Kooperativer Bibliotheksverbund

In: Diabetologia, Springer Science and Business Media LLC, Vol. 47, No. S1 ( 2004-8), p. A1-A464

Type of Medium: Online Resource

ISSN: 0012-186X , 1432-0428

URL: Article

DOI: 10.1007/BF03375463

Language: English

Publisher: Springer Science and Business Media LLC

Publication Date: 2004

detail.hit.zdb_id: 1458993-X

In: Journal of Clinical Oncology, American Society of Clinical Oncology (ASCO), Vol. 34, No. 34 ( 2016-12-01), p. 4086-4093

Abstract: Few treatment options exist for adult T-cell leukemia/lymphoma (ATL), and the prognosis for this disease is poor. A phase I study of lenalidomide demonstrated preliminary antitumor activity in patients with relapsed ATL. The current phase II study evaluated the efficacy and safety of lenalidomide monotherapy in patients with relapsed or recurrent ATL. Patients and Methods Patients 20 years of age or older with acute, lymphoma, or unfavorable chronic subtype ATL, who had received one or more prior anti-ATL systemic chemotherapy and achieved stable disease or better on their last anti-ATL therapy with subsequent relapse or recurrence, were eligible. Patients received oral lenalidomide 25 mg/d continuously until disease progression or unacceptable toxicity. The primary end point was overall response rate; secondary end points included safety, tumor control rate (stable disease or better), time to response, duration of response, time to progression, progression-free survival, and overall survival. Results Objective responses were noted in 11 of 26 patients (overall response rate, 42%; 95% CI, 23% to 63%), including four complete responses and one unconfirmed complete response. The tumor control rate was 73%. The median time to response and duration of response were 1.9 months and not estimable, respectively, and the median time to progression was 3.8 months. The median progression-free survival and overall survival were 3.8 and 20.3 months, respectively. The most frequent grade ≥ 3 adverse events were neutropenia (65%), leukopenia (38%), lymphopenia (38%), and thrombocytopenia (23%), which were all manageable and reversible. Conclusion Lenalidomide demonstrated clinically meaningful antitumor activity and an acceptable toxicity profile in patients with relapsed or recurrent aggressive ATL, hinting at its potential to become a treatment option. Further investigations of lenalidomide in ATL and other mature T-cell neoplasms are warranted.

Type of Medium: Online Resource

ISSN: 0732-183X , 1527-7755

URL: Article

DOI: 10.1200/JCO.2016.67.7732

Language: English

Publisher: American Society of Clinical Oncology (ASCO)

Publication Date: 2016

detail.hit.zdb_id: 2005181-5

In: Journal of Clinical Investigation, American Society for Clinical Investigation, Vol. 129, No. 5 ( 2019-4-2), p. 1878-1894

Type of Medium: Online Resource

ISSN: 0021-9738 , 1558-8238

URL: Article

URL: Article

DOI: 10.1172/JCI120654

DOI: 10.1172/JCI120654DS1

Language: English

Publisher: American Society for Clinical Investigation

Publication Date: 2019

detail.hit.zdb_id: 2018375-6

In: Blood, American Society of Hematology, Vol. 140, No. Supplement 1 ( 2022-11-15), p. 8785-8786

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood-2022-162354

Language: English

Publisher: American Society of Hematology

Publication Date: 2022

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 138, No. Supplement 1 ( 2021-11-05), p. 505-505

Abstract: The BCL-2 inhibitor venetoclax (ven) in combination with hypomethylating agents (HMA) has revolutionized acute myeloid leukemia (AML) therapy. However, the majority of patients with AML who received ven/HMA eventually relapse and cures are still elusive. We previously generated ven-resistant MV4;11 (MV4;11 VR) cells and found them to express elevated c-Myc protein levels. The dual inhibition of MDM2 and XPO1 significantly increased nuclear p53 protein and dramatically upregulated p53 target genes. On the other hand, dual inhibition of MDM2 and XPO1 profoundly decreased c-Myc protein levels in a p53-dependent manner, resulting in the profound downregulation of MYC transcriptional program. Clinical grade MDM2 and XPO1 inhibitors milademetan (mil) and selinexor (sel) significantly reduced leukemia burden and prolonged survival in the xenograft model injected with MV4;11 VR cells. However, the treatment effect did not result in very long relapse free responses. To investigate potential resistance mechanisms to dual inhibition of MDM2 and XPO1, we utilized our previously reported (Muftuoglu, ASH 2020) multiparameter flow cytometry to assess multiple stress responses pathways including p21, ATF4, PARP, LC3B, Ki-67, active caspase-3 and amine-reactive viability dye at the single cell level in ven-resistant AML cells. We found that residual cells, after combined MDM2 and XPO1 inhibition, expressed high levels of p21, ATF4 and LC3B and low Ki-67 levels, suggesting that the resistant population is in a cell kinetic quiescent state with activation of autophagy and ER stress. Interestingly, ven-resistant MV4;11 cells with in vivo acquired resistance to dual inhibition of MDM2 and XPO1 demonstrated elevated protein levels of respiratory chain complex proteins (NDUFB8, MTCO1, UQCRC2 and ATP5A), suggesting increased OXPHOS dependency. Intriguingly, these MV4;11VR cells with subsequently acquired resistance to MDM2 and XPO1 inhibition in vivo have restored sensitivity to ven. We previously reported that the combination of MDM2 and BCL-2 inhibitors induces synergistic apoptosis through the elimination of dormant p21 high residual AML cell (Pan, Cancer Cell 2017). In a clinical trial of Idasanutlin + Venetoclax we observed over 45% response rates in relapsed/refractory AML patients (Daver, ASH 2020). Therefore, we hypothesized that combining MDM2/XPO1 inhibition with BCL-2 inhibition further induces cell killing in ven/HMA resistant AML cells. To address this, we treated ven-resistant AML cells with triple combination of mil, sel and ven, resulting in a profound cytoreduction in vitro compared with other single and doublet treatments. Next, we treated NSG mice injected with PDX AML cells with FLT3-ITD, GATA2 and NRAS mutations obtained from a patient who relapsed after ven/decitabine treatment. The doublet combinations especially of mil + ven reduced circulating blasts and significantly prolonged survival. Of note, no mice have died at day 180 after treatment in the group receiving triple combination of mil, ven and sel, with profound and sustained cytoreduction (more than 2 log 10 difference) and significantly prolonged survival compared with any other treatment (Fig. A, B). The triple combination was well-tolerated and did not decrease mouse CD45+ cells, platelets and hemoglobin. In conclusion, the concomitant inhibition of MDM2, XPO1 and BCL-2 was feasible and exerted dramatic and sustained anti-leukemia activity in ven/HMA resistant AML cell in vitro and in vivo. Milademetan (RAIN-32) is currently being developed by Rain Therapeutics. Figure 1 Figure 1. Disclosures Carter: Syndax: Research Funding; Ascentage: Research Funding. Daver: Pfizer: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy, Other: Data Monitoring Committee member; Novartis: Consultancy; Gilead Sciences, Inc.: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; ImmunoGen: Consultancy, Research Funding; Trillium: Consultancy, Research Funding; Glycomimetics: Research Funding; Sevier: Consultancy, Research Funding; Astellas: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Abbvie: Consultancy, Research Funding; Novimmune: Research Funding; Hanmi: Research Funding; Amgen: Consultancy, Research Funding; Trovagene: Consultancy, Research Funding; FATE Therapeutics: Research Funding; Dava Oncology (Arog): Consultancy; Celgene: Consultancy; Syndax: Consultancy; Shattuck Labs: Consultancy; Agios: Consultancy; Kite Pharmaceuticals: Consultancy; SOBI: Consultancy; STAR Therapeutics: Consultancy; Karyopharm: Research Funding; Newave: Research Funding. Lesegretain: Daiichi-Sankyo Inc.: Current Employment. Seki: Daiichi-Sankyo Inc.: Current Employment. Shacham: Karyopharm: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties: (8999996, 9079865, 9714226, PCT/US12/048319, and I574957) on hydrazide containing nuclear transport modulators and uses, and pending patents PCT/US12/048319, 499/2012, PI20102724, and 2012000928) . Konopleva: Ascentage: Other: grant support, Research Funding; Reata Pharmaceuticals: Current holder of stock options in a privately-held company, Patents & Royalties: intellectual property rights; Eli Lilly: Patents & Royalties: intellectual property rights, Research Funding; Novartis: Other: research funding pending, Patents & Royalties: intellectual property rights; AstraZeneca: Other: grant support, Research Funding; Rafael Pharmaceuticals: Other: grant support, Research Funding; F. Hoffmann-La Roche: Consultancy, Honoraria, Other: grant support; AbbVie: Consultancy, Honoraria, Other: Grant Support, Research Funding; Genentech: Consultancy, Honoraria, Other: grant support, Research Funding; Forty Seven: Other: grant support, Research Funding; Cellectis: Other: grant support; Calithera: Other: grant support, Research Funding; Agios: Other: grant support, Research Funding; KisoJi: Research Funding; Ablynx: Other: grant support, Research Funding; Stemline Therapeutics: Research Funding; Sanofi: Other: grant support, Research Funding. Andreeff: Medicxi: Consultancy; Glycomimetics: Consultancy; AstraZeneca: Research Funding; Syndax: Consultancy; Amgen: Research Funding; Oxford Biomedica UK: Research Funding; Karyopharm: Research Funding; Daiichi-Sankyo: Consultancy, Research Funding; Breast Cancer Research Foundation: Research Funding; Novartis, Cancer UK; Leukemia & Lymphoma Society (LLS), German Research Council; NCI-RDCRN (Rare Disease Clin Network), CLL Foundation; Novartis: Membership on an entity's Board of Directors or advisory committees; Senti-Bio: Consultancy; Reata, Aptose, Eutropics, SentiBio; Chimerix, Oncolyze: Current holder of individual stocks in a privately-held company; ONO Pharmaceuticals: Research Funding; Aptose: Consultancy.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood-2021-149260

Language: English

Publisher: American Society of Hematology

Publication Date: 2021

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 126, No. 23 ( 2015-12-03), p. 1266-1266

Abstract: Internal tandem duplication (ITD) mutations of the fms-like tyrosine kinase-3 (FLT3) gene are common in patients with acute myeloid leukemia (AML) and associated with poor disease prognosis (Kottaridis et al., 2003). FLT3-targeted therapies using novel small-molecule inhibitors such as sorafenib are under development with some notable effects (Adnane et al., 2005). However, acquired resistance to these agents is frequent following long-term therapy. One of the main resistance mechanisms is the acquisition of point mutations in the tyrosine kinase (TK) domains (TKD) ofFLT3, in addition to ITD mutations (Smith et al., 2012; Zhang et al., 2014). Another novel target for AML therapy is exportin-1 (XPO1), one of the major transporters of proteins and mRNAs from the nucleus. High expression of XPO1 is associated with decreased overall survival in AML patients (Kojima et al., 2013). and its levels were increased in patients with FLT3-ITD mutations. Interestingly, targeting CRM1 with the small molecule inhibitor KPT-185 down-regulated FLT3 expression (Ranganathan et al., 2012). The XPO1 inhibitor selinexor (PKT-330) has been developed recently and has shown encouraging anti-tumor effects against AML-initiating cells in primary human AML-engrafted NSG mice (Etchin et al., 2015) and in ongoing clinical trials. In this study, we investigated the anti-tumor activity of combined treatment with sorafenib and selinexor in FLT3-mutant AML cells, including those with ITD and TKD point mutations, (Smith et al., 2012; Zhang et al., 2014). Selinexor alone exerted marked cell killing in human and murineFLT3-mutant AML cells, including those harboring D835Y or ITD with Y842C or F691L mutations. Interestingly, FLT3 and MAPK or FLT3 and AKT signaling pathways were activated by the selinexor treatment for 24 h in murine FLT3 -mutant AML cells. Combination treatment with selinexor and sorafenib demonstrated synergistic or additive pro-apoptotic effects in TKD point- and FLT3-ITD-mutanted AML cells, respectively. Mechanistically, the combination treatment triggered significant suppression of phospho-ERK and phospho-AKT compared to selinexor treatment alone in all examined AML cell lines. Of note, the combination treatment promoted the retention of ERK, AKT, NFκB, and FOXO3a the in nucleus, which could explain the low levels of phospho-ERK and phospho-AKT observed in the cytosol. In addition, a 5-day in vitro combination treatment with extremely low 5 to 10 nM concentrations of each agent promoted myeloid differentiation of FLT3-ITD mutant MOLM14 cells. No cell killing was observed using this treatment scenario. Furthermore, the combination treatment demonstrated profound anti-tumor efficacy in a human FLT3-mutated leukemia model in NOG mice. The median survival was doubled from 16 days in the vehicle-treated mice to 31 or 23 days with selinexor or sorafenib treatments, respectively, and this effect was increased to 51 days in the mice that received the combination treatment (p = 0.0001). Our findings provide a pre-clinical basis for a combinatorial treatment strategy targeting XPO1 and FLT3-ITD in FLT3 -mutated AML patients, especially in those patients who have acquired resistance to FLT3-targeted therapy. Disclosures Zhang: Karyopharm: Research Funding. Ishizawa:Karyopharm: Research Funding. Daver:ImmunoGen: Other: clinical trial, Research Funding.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood.V126.23.1266.1266

Language: English

Publisher: American Society of Hematology

Publication Date: 2015

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 126, No. 23 ( 2015-12-03), p. 2051-2051

Abstract: The clinical challenge posed by p53-deficiency in hematological malignancies needs novel therapeutic strategies. ONC201, a first-in-class small molecule, was discovered as a p53-independent activator of apoptosis with a benign preclinical safety profile (Allen et al, Sci Transl Med 2013). Here we report that ONC201 exerts anti-tumor effects in hematological malignancies and leukemia stem cells (LSC) via the induction of ATF4 in the integrated stress response (ISR). ONC201 induced p53-independent apoptosis in cell lines and primary patient samples from mantle cell lymphomas (MCL) and acute myeloid leukemias (AML), independent of genetic alterations that correlate with poor prognosis (e.g., TP53 mutation, FLT3-ITD, or complex karyotype). ONC201 also induced apoptosis in LSC, while sparing normal bone marrow progenitors, as measured in vitro and after transplantation. Specifically, we recovered unfractionated LSC-containing populations of AML cells (t(9;11)(p22; q23), CEBPA and ATM mutant) from secondarily-engrafted mice and cultured them in vitro for 48 hr in two groups, ONC201-treated (5 uM) or Control. For both groups, the same number of Trypan Blue-negative cells was then re-transplanted. The frequency of human CD45+ cells 4 weeks after transplantation was 38.09 ± 2.59 % of tibial BM cells in untreated mice and 0.10 ± 0.05% in treated mice (n = 3 for each, p 〈 0.01). The survival of the treated group was dramatically prolonged (Figure 1). Gene Set Enrichment Analysis (GSEA) of gene expression profiling (GEP) data of Jeko-1 and Z-138 cells treated with ONC201 implicated up-regulated Endoplasmic Reticulum (ER) stress-related genes, such as targets of the ER stress-induced transcription factor CHOP (DDIT3; FDR q = 0.016), and ER component proteins (FDR q = 0.039). We confirmed the ONC201-induced increased mRNA levels of DDIT3, GADD34, DR5 and TRIB3, and increased protein levels of CHOP, ATF4 and IRE1-a in Jeko-1 cells. Knockdown of ATF4 and IRE1-a revealed that ATF4 is an essential protein for ONC201-induced apoptosis in most AML cells and MCL cells, while IRE1-a is only necessary for apoptosis in MCL cells. However, unlike other ER stress inducers, ONC201 did not cause phosphorylation of an eIF2a kinase PERK, a hallmark of classical ER stress. Importantly, ONC201 was also effective in lymphoma cells resistant to bortezomib, an ER stress inducer, supporting the notion that ONC201 uses a unique mechanism to trigger the ISR. In addition, ONC201 inhibited mTORC1 signaling, likely secondary to ATF4 activation via the induction of DDIT4, a negative regulator of mTORC1, which presumably increases the cytotoxicity of ONC201 by global translation inhibition. Investigating the type of apoptosis induced by ONC201, we examined protein levels of anti-apoptotic BCL-2 family members (BCL-2, BCL-XL and MCL-1) after ONC201 treatment. MCL-1 was reduced most notably after 12 hr. We then tested the effect of ONC201 on cells with overexpression or knockdown of BCL-2 family proteins. MCL-1 knockdown in OCI-AML3 cells increased their sensitivity to ONC201 only slightly, but ONC201 efficacy was dramatically reduced in BCL-2-overexpressing HL-60 cells, even more so than in BCL-XL-overexpressing HL-60 cells. Therefore, we investigated whether ONC201 sensitivity could be increased by ABT-199, a small-molecule BH3 mimetic that specifically inhibits BCL-2, and is known to be ineffective in cell lines with MCL-1 overexpression; accordingly, the combination of ONC201 and BCL-2 antagonist ABT-199 was highly synergistic (Figure 2). In conclusion, ONC201 induces p53-independent apoptosis and abrogates LSC function by ATF4 induction, via the ISR. By suppressing MCL-1, ONC201 can also increase the effectiveness of the Bcl-2 inhibitor ABT-199. These findings suggest that ONC201 may provide promising novel therapeutic strategies for TP53 -wild type and TP53 -mutant hematological malignancies. Phase I/II clinical trials have been initiated at the MD Anderson Cancer Center in leukemias and lymphomas to determine safety, efficacy and further characterize mechanism of action. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Ishizawa: Karyopharm: Research Funding. Allen:Oncoceutics, Inc: Employment, Equity Ownership. Orlowski:Janssen Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Array BioPharma: Consultancy, Research Funding; Millennium Pharmaceuticals: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; BioTheryX, Inc.: Membership on an entity's Board of Directors or advisory committees; Acetylon: Membership on an entity's Board of Directors or advisory committees; Genentech: Consultancy; Forma Therapeutics: Consultancy; Onyx Pharmaceuticals: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Spectrum Pharmaceuticals: Research Funding. Wang:Celgene: Research Funding. Konopleva:Novartis: Research Funding; AbbVie: Research Funding; Stemline: Research Funding; Calithera: Research Funding; Threshold: Research Funding. Andreeff:Oncoceutics, Inc.: Membership on an entity's Board of Directors or advisory committees.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood.V126.23.2051.2051

Language: English

Publisher: American Society of Hematology

Publication Date: 2015

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 128, No. 22 ( 2016-12-02), p. 2893-2893

Abstract: Development of MDM2 inhibitors enabled successful induction of p53-mediated apoptosis in tumor cells without a risk of DNA damage. Early clinical trials of MDM2 inhibitors demonstrated proof-of-concept (Andreeff et al., Clin Can Res, 2015). However, a clinical challenge is that not all the tumors bearing wild-type TP53 are sensitive to MDM2 inhibition. We here discovered novel gene profiling-based algorithms for predicting tumor sensitivity to MDM2 inhibition, using DS-3032b, a novel potent MDM2 inhibitor, which is currently in early clinical trials. In vitro inhibitory effects of DS-3032b on MDM2-p53 interaction was demonstrated using the homogeneous time resolved fluorescence (HTRF) assay (IC50 5.57 nM). DS-3032b treatment (30-1000 nM) indeed increased p53 protein in a dose-dependent manner, and also the p53 targets MDM2 and p21, in cancer cell lines with wild-type TP53 (SJSA-1, MOLM-13, DOHH-2, and WM-115), showing around 10-fold potent growth inhibition effects compared to Nutlin-3a (Table 1). The xenograft mouse models with SJSA-1 and MOLM-13 cells showed 〉 90% reduction in tumor growth with oral administrations of 25 and 50 mg/kg/day. For discovering predictive gene signatures, we performed two different approaches. In the first approach, 240 cell lines available as OncoPanel were treated with DS-3032b, another prototypic MDM2 inhibitor DS-5272, and Nutlin-3a, and determined 62 sensitive and 164 resistant lines, based on GI50s. Using gene expression profiling (GEP) publicly available for all the cell lines, we selected 175 top-ranked genes with highest expression in the 62 sensitive cell lines. We thus defined the average of Z-scores of the 175 gene expression as "sensitivity score". To validate the 175-gene signature, we evaluated in vivo anti-tumor activities of DS-3032b in 13 patient-derived tumor xenografts (melanoma, NSCLC, colorectal and pancreatic cancers). The prediction accuracy, sensitivity, positive predictive value (PPV), and negative predictive value (NPV) were 85, 88, 88 and 80% respectively. As another validation set, 41 primary AML samples were treated with DS-3032b to define the top and bottom one-third most sensitive or resistant samples (14 each), and GEP was performed in every sample. TP53 mutations were detected in 8 specimens by next generation sequencing and confirmed by Sanger sequencing. The 175-gene signature was applied to the AML dataset, and the accuracy, sensitivity, PPV and NPV to predict the 14 sensitive or resistant samples were 79, 93, 72 and 90% respectively. Importantly, this signature was more predictive than the TP53 mutation status alone applied (68, 93, 62 and 86%). (Table 2A-B) In contrast to the cell line-based approach, the second approach defined an AML-specific gene signature. Specifically, we used the same dataset of 41 primary AML samples described above as training and validation set, by performing random forest methods with cross validation. Using a routine way in bioinformatics analysis of classifying gene signature, we first selected the 1500 top-ranked genes with highest expression variance among all the specimens. In addition, p53-related 32 genes that potentially have predictive values were also selected based on the previous reports. Classification was performed using the random forest method to identify a predictive algorithm with the 1500-gene set, 32-gene set or combined 1525-gene set (7 genes were overlapped), thus we found that the 1525-gene set had highest performance than each gene set alone. However, applying this method to all the 41 samples showed inferior predictive performance than applied only to the 33 wild-type TP53 samples (the prediction accuracy, sensitivity, PPV and NPV were 68, 72, 67 and 69%, vs. 77, 82, 75 and 80%).(Table 2C) Finally, we combined each of the two algorithms (Table 2B-C) with TP53 mutation status. Specifically, the samples with TP53 mutations were predicted as resistant, then either of gene signatures was applied to the rest of the samples with wild-type TP53. Predictive performance (Table 2D-E) was improved in both signatures compared to the others, especially showing the highest PPVs (80 and 82%, respectively). Taken together, gene signatures discovered in the present study, by combining with TP53 mutation status, provided new highly predictive algorithms for therapy of MDM2 inhibition. Our findings will be tested in ongoing clinical trials of DS-3032b. Disclosures Nakamaru: Daiichi Sankyo Co., Ltd: Employment. Seki:2Daiichi Sankyo Co., Ltd.: Employment. Tazaki:2Daiichi Sankyo Co., Ltd.: Employment. DiNardo:Celgene: Research Funding; Novartis: Other: advisory board, Research Funding; Abbvie: Research Funding; Agios: Other: advisory board, Research Funding; Daiichi Sankyo: Other: advisory board, Research Funding. Tse:Daiichi Sankyo, Inc.: Employment.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood.V128.22.2893.2893

Language: English

Publisher: American Society of Hematology

Publication Date: 2016

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 134, No. Supplement_1 ( 2019-11-13), p. 2556-2556

Abstract: Background. MDM2 is frequently overexpressed in acute myeloid leukemias (AML) and suppresses p53-mediated apoptosis while p53 mutations are relatively rare in AML. MDM2 inhibitors as a monotherapy have shown limited efficacy in clinical trials in AML (~25% response rate) (Andreeff, Clin Cancer Res 2015). XPO1 transports around 300 proteins, including p53 and other tumor suppressors, from the nucleus to the cytoplasm. Overexpression of XPO1 is associated with unfavorable outcomes in AML (Kojima, Blood 2013). p53 activation or XPO1 inhibition have been reported to decrease c-Myc protein levels through diverse mechanisms (Porter Mol Cell 2017 and Tabe PLoSOne 2015). Objective: We investigated anti-leukemia effect of dual MDM2 and XPO1 inhibition, with the intent to maximize the pro-apoptotic functions of p53, using the MDM2 inhibitor milademetan (Daiichi-Sankyo), and selinexor, a recently FDA-approved XPO1 inhibitor or its analog eltanexor (Karyopharm). Results: Treatment with milademetan and selinexor (1:1 molar ratio) induced synergistic apoptosis in AML cell lines with wild-type p53 (ED50, 89.3 ± 18.6 nM, combination index (CI), 0.60 ± 0.08). Activity in p53 mutant AML required 40-fold higher ED50 (3572 ± 1986 nM), reflected in an antagonistic CI of 6.94 ± 3.06. Knockdown of wild-type p53 by shRNA in OCI-AML3 (OCI-AML3 shp53) cells or presence of TP53 mutation (p.R248W) in MOLM-13 cells eliminated the synergistic effects, suggesting that normal p53 function is a major determinant of sensitivity to combined treatment. Next, we treated primary AML samples with milademetan and selinexor or eltanexor and observed that effects were mutation-agnostic (e.g. RAS and FLT3) except for TP53. Combined treatment significantly reduced AUC determined by absolute live cell numbers compared to each drug alone, and induced synergistic apoptosis in primary AML samples with wild-type p53 (ED50 values, 27.2 - 937.4 nM, CI, 0.51 ± 0.07), with similar efficacies in complex and non-complex karyotype AMLs (279.6 ± 94.7 vs 256.6 ± 56.4 nM, P = 0.84). In contrast, combined treatment showed antagonistic effects in primary AML samples with loss-of-function TP53 mutations (CI 〉 1.0). Immature CD34+CD38- AML cells were more susceptible to combined treatment than CD34- AML cells (apoptosis induction, 76.2 ± 6.7% vs 47.5 ± 6.8%, P = 0.0002) Mechanistically, combined inhibition increased p53 protein levels and accumulated p53 but not MDM2 protein in the nucleus compared to each drug alone. Combined treatment induced more TP53 target genes (MDM2, CDKN1A, BBC3, FAS and Bax) in OCI-AML3 cells with control shRNA compared with OCI-AML3 shp53 cells. Combinatorial inhibition showed much enhanced reduction of c-Myc mRNA and protein levels in OCI-AML3 shC cells compared with OCI-AML3 shp53 cells (82% vs 32%). In confirmation, combined inhibition reduced c-Myc protein levels profoundly in wild-type p53 primary AMLs (ANOVA P 〈 0.0001). In contrast, c-Myc reduction was not observed in primary AMLs with p53-inactivating mutations. Intriguingly, OCI-AML3 cells overexpressing c-Myc by lentiviral transduction showed greater sensitivity to XPO1 inhibitors and the combination compared to empty-vector controls, and baseline levels of c-Myc protein also negatively correlated with ED50 for combined treatment in primary AML samples (Spearman R = -0.5357, P = 0.0422). Conclusion: These preclinical data suggest that dual inhibition of MDM2 and XPO1 induces synergistic apoptosis through accumulation of nuclear p53 and suppression of c-Myc in wild-type p53 AMLs. A clinical trial testing this concept in AML is under development. Disclosures Ishizawa: Daiichi Sankyo: Patents & Royalties: Joint submission with Daiichi Sankyo for a PTC patent titled "Predictive Gene Signature in Acute Myeloid Leukemia for Therapy with the MDM2 Inhibitor DS-3032b," United States, 62/245667, 10/23/2015, Filed. Daver:Novartis: Consultancy, Research Funding; Agios: Consultancy; Jazz: Consultancy; Hanmi Pharm Co., Ltd.: Research Funding; Pfizer: Consultancy, Research Funding; Astellas: Consultancy; Immunogen: Consultancy, Research Funding; Forty-Seven: Consultancy; Abbvie: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Servier: Research Funding; Incyte: Consultancy, Research Funding; NOHLA: Research Funding; Glycomimetics: Research Funding; BMS: Consultancy, Research Funding; Karyopharm: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Sunesis: Consultancy, Research Funding; Celgene: Consultancy; Otsuka: Consultancy. Lesegretain:Daiichi-Sankyo Inc.: Employment, Equity Ownership. Shacham:Karyopharm Therapeutics Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Andreeff:NIH/NCI: Research Funding; Center for Drug Research & Development: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy; Amgen: Consultancy; AstaZeneca: Consultancy; 6 Dimensions Capital: Consultancy; Reata: Equity Ownership; Aptose: Equity Ownership; Eutropics: Equity Ownership; Senti Bio: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Oncoceutics: Equity Ownership; Oncolyze: Equity Ownership; Breast Cancer Research Foundation: Research Funding; CPRIT: Research Funding; NCI-RDCRN (Rare Disease Cliln Network): Membership on an entity's Board of Directors or advisory committees; CLL Foundation: Membership on an entity's Board of Directors or advisory committees; BiolineRx: Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo, Inc.: Consultancy, Patents & Royalties: Patents licensed, royalty bearing, Research Funding; Leukemia Lymphoma Society: Membership on an entity's Board of Directors or advisory committees; Cancer UK: Membership on an entity's Board of Directors or advisory committees; NCI-CTEP: Membership on an entity's Board of Directors or advisory committees; German Research Council: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Consultancy.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood-2019-126950

Language: English

Publisher: American Society of Hematology

Publication Date: 2019

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7

In: Blood, American Society of Hematology, Vol. 124, No. 21 ( 2014-12-06), p. 3617-3617

Abstract: A minor fraction of leukemia cells, leukemia stem cells, have been shown to be highly resistant to current therapies and thought to be responsible for recurrence. BMI-1, a part of polycomb repressive complex 1 (PRC1) is essential for the self-renewal of normal hematopoietic and leukemia stem cells. PTC-209 is a novel selective transcriptional inhibitor of BMI-1, which has been shown to have antitumor activity against cancer-initiating cells in colorectal cancer. We investigated the prognostic significance of BMI-1 in acute myeloid leukemia (AML) using reversed phase protein array and effects of the BMI-1 inhibitor PTC-209 on primary and leukemia cell lines. BMI-1 protein expression was determined in bulk AML blasts from 511 newly diagnosed patients. BMI-1 expression was higher in unfavorable cytogenetics (n=252, median 0.068) compared to intermediate (n=225, median -0.116, P = 0.017) or favorable cytogenetics (n=34, median -0.338, P = 0.0007 versus unfavorable, 0.05 versus intermediate). Higher BMI-1 levels were associated with shorter median overall survival (42.8 versus 55.3 weeks, P = 0.046 Log Rank test). There was no correlation between BMI-1 levels and percentages of CD34 -positive cells (r = 0.07). A total of 6 AML (MOLM-13, OCI-AML3, MV4-11, NB4, HL60 and U-937) and 5 ALL (Reh, NALM6, Jurkat, Raji and MOLT-4) cell lines were exposed to PTC-209 for 48 hours. PTC-209 exhibited dose- and time-dependent anti-proliferative and cytotoxic activities. The IC50 values (concentration at which cell growth is inhibited by 50% at 48 hours of exposure) were 0.33 ± 0.04 µM (mean ± SEM) for AML and 0.55 ± 0.09 µM for ALL, indicating potent anti-proliferative effects. In contrast, PTC-209 showed differential cytotoxic effects between AML and ALL cells. The ED50 values (effective concentration inducing 50% killing as measured by Annexin V positivity) were no more than 2.5 µM in 5 of 6 AML lines while they were higher than 10 µM in 3 out of 5 ALL cell lines, implicating that BMI-1 is more critical in AML than ALL. Treatment with PTC-209 triggered several molecular events consistent with induction of apoptosis in sensitive lines (e.g. MV4-11 and MOLM-13): conformational change of BAX (i.e., BAX activation), loss of mitochondrial membrane potential (MMP), caspase-3 activation and DNA fragmentation in addition to phosphatidylserine (PS) externalization. Eighteen-hour treatment of MV4-11 cells with 2.5 µM PTC-209 led to compound-specific induction of conformationally active BAX (31%), MMP loss (80%) and caspase-3 cleavage (38%). qRT-PCR showed reduced transcript level of BMI-1 (61% reduction) after 6-hour PTC-209 exposure in MV4-11 cells. PTC-209 induced PS externalization in primary AML cells (82.5 ± 4.3% after 48-hour treatment with 2 µM PTC-209, n = 6) and to a lesser degree, in ALL cells (33.7 ± 13.4%, n = 4, p 〈 0.05). Importantly, CD34+CD38– AML progenitor cells were as sensitive to PTC-209 as C34– more mature AML cells. Normal lymphocytes were resistant to PTC-209 (9.1 ± 4.6% even at 10 µM). Collectively, BMI-1 inhibition by small molecule inhibitors could be developed into a novel therapeutic strategy. Disclosures No relevant conflicts of interest to declare.

Type of Medium: Online Resource

ISSN: 0006-4971 , 1528-0020

URL: Article

DOI: 10.1182/blood.V124.21.3617.3617

Language: English

Publisher: American Society of Hematology

Publication Date: 2014

detail.hit.zdb_id: 1468538-3

detail.hit.zdb_id: 80069-7