN4924 and standard-of-care agent cytarabine as examples. Utilizing a multicolor flow cytometry antibody panel for identification of hematopoietic stem cells, multipotent progenitors, common myeloid progenitors, granulocyte-monocyte progenitors, and megakaryocyte-erythroid progenitors present in mononuclear cell fractions isolated from bone marrow aspirates, we compare stem and progenitor cell counts after treatment for 24 hours with drug versus diluent. We demonstrate that MLN4924 exerts a cytotoxic effect on MDS and AML stem and progenitor cell populations, whereas cytarabine has more limited effects. Further application of this method for evaluating drug effects on these populations ex vivo and in vivo may inform rational design and selection of therapies in the clinical setting. Stem Cells Translational Medicine 2017;6:840-850.

Topics: Cell Count; Cell Death; Cell Survival; Cyclopentanes; Cytarabine; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid, Acute; Myelodysplastic Syndromes; Pyrimidines

Topics: Azacitidine; Cell Line, Tumor; Cyclopentanes; Humans; Leukemia, Myeloid, Acute; Neoplasm Proteins; Proteomics; Pyrimidines

Two classes of novel agents, NEDD8-activating enzyme (NAE) and histone deacetylase (HDAC) inhibitors, have shown single-agent activity in acute myelogenous leukemia (AML)/myelodysplastic syndrome (MDS). Here we examined mechanisms underlying interactions between the NAE inhibitor pevonedistat (MLN4924) and the approved HDAC inhibitor belinostat in AML/MDS cells. MLN4924/belinostat coadministration synergistically induced AML cell apoptosis with or without p53 deficiency or FLT3-internal tandem duplication (ITD), whereas p53 short hairpin RNA (shRNA) knockdown or enforced FLT3-ITD expression significantly sensitized cells to the regimen. MLN4924 blocked belinostat-induced antiapoptotic gene expression through nuclear factor-κB inactivation. Each agent upregulated Bim, and Bim knockdown significantly attenuated apoptosis. Microarrays revealed distinct DNA damage response (DDR) genetic profiles between individual vs combined MLN4924/belinostat exposure. Whereas belinostat abrogated the MLN4924-activated intra-S checkpoint through Chk1 and Wee1 inhibition/downregulation, cotreatment downregulated multiple homologous recombination and nonhomologous end-joining repair proteins, triggering robust double-stranded breaks, chromatin pulverization, and apoptosis. Consistently, Chk1 or Wee1 shRNA knockdown significantly sensitized AML cells to MLN4924. MLN4924/belinostat displayed activity against primary AML or MDS cells, including those carrying next-generation sequencing-defined poor-prognostic cancer hotspot mutations, and CD34(+)/CD38(-)/CD123(+) populations, but not normal CD34(+) progenitors. Finally, combined treatment markedly reduced tumor burden and significantly prolonged animal survival (P < .0001) in AML xenograft models with negligible toxicity, accompanied by pharmacodynamic effects observed in vitro. Collectively, these findings argue that MLN4924 and belinostat interact synergistically by reciprocally disabling the DDR in AML/MDS cells. This strategy warrants further consideration in AML/MDS, particularly in disease with unfavorable genetic aberrations.

Topics: Animals; Apoptosis; Bcl-2-Like Protein 11; Cell Cycle Proteins; Cells, Cultured; Checkpoint Kinase 1; Cyclopentanes; DNA Damage; DNA Repair; Drug Synergism; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Kaplan-Meier Estimate; Leukemia, Myeloid, Acute; Mice; Molecular Targeted Therapy; Myelodysplastic Syndromes; Neoplasm Proteins; NF-kappa B; Nuclear Proteins; Protein Processing, Post-Translational; Protein-Tyrosine Kinases; Pyrimidines; RNA Interference; RNA, Small Interfering; S Phase Cell Cycle Checkpoints; Sulfonamides; U937 Cells; Ubiquitin-Activating Enzymes; Xenograft Model Antitumor Assays

High levels of microRNA-155 (miR-155) are associated with poor outcome in acute myeloid leukemia (AML). In AML, miR-155 is regulated by NF-κB, the activity of which is, in part, controlled by the NEDD8-dependent ubiquitin ligases. We demonstrate that MLN4924, an inhibitor of NEDD8-activating enzyme presently being evaluated in clinical trials, decreases binding of NF-κB to the miR-155 promoter and downregulates miR-155 in AML cells. This results in the upregulation of the miR-155 targets SHIP1, an inhibitor of the PI3K/Akt pathway, and PU.1, a transcription factor important for myeloid differentiation, leading to monocytic differentiation and apoptosis. Consistent with these results, overexpression of miR-155 diminishes MLN4924-induced antileukemic effects. In vivo, MLN4924 reduces miR-155 expression and prolongs the survival of mice engrafted with leukemic cells. Our study demonstrates the potential of miR-155 as a novel therapeutic target in AML via pharmacologic interference with NF-κB-dependent regulatory mechanisms. We show the targeting of this oncogenic microRNA with MLN4924, a compound presently being evaluated in clinical trials in AML. As high miR-155 levels have been consistently associated with aggressive clinical phenotypes, our work opens new avenues for microRNA-targeting therapeutic approaches to leukemia and cancer patients.

Topics: Animals; Apoptosis; Blotting, Western; Cell Differentiation; Cell Proliferation; Chromatin Immunoprecipitation; Cyclopentanes; Drug Resistance, Neoplasm; Female; fms-Like Tyrosine Kinase 3; Gene Expression Regulation, Leukemic; Humans; Leukemia, Myeloid, Acute; Mice; Mice, Inbred NOD; Mice, SCID; MicroRNAs; Monocytes; NEDD8 Protein; NF-kappa B; Promoter Regions, Genetic; Pyrimidines; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Tandem Repeat Sequences; Tumor Cells, Cultured; Ubiquitins; Xenograft Model Antitumor Assays

MLN4924 (pevonedistat), an inhibitor of the Nedd8 activating enzyme (NAE), has exhibited promising clinical activity in acute myelogenous leukemia (AML). Here we demonstrate that MLN4924 induces apoptosis in AML cell lines and clinical samples via a mechanism distinct from those observed in other malignancies. Inactivation of E3 cullin ring ligases (CRLs) through NAE inhibition causes accumulation of the CRL substrate c-Myc, which transactivates the PMAIP1 gene encoding Noxa, leading to increased Noxa protein, Bax and Bak activation, and subsequent apoptotic changes. Importantly, c-Myc knockdown diminishes Noxa induction; and Noxa siRNA diminishes MLN4924-induced killing. Because Noxa also neutralizes Mcl-1, an anti-apoptotic Bcl-2 paralog often upregulated in resistant AML, further experiments have examined the effect of combining MLN4924 with BH3 mimetics that target other anti-apoptotic proteins. In combination with ABT-199 or ABT-263 (navitoclax), MLN4924 exerts a synergistic cytotoxic effect. Collectively, these results provide new insight into MLN4924-induced engagement of the apoptotic machinery that could help guide further exploration of MLN4924 for AML.

Topics: Antineoplastic Agents; Apoptosis; Bridged Bicyclo Compounds, Heterocyclic; Cell Line, Tumor; Cyclopentanes; Drug Synergism; HL-60 Cells; Humans; Leukemia, Myeloid, Acute; Myeloid Cell Leukemia Sequence 1 Protein; Proto-Oncogene Proteins c-bcl-2; Proto-Oncogene Proteins c-myc; Pyrimidines; RNA Interference; RNA, Small Interfering; Sulfonamides; Up-Regulation

Sensitive to Apoptosis Gene (SAG), also known as RBX2 (RING box protein-2), is the RING component of SCF (SKP1, Cullin, and F-box protein) E3 ubiquitin ligase. Our previous studies have demonstrated that SAG is an anti-apoptotic protein and an attractive anti-cancer target. We also found recently that Sag knockout sensitized mouse embryonic stem cells (mES) to radiation and blocked mES cells to undergo endothelial differentiation. Here, we reported that compared to wild-type mES cells, the Sag(-/-) mES cells were much more sensitive to all-trans retinoic acid (RA)-induced suppression of cell proliferation and survival. While wild-type mES cells underwent differentiation upon exposure to RA, Sag(-/-) mES cells were induced to death via apoptosis instead. The cell fate change, reflected by cellular stiffness, can be detected as early as 12 hrs post RA exposure by AFM (Atomic Force Microscopy). We then extended this novel finding to RA differentiation therapy of leukemia, in which the resistance often develops, by testing our hypothesis that SAG inhibition would sensitize leukemia to RA. Indeed, we found a direct correlation between SAG overexpression and RA resistance in multiple leukemia lines. By using MLN4924, a small molecule inhibitor of NEDD8-Activating Enzyme (NAE), that inactivates SAG-SCF E3 ligase by blocking cullin neddylation, we were able to sensitize two otherwise resistant leukemia cell lines, HL-60 and KG-1 to RA. Mechanistically, RA sensitization by MLN4924 was mediated via enhanced apoptosis, likely through accumulation of pro-apoptotic proteins NOXA and c-JUN, two well-known substrates of SAG-SCF E3 ligase. Taken together, our study provides the proof-of-concept evidence for effective treatment of leukemia patients by RA-MLN4924 combination.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Differentiation; Cell Line, Tumor; Cyclopentanes; Drug Interactions; Embryonic Stem Cells; Enzyme Activation; Enzyme Inhibitors; Female; Gene Deletion; Humans; JNK Mitogen-Activated Protein Kinases; Leukemia, Myeloid, Acute; Male; Mice; Proto-Oncogene Proteins c-bcl-2; Pyrimidines; Tretinoin; Ubiquitin-Protein Ligases

NEDD8 activating enzyme (NAE) has been identified as an essential regulator of the NEDD8 conjugation pathway, which controls the degradation of many proteins with important roles in cell-cycle progression, DNA damage, and stress responses. Here we report that MLN4924, a novel inhibitor of NAE, has potent activity in acute myeloid leukemia (AML) models. MLN4924 induced cell death in AML cell lines and primary patient specimens independent of Fms-like tyrosine kinase 3 expression and stromal-mediated survival signaling and led to the stabilization of key NAE targets, inhibition of nuclear factor-kappaB activity, DNA damage, and reactive oxygen species generation. Disruption of cellular redox status was shown to be a key event in MLN4924-induced apoptosis. Administration of MLN4924 to mice bearing AML xenografts led to stable disease regression and inhibition of NEDDylated cullins. Our findings indicate that MLN4924 is a highly promising novel agent that has advanced into clinical trials for the treatment of AML.

Topics: Animals; Blotting, Western; Cell Line, Tumor; Cullin Proteins; Cyclopentanes; DNA Damage; Fluorescent Antibody Technique; fms-Like Tyrosine Kinase 3; Humans; Leukemia, Myeloid, Acute; Mice; NEDD8 Protein; NF-kappa B; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Pyrimidines; Reactive Oxygen Species; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Ubiquitins; Xenograft Model Antitumor Assays