Molecular Biology and Genetics / Moleküler Biyoloji ve Genetik

Permanent URI for this collectionhttps://hdl.handle.net/11147/9

Browse

Filters

2011 - 20143

Settings

Author: search.filters.author.Öğretmen, Besim

Search Results

Now showing 1 - 3 of 3
  • Conference Object
    Targeting Sphingosine Kinase-1/Spingosine-1-phosphate Receptor 2 Signalling Pathway To Overcome T315i Mutation in 32dcl3 Cells
    (Elsevier Ltd., 2014) Adan Gökbulut, Aysun; Öğretmen, Besim; Baran, Yusuf
    The main problem in chronic myeloid leukemia patients is the development of resistance against tyrosine kinase inhibitors. The expression of BCR-ABL1 having T315 mutation is responsible for the development of nilotinib resistance. The alterations in sphingolipid signalling pathway is a significant BCR-ABL1-dependent resistance mechanism. Recently, we showed that sphingosine kinase-1 (SK-1)/sphingosine-1 phosphate (S1P)-mediated drug resistance is transduced via sphingosine-1 phosphate receptor 2 (S1P2) that inhibits protein phosphatase 2A (PP2A), causing increased stability of BCR-ABL1. However, specific signaling cascade involved in this process remain unkown. In this study, BCR-ABL1 expressing 32Dcl3 cells, 32D-p210Bcr-Abl(wt) and 32D-p210Bcr-Abl (T315I) were used. The antiproliferative effects of nilotinib, SK-1 inhibitor (PF-543), S1P2 inhibitor (JTE-013), phospholipase C inhibitor (U-73122) and nilotinib/PF-543 and nilotinib/JTE-013 combinations on wt and resistant cells were determined by MTT assay. Isobologram analysis was performed using CompuSyn program.
  • Article
    Citation - WoS: 108
    Citation - Scopus: 119
    Sphingosine Kinase-1 and Sphingosine 1-Phosphate Receptor 2 Mediate Bcr-Abl1 Stability and Drug Resistance by Modulation of Protein Phosphatase 2a
    (American Society of Hematology, 2011) Salas, Arelis; Ponnusamy, Suriyan; Senkal, Can E.; Meyers-Needham, Marisa; Selvam, Shanmugam Panneer; Saddoughi, Sahar A.; Apohan, Elif; Sentelle, R. David; Smith, Charles; Gault, Christopher R.; Obeid, Lina M.; El-Shewy, Hesham M.; Oaks, Joshua; Santhanam, Ramasamy; Marcucci, Guido; Baran, Yusuf; Mahajan, Sandeep; Fernandes, Daniel; Stuart, Robert; Perrotti, Danilo; Öğretmen, Besim
    The mechanisms by which sphingosine kinase-1 (SK-1)/sphingosine 1-phosphate (S1P) activation contributes to imatinib resistance in chronic myeloid leukemia (CML) are unknown. We show herein that increased SK-1/S1P enhances Bcr-Abl1 protein stability, through inhibition of its proteasomal degradation in imatinib-resistant K562/IMA-3 and LAMA-4/IMA human CML cells. In fact, Bcr-Abl1 stability was enhanced by ectopic SK-1 expression. Conversely, siRNA-mediated SK-1 knockdown in K562/IMA-3 cells, or its genetic loss in SK-1-/- MEFs, significantly reduced Bcr-Abl1 stability. Regulation of Bcr-Abl1 by SK-1/S1P was dependent on S1P receptor 2 (S1P2) signaling, which prevented Bcr-Abl1 dephosphorylation, and degradation via inhibition of PP2A. Molecular or pharmacologic interference with SK-1/S1P2 restored PP2A-dependent Bcr-Abl1 dephosphorylation, and enhanced imatinib- or nilotinib-induced growth inhibition in primary CD34+ mononuclear cells obtained from chronic phase and blast crisis CML patients, K562/IMA-3 or LAMA4/IMA cells, and 32Dcl3 murine progenitor cells, expressing the wild-type or mutant (Y253H or T315I) Bcr-Abl1 in situ. Accordingly, impaired SK-1/S1P2 signaling enhanced the growth-inhibitory effects of nilotinib against 32D/T315I-Bcr-Abl1-derived mouse allografts. Since SK-1/S1P/S1P2 signaling regulates Bcr-Abl1 stability via modulation of PP2A, inhibition of SK-1/S1P2 axis represents a novel approach to target wild-type- or mutant-Bcr-Abl1 thereby overcoming drug resistance. © 2011 by The American Society of Hematology.
  • Article
    Citation - WoS: 49
    Citation - Scopus: 50
    Targeting glucosylceramide synthase sensitizes imatinib-resistant chronic myeloid leukemia cells via endogenous ceramide accumulation
    (Springer Verlag, 2011) Baran, Yusuf; Bielawski, Jacek; Gündüz, Ufuk; Öğretmen, Besim
    Purpose: Drug resistance presents a major obstacle for the treatment of some patients with chronic myeloid leukemia (CML). Pro-apoptotic ceramide mediates imatinib-induced apoptosis, and metabolism of ceramide by glucosylceramide synthase (GCS) activity, converting ceramide to glucosyl ceramide, might contribute to imatinib resistance. In this study, we investigated the role of ceramide metabolism by GCS in the regulation of imatinib-induced apoptosis in drug-sensitive and drug-resistant K562 and K562/IMA-0.2 and K562/IMA-1 human CML cells, which exhibit about 2.3- and 19-fold imatinib resistance, respectively. Methods: Cytotoxic effects of PDMP and imatinib were determined by XTT cell proliferation assay. Expression levels of GCS were determined by RT-PCR and western blot. Intracellular ceramide levels were determined by LC-MS. Cell viability analyses was conducted by Trypan blue dye exclusion assay. Cell cycle and apoptosis analyses were examined by flow cytometry. Results: We first showed that mRNA and protein levels of GCS are increased in drug-resistant K562/IMA as compared to sensitive K562 cells. Next, forced expression of GCS in sensitive K562 cells conferred resistance to imatinib-induced apoptosis. In reciprocal experiments, targeting GCS using its known inhibitor, PDMP, enhanced ceramide accumulation and increased cell death in response to imatinib in K562/IMA cells. Conclusion: Our data suggest the involvement of GCS in resistance to imatinib-induced apoptosis, and that targeting GCS by PDMP increased imatinib-induced cell death in drug-sensitive and drug-resistant K562 cells via enhancing ceramide accumulation.