scholarly journals 1-Hydroxy-8-methoxy-anthraquinon reverses cisplatin resistance by inhibiting 6PGD in cancer cells

2019 ◽  
Vol 14 (1) ◽  
pp. 454-461 ◽  
Author(s):  
Huamin Zhang ◽  
Haowei Zhang ◽  
Sihui Wang ◽  
Zhihai Ni ◽  
Tiejun Wang
Keyword(s):  
Lung Cancer ◽  
Cell Lines ◽  
Small Molecule ◽  
Cisplatin Resistance ◽  
Small Molecule Inhibitor ◽  
Resistant Cell ◽  
Expression Level ◽  
Sensitive Cell ◽  
Molecule Inhibitor ◽  
Resistant Cells

AbstractTargeting 6-phosphogluconate dehydrogenase (6PGD) can inhibit cancer cell proliferation and tumor growth. However, the relationship between 6PGD and cisplatin resistance still needs further study. Cisplatin-sensitive and cisplatin-resistant ovarian cancer OV2008 and C13* lines and lung cancer A549 and A549DDP lines were treated with different concentrations of cisplatin and cell viability was evaluated. We also compared the growth rates and the cell cycle distributions between cisplatin-sensitive and cisplatin-resistant cells. The expression level of 6PGD was detected by immunoblotting. The Chou-Talalay method was used to evaluate the effect of a combination treatment using cisplatin and the small molecule inhibitor 1-Hydroxy-8-methoxy-anthraquinon (S3) that targets 6PGD. The cisplatin-resistant ovarian and lung cancer cell lines grew faster than the cisplatin- sensitive cell lines, with more cells in S and G2 phases in cisplatin-resistant cell lines. The expression level of 6PGD in cisplatin-resistant cell lines was significantly increased compared with cisplatin-sensitive cell lines. Furthermore, treatment of cells with the S3 small molecule inhibitor of 6PGD together with cisplatin could overcome cisplatin resistance. The expression level of 6PGD in cisplatin-resistant cells lines was significantly upregulated, and the resistance to cisplatin of drug-resistant cells lines could be overcome when treated with the small molecule inhibitor S3 that specifically targets 6PGD.

scholarly journals Small molecule inhibitor of OGG1 blocks oxidative DNA damage repair at telomeres and potentiates Methotrexate anticancer effects

2020 ◽  
Author(s):  
Juan Miguel Baquero ◽  
Carlos Benítez-Buelga ◽  
Varshni Rajagopal ◽  
Zhao Zhenjun ◽  
Raúl Torres-Ruiz ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Cell Lines ◽  
Small Molecule ◽  
Oxidative Dna Damage ◽  
Genome Instability ◽  
Excision Repair ◽  
Small Molecule Inhibitor ◽  
Osteosarcoma Cell ◽  
Molecule Inhibitor

Abstract Background: The most common oxidative DNA lesion is 8-oxoguanine (8-oxoG) which is mainly recognized and excised by the glycosylase OGG1, initiating the Base Excision Repair (BER) pathway. Telomeres are particularly sensitive to oxidative stress which disrupts telomere homeostasis triggering genome instability. Methods: We used U2OS OGG1-GFP osteosarcoma cell line to study the role of OGG1 at the telomeres in response to oxidative stress. Next, we investigated the effects of inactivating pharmacologically the BER during oxidative stress (OS) conditions by using a specific small molecule inhibitor of OGG1 (TH5487) in different human cell lines. Results: We have found that during OS, TH5487 effectively blocks BER initiation at telomeres causing accumulation of oxidized bases at this region, correlating with other phenotypes such as telomere losses, micronuclei formation and mild proliferation defects. Besides, the antimetabolite Methotrexate synergizes with TH5487 through induction of intracellular ROS formation, which potentiates TH5487 mediated telomere and genome instability in different cell lines. Conclusions: Our findings demonstrate that OGG1 is required to protect telomeres from OS and present OGG1 inhibitors as a tool to induce oxidative DNA damage at telomeres, with the potential for developing new combination therapies for cancer treatment.

Abstract 2717: DMH1, a small molecule inhibitor of BMP type I receptors, suppresses growth and invasion of lung cancer

2014 ◽  
Author(s):  
Ying Huang ◽  
Rachel Lee ◽  
Andy Chang ◽  
Jeffery Fan ◽  
Chantelle Labib ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Small Molecule ◽  
Small Molecule Inhibitor ◽  
Type I ◽  
Molecule Inhibitor

70 A Selective Small Molecule Inhibitor of Mcl-1 Induces Bak Dependent Apoptosis in Cancer Cell Lines

2012 ◽  
Vol 48 ◽  
pp. 23
Author(s):  
D.C. Phillips ◽  
Y. Xiao ◽  
M. Bruncko ◽  
C. Park ◽  
H. Zhang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cancer Cell ◽  
Small Molecule ◽  
Cancer Cell Lines ◽  
Small Molecule Inhibitor ◽  
Molecule Inhibitor

The Tak-1 Inhibitor AZ-Tak1 Inhibits XIAP, Activates Caspase-9, and Induces Apoptosis in Mantle Cell Lymphoma.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1692-1692 ◽  
Author(s):  
Daniela Buglio ◽  
Sangeetha Palakurthi ◽  
Katharine F. Byth ◽  
Anas Younes
Keyword(s):  
Mantle Cell Lymphoma ◽  
Cell Lines ◽  
Small Molecule ◽  
Cell Lymphoma ◽  
Small Molecule Inhibitor ◽  
Caspase 9 ◽  
Lymphoma Cells ◽  
Molecule Inhibitor ◽  
Mantle Cell ◽  
Induced Apoptosis

Abstract Abstract 1692 Poster Board I-718 Transforming growth factor-b-activated kinase 1 (TAK1) is a key regulator of NF-kB activation. TAK1 can be activated by a variety of pro-inflammatory cytokines and T and B cell receptors. Recent experiments demonstrated that deletion of TAK1 results in inactivation of both JNK and NF-kB signaling resulting in massive apoptotic death of hematopoietic cells in mice. In this study, we examined the expression pattern of TAK1 and its role as a potential therapeutic target for lymphoma. First, we examined TAK1 expression in a panel of lymphoid cell lines by western blot, and found it to be highly expressed in mantle cell lymphoma cell lines (Mino, SP53, and Jeko-1). These lines expressed relatively low levels of the tumor suppressor protein A20. Mino and SP53 expressed high level of p-p38. Subsequently, we investigated the in vitro activity of the novel TAK1 small molecule inhibitor AZ-Tak1 in these cell lines. AZ-Tak1 is a potent and a relatively selective inhibitor of TAK1 kinase activity, with an IC50 of 0.009 mM. It also inhibits Jak2 but at a much higher concentration (IC50=0.18 mM). AZ-Tak1 treatment decreased the level of p38 and ERK in mantle cell lymphoma cells, and induced apoptosis in a dose and time dependent manner, with an IC50 of 0.1-0.5 mM. Using the annexin-V and PI staining and FACS analysis, After 48 hours of incubation, AZ-Tak1 (0.1 mM) induced apoptosis in 28%, 34% and 86% of Mino, SP53, and Jeko cells, respectively, which was increased to 32%, 42%, and 86% when 0.5 mM concentration was used. Similar activity was also observed when primary mantle cell lymphoma cells were examined. Using pathway-specific protein arrays focusing on apoptosis, kinases, and transcription factors, AZ-Tak1 (0.5 mM) altered the level of several proteins that regulate cell growth and survival, especially members of the inhibitors of apoptosis (IAP) family. Specifically, AZ-Tak1 decreased the level of SMAC/DIABOLO and cytochrome –C in the mitochondria, which was associated with a decrease in the level of the anti-apoptotic protein X-linked IAP (XIAP) and activation of the intrinsic apoptotic pathway as evident by activation of caspase 9, cleavage of caspase 3, and induction of apoptosis. Furthermore, and consistant with its ability to inhibit Jak2 activity, AZ-Tak1 reduced STAT2 and STAT6 levels. AZ-Tak1 demonstrated no significant effect on bcl-2 family members. Finally, co-treatment with HDAC inhibitors demonstrated synergistic effect with low concentrations of AZ-Tak1. Collectively, our data demonstrate that targeting TAK1 by the small molecule inhibitor AZ-Tak1 induces cell death in mantle cell lymphoma by activating the intrinsic apoptosis pathway, suggesting that targeting TAK1 may have a therapeutic value for the treatment of mantle cell lymphoma. Disclosures Palakurthi: Astra Zeneca: Employment. Byth:Astra Zeneca : Employment.

Pharmacological Blockade of NF-kB with AS602868, a Potent Small Molecule Inhibitor of IKK2 Leads to Apoptosis of Sensitive and Resistant Chronic Myeloid Leukemia Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2882-2882
Author(s):  
Veronique Imbert ◽  
Nadege Gonthier ◽  
Catherine Frelin ◽  
Nicolas Sirvent ◽  
Michael Hummelsberger ◽  
...  
Keyword(s):  
Cell Lines ◽  
Small Molecule ◽  
Hematologic Malignancies ◽  
Cytogenetic Response ◽  
Small Molecule Inhibitor ◽  
Cellular Targets ◽  
Promising Target ◽  
Molecule Inhibitor ◽  
First Choice ◽  
Dependent Growth

Abstract The Bcr-Abl inhibitor imatinib mesylate is now the first choice treatment for all newly diagnosed CML patients. Nevertheless not all patients reach a complete cytogenetic response (CCR) and many develop resistance for the drug upon disease progression. Additional cellular targets should be thus identified to develop alternative therapeutic strategies. The transcription factor NF-kB appears as a promising target for the treatment of cancer since it is a pro-survival factor, found abnormaly active in numerous hematologic malignancies and solid tumors. NF-kB activation relies on ubiquitin-mediated degradation of its inhibitor IkB after phosphorylation by 2 specific kinases, IKK1 and IKK2. We disrupted NF-kB signaling pathway by the use of AS602868, a small molecule inhibitor of IKK2 on two CML cell lines, LAMA84 and BaF3/Bcr-Abl and on primary CML cells. First we showed that NF-kB is constitutively activated in both CML cell lines and that this constitutive activation is under the control of the Bcr-Abl kinase. We next demonstrated that IKK2 targeting by AS602868 led to apoptosis of CML cells. AS602868 induced a dose dependent growth inhibition (IC50 3–4μM) of CML cell lines. This inhibition was associated with activation of cellular caspases 3, 8 and 9 and induction of DNA fragmentation. Interestingly AS602868 could induce death of imatinib resistant cellular clones derived from LAMA84 and BaF3-Bcr-Abl cells (IC50 1–2μM). Finally AS602868 selectively inhibited the proliferation and the hematopoietic colony formation of primary CML cells derived from imatinib sensitive or resistant patients. Although the exact mechanism of NF-kB activation by Bcr-Abl remains to be elucidated, our data strongly suggest that targeting NF-kB with the IKK2 inhibitor AS602868 may represent a new promising therapeutic tool for the treatment of CML.

Simultaneous Targeting of Aurora Kinases and Bcr-Abl by the Small Molecule Inhibitor PHA-739358 Is Effective in Imatinib-Resistant Mutations Including T315I.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1042-1042 ◽  
Author(s):  
Artur Gontarewicz ◽  
Stefan Balabanov ◽  
Gunhild Keller ◽  
Riccardo Colombo ◽  
Alessio Graziano ◽  
...  
Keyword(s):  
Cell Lines ◽  
Small Molecule ◽  
Leukemia Cell ◽  
Chronic Phase ◽  
Adaptor Protein ◽  
Small Molecule Inhibitor ◽  
Aurora Kinases ◽  
Molecule Inhibitor ◽  
Clinical Resistance ◽  
T315i Mutation

Abstract The emergence of resistance to Imatinib (IM) mediated by mutations in the BCR-ABL domain became a major challenge in the treatment of chronic myeloid leukemia (CML). Second generation Bcr-Abl inhibitors overcome most mutations except T315I, a frequent cause of clinical resistance, emphasising the importance of developing alternative therapeutic strategies. Here, we report studies performed with a novel small molecule inhibitor, PHA-739358, which selectively targets Bcr-Abl and Aurora kinases. PHA-739358 exhibited strong antiproliferative and pro-apoptotic activity against a broad panel of human BCR-ABL positive and negative cell lines and against mouse BaF3 cells ectopically expressing wild type (wt) or IM-resistant BCR-ABL mutants, including T315I. Pharmacological synergism of IM and PHA-739358 was observed in leukemia cell lines with low-level resistance to IM, whereas no synergistic effects were observed in BCR-ABL negative or highly IM resistant T315I mutated cells. Treatment with PHA-739358 significantly decreased phosphorylation of histone H3-(Ser10), a marker of Aurora B activity and of the small adaptor protein CrkL, an accepted down-stream target of Bcr-Abl. Hence, PHA-739358 acts via combined inhibition of Bcr-Abl and Aurora kinases. Moreover, strong antiproliferative effects of PHA-739358 were observed in CD34+ cells derived from newly diagnosed CML patients and from IM-resistant individuals in chronic phase or blast crisis including those harbouring a T315I mutation. Thus, PHA-739358 represents a promising new strategy for treatment of IM resistant BCR-ABL positive leukemias, including those harbouring the T315I mutation. Clinical trials investigating this compound in IM-resistant CML have recently been initiated.

Evaluation of TW-37, a pan Bcl-2 Proteins Small-Molecule Inhibitor, Against Spectrum of Human B-Cell Lines and Patient-Derived Samples.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4521-4521
Author(s):  
Ramzi M. Mohammad ◽  
Yuan Sun ◽  
Shaomeng Wang ◽  
Amro Aboukameel ◽  
Ayad M. Al-Katib
Keyword(s):  
Tumor Growth ◽  
B Cell ◽  
Cell Lines ◽  
Small Molecule ◽  
Cell Lymphoma ◽  
Lymphoblastic Leukemia ◽  
B Cell Lymphoma ◽  
Small Molecule Inhibitor ◽  
Molecule Inhibitor ◽  
Human B Cell

Abstract Non-Hodgkin’s lymphoma (NHL) tumors include a group of heterogeneous diseases with varying natural histories and responsiveness to therapy; nonetheless, overexpression of Bcl-2 protein is seen in more than 80% of NHL. Throughout the years our laboratory succeeded in establishing a panel of B-cell lines representing various maturational stages of NHL. In this study, we have utilized a structure-based strategy to design a new class of potent nonpeptidic small-molecule inhibitor (SMI) of Bcl-2 family. TW-37, a lead compound that was designed to target the BH3 binding groove of antiapopototic Bcl-2 proteins. It binds to Bcl-2, Bcl-XL and Mcl-1 with Ki values of 290 nM, 1110 nM and 260 nM, respectively. TW-37 showed significant antiproliferative effect against Pre-B-Acute Lymphoblastic Leukemia (WSU-pre-B-ALL), Diffuse Large Cell Lymphoma (WSU-DLCL2), Follicular Small Cleaved Cell Lymphoma (WSU-FSCCL), Waldenstrom’s Macroglobulinemia (WSU-WM) and primary cells obtained from lymphoma patients, despite variations in their anti- and pro-apoptotic Bcl-2 proteins (Bcl-2, Bcl-XL, Mcl-1, Bax, Bak, Bim, Bad, BUMA and Bok). The IC50 for TW-37 varied from 165 nM in the WSU-FSCCL to 300 nM in WSU-DLCL2 cells. Apoptosis was independent of proliferative status or pathological classification of B-cell tumor. TW-37 was able to block Bim-Bcl-XL and Bim-Mcl-1 eterodimerization and induces apoptosis via activation of caspases -9, -3, PARP and DNA fragmentation. Although cell lines and patient samples expressed multiple Bcl-2 family proteins at various levels, TW-37 induced apoptosis was only strongly associated with Bax:Mcl-1 ratio. TW-37 administered to tumor-bearing SCID mice led to significant tumor growth inhibition (T/C), tumor growth delay (T-C) and Log10kill, when used at its maximum tolerated dose (40 mg/kg x 3days) via tail vein. failed to induce changes in the Bcl-2 proteins levels suggests that assessment of baseline Bcl-2 family proteins can be used to prognosticate the response to drug. These findings indicate activity of TW-37 across the spectrum of human B-cell tumors and support the concept of targeting the Bcl-2 system as a therapeutic strategy in the treatment of B-cell lymphoma.

Development of a Small-Molecule Inhibitor Screen to Rapidly Identify Key Signaling Pathways in Leukemogenesis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 708-708
Author(s):  
Jeffrey W Tyner ◽  
Luke Fletcher ◽  
Wayne Yang ◽  
Stephen T Oh ◽  
Jason R. Gotlib ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Cell Viability ◽  
Signaling Pathways ◽  
Cell Lines ◽  
Small Molecule ◽  
Tyrosine Kinases ◽  
Leukemic Cell ◽  
Small Molecule Inhibitor ◽  
Kinase Activation ◽  
Molecule Inhibitor

Abstract Abstract 708 Aberrantly activated tyrosine kinases and their associated signaling pathways are critical to leukemogenesis and primary acute myeloid leukemia (AML) cell viability. While aberrant kinase activation has been confirmed in a significant percentage of AML, constitutive phosphorylation of STAT5, a marker of tyrosine kinase activation, is present in the majority of AML samples indicating that as yet unidentified tyrosine kinases can be aberrantly activated and contribute to leukemogenesis. Efforts to identify activating tyrosine kinase mutations using high-throughput sequencing have identified low frequency mutations of uncertain functional significance. Because these studies failed to detect additional high-frequency kinase mutations, the identity and mechanism of tyrosine kinase activation may be unique in many AMLs. To avoid the imitations of high-throughput sequencing, we have developed a functional assay that can rapidly and simultaneously identify therapeutic targets while providing therapeutic options. Methods: To rapidly identify activated kinase pathways in individual, primary AML samples, we have developed a small-molecule inhibitor array which includes 90 small-molecule, cell-permeable inhibitor compounds including a core of 36 tyrosine kinase inhibitors that covers the majority of the tyrosine kinome. Many of the inhibitors are available for clinical use or are in clinical development. In this assay, inhibitors were placed in 96-well plates at four serial dilutions to allow IC50 calculations. Three days after adding primary AML cells to each well, we performed an MTS cell viability assay to evaluate the effect of each inhibitor on cell viability. Because most inhibitors affect multiple kinases, we compared target specificities of compounds that decrease primary AML cell viability with those that have no effect to identify potential targets. Results: In preliminary proof-of-principal experiments, we tested leukemia cell lines with known activating tyrosine kinase mutations and Ba/F3 cell lines expressing activated tyrosine kinases. Appropriate inhibitor sensitivity profiles were obtained in CMK cells which depend on a JAK3 A572V mutation for viability, MKPL-1 cells with an activating CSF1R translocation, and in a Ba/F3 line expressing JAK2 V617F. In addition to the primary target, downstream targets were frequently identified; MKPL-1 cells also showed sensitivity to phosphoinositol 3-kinase and NFKB inhibitors. Thus, not only primary targets but the downstream signaling pathways critical to leukemic cell viability can be highlighted using this assay. To date, we have analyzed approximately 150 primary leukemia and lymphoma samples. In some cases, targets could be identified by comparison of overlapping kinase specificities for compounds that decreased leukemic cell viability and subtraction of possible kinase targets inhibited by compounds that had no effect on viability. However, many cases exhibited complex, often unique, inhibitor sensitivity profiles that complicated target identification. Comparison with sensitivity profiles for known aberrantly activated kinases was useful when available. Accordingly, additional leukemia cell lines and Ba/F3 lines that depend on a single aberrantly activated tyrosine kinase for viability are being evaluated. Automated scripts that correlate the leukemic cell inhibitor sensitivity with the inhibitor target specificity are also in preparation. Conclusions: These preliminary data demonstrate that the small-molecule inhibitor functional assays can rapidly identify disease causing genes, provide insights into their mechanism of action, and suggest therapeutic options. The distinct patterns of tyrosine kinase sensitivity in these samples support the hypothesis that tyrosine kinases and related pathways contributing to leukemogenesis in each patient may be different and that targeted therapy will be most effective when administered on an individualized basis. Disclosures: Druker: OHSU patent #843 - Mutate ABL Kinase Domains: Patents & Royalties; MolecularMD: Equity Ownership; Roche: Consultancy; Cylene Pharmaceuticals: Consultancy; Calistoga Pharmaceuticals: Consultancy; Avalon Pharmaceuticals: Consultancy; Ambit Biosciences: Consultancy; Millipore via Dana-Farber Cancer Institute: Patents & Royalties; Novartis, ARIAD, Bristol-Myers Squibb: Research Funding.

Mechanisms of Acquired Resistance to Venetoclax in Preclinical AML Models

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 328-328 ◽  
Author(s):  
Qi Zhang ◽  
Rongqing Pan ◽  
Lina Han ◽  
Ce Shi ◽  
Stephen E. Kurtz ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Lines ◽  
Small Molecule ◽  
Research Funding ◽  
Synergistic Effects ◽  
Acquired Resistance ◽  
Parental Cell ◽  
Resistant Cell ◽  
Flt3 Inhibitors ◽  
Resistant Cells

Abstract BH3-mimetic ABT-199 (venetoclax, VEN) is a selective small-molecule antagonist of the anti-apoptotic BCL-2 protein. It binds to BCL-2 specifically, causing the release of pro-apoptotic BAX and BH3-only proteins and induction of cell death. Our studies indicated that AML is a BCL-2 dependent disease that, in pre-clinical studies, responds robustly to VEN by induction of apoptotic cell death (Pan et al., Cancer Discovery 2014). As a single agent, VEN demonstrated clinical activity in relapsed/refractory AML, yet patients who initially responded ultimately developed resistance and progressed. In this study we investigated mechanisms of acquired resistance to VEN in preclinical AML models. First, we generated 5 VEN-resistant cell lines (OCI-AML2, Kasumi, KG-1, MV4;11 and Molm13; with VEN cell-killing IC50s of 0.021µM, 0.046µM, 0.073µM, 0.020µM and 0.050µM, respectively) by exposing the cells to gradually increasing VEN concentrations. The IC50s of resistant cells are 15.2µM, 5.7µM, 31.6µM, 11.4µM and 15.4µM (124-723-fold greater than their parental counterparts). Protein analysis of resistant cells using immunoblotting demonstrated increased expression of MCL-1, a known resistance factor to VEN, in 4 resistant cell lines (OCI-AML2, KG-1, Mv4;11 and Molm13); and BCL-XL increase in MV4;11 and Molm13 resistant cells. To characterize the functional role of MCL-1 and BCL-XL in resistance to VEN, we co-treated parental and resistant cells with novel MCL-1 and BCL-XL- selective inhibitors (A-1210477 and A-1155463). The combination of VEN with A-1210477 or A-1155463 showed synergistic growth inhibition in all 5 parental cell lines (combination indices (CI) for A-1210477 were 0.15-0.62; CI for A-1155463 were 0.33-0.51, except >3 for KG-1). Notably, 4 out of 5 resistant cell lines (OCI-AML2, Kasumi, MV4;11, Molm13) became more sensitive to MCL-1 selective inhibitor A-1155463 but not to BCL-XL inhibitor A-1210477. However, no further effects were seen in resistant cells when combined with VEN. We next compared sensitivity of three paired parental and resistant cell lines (OCI-AML2, MV4;11 and Molm13) to a library of 130 specific small-molecule inhibitors (Tyner, et.al.. Cancer Res. 2013). Cells were co-treated with VEN and each specific inhibitor, and drug target scores were calculated based on the IC50 of measured effectiveness of panel drugs against the cells. The screening revealed modulation of sensitivity to mTOR, MEK, and FLT3 pathways in resistant cells (Fig.1C). To confirm these findings, we next co-treated AML cells with VEN and specific inhibitors of the mTOR pathway (rapamycin and AZD2014) or MEK pathway (CI1040) in all 5 paired parental and resistant cell lines; or with FLT3 inhibitors (quizartinib and sorafenib) in parental and resistant MV4;11 and Molm13, which harbor FLT3-ITD. The combination of VEN and AZD2014 achieved synergistic effects in all 5 parental cell lines (CI AZD2014: 0.08-0.94), and VEN/rapamycin were synergistic in 3 parental cell lines (CI rapamycin: 0.00-0.55, except 1.76 for KG-1 and 1.59 for Molm13). Combination of VEN with CI1040 achieved synergy in OCI-AML2, Kasumi, MV4;11 and Molm13 parental cell lines (CI: 0.14-0.61). Finally, VEN/FLT3 inhibitors achieved synergistic effects in MV4;11 and Molm13 parental cell lines (CI quizartinib: 0.66-0.69; CI sorafenib: 0.64-0.71). The resistant cell lines exhibited sensitivity to these inhibitors as single agents, and no synergistic effects were seen when combined with VEN. We have further induced in vivo resistance in two primary AML xenografts by treating NSG mice engrafted with 2nd passage AML cells with 100 mg/kg Q.D. VEN for 4 weeks followed by harvest of leukemic cells that repopulated the mouse after treatment discontinuation. While the proteomics, gene expression (RNAseq) and drug screening assays are in progress, preliminary immunoblotting studies demonstrated decreased expression of BCL-XL and BCL-2 (Fig.1B). In summary, we identified multiple mechanisms of acquired resistance to VEN, which ultimately modulate the balance between pro- and anti-apoptotic BCL-2 family members. Our studies indicate that upfront combination of VEN with selective inhibitors of MCL-1, or with inhibitors of specific signaling pathways, can synergistically induce apoptosis in AML cells and conceivably prevent emergence of VEN resistance. Disclosures Leverson: AbbVie: Employment, Equity Ownership. Tyner:Aptose Biosciences: Research Funding; Constellation Pharmaceuticals: Research Funding; Janssen Pharmaceuticals: Research Funding; Array Biopharma: Research Funding; Incyte: Research Funding. Konopleva:Novartis: Research Funding; AbbVie: Research Funding; Stemline: Research Funding; Calithera: Research Funding; Threshold: Research Funding.

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