scholarly journals Upregulation of Thymidylate Synthase Induces Pemetrexed Resistance in Malignant Pleural Mesothelioma

2021 ◽  
Vol 12 ◽  
Author(s):  
Yuzo Sato ◽  
Masaru Tomita ◽  
Tomoyoshi Soga ◽  
Atsushi Ochiai ◽  
Hideki Makinoshima
Keyword(s):  
Drug Resistance ◽  
Cell Lines ◽  
Malignant Pleural Mesothelioma ◽  
Quantitative Polymerase Chain Reaction ◽  
Resistance Mechanism ◽  
Parental Cell ◽  
Resistant Cell ◽  
Pleural Mesothelioma ◽  
Pyrimidine Synthesis ◽  
Resistant Cells

Malignant pleural mesothelioma (MPM) is an invasive malignancy that develops in the pleural cavity, and antifolates are used as chemotherapeutics for treating. The majority of antifolates, including pemetrexed (PMX), inhibit enzymes involved in purine and pyrimidine synthesis. MPM patients frequently develop drug resistance in clinical practice, however the associated drug-resistance mechanism is not well understood. This study was aimed to elucidate the mechanism underlying resistance to PMX in MPM cell lines. We found that among the differentially expressed genes associated with drug resistance (determined by RNA sequencing), TYMS expression was higher in the established resistant cell lines than in the parental cell lines. Knocking down TYMS expression significantly reduced drug resistance in the resistant cell lines. Conversely, TYMS overexpression significantly increased drug resistance in the parental cells. Metabolomics analysis revealed that the levels of dTMP were higher in the resistant cell lines than in the parental cell lines; however, resistant cells showed no changes in dTTP levels after PMX treatment. We found that the nucleic acid-biosynthetic pathway is important for predicting the efficacy of PMX in MPM cells. The results of chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR) assays suggested that H3K27 acetylation in the 5′-UTR of TYMS may promote its expression in drug-resistant cells. Our findings indicate that the intracellular levels of dTMP are potential biomarkers for the effective treatment of patients with MPM and suggest the importance of regulatory mechanisms of TYMS expression in the disease.

Analysis of mRNAs and ncRNAs and Prediction Competing Endogenous RNA Networks in Colorectal Cancer Chemo-Resistance by Whole-Transcriptome Sequencing

2020 ◽  
Author(s):  
Fei Yao ◽  
Chuanren Zhou ◽  
Qiyou Huang ◽  
Xiaoying Huang ◽  
Jie Chen ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Drug Resistance ◽  
Cell Lines ◽  
Transcriptome Sequencing ◽  
Regulatory Networks ◽  
Parental Cell ◽  
Resistant Cell ◽  
Competing Endogenous Rna ◽  
Whole Transcriptome Sequencing ◽  
Whole Transcriptome

Abstract Background: Chemo-resistance is a major clinical obstacle to the treatment of colorectal cancer (CRC), mRNAs and non-coding RNAs (ncRNAs) have been reported to modulate the development of chemo-resistance. However, the profiles of mRNAs and ncRNAs as well as competing endogenous RNA (ceRNA) networks in CRC chemo-resistance are still unclear, and whether different drug resistance of CRC have the same mechanisms also needs to be explored. This study aims to uncover the expression of mRNAs and ncRNAs in parental cell lines and different chemo-resistant cell lines, and construct ceRNA regulatory networks by whole-transcriptome sequencing.Methods: The expression of mRNAs and ncRNAs in parental cell lines and drug-resistant cell lines were identified by whole-transcriptome sequencing and bioinformatics methods.Results: A total of 1779 mRNAs, 64 miRNAs, 11 circRNAs and 295 lncRNAs were common differentially expressed in two different chemo-resistant cell lines when compared with the control. In addition, 5,767 lncRNA-miRNA-mRNA relationship pairs and 47 circRNA-miRNA-mRNA pathways were constructed according to ceRNA regulatory rules, in which AC109322.2-hsa-miR-371a-5p-BTNL3 and hsacirc_027876-hsa-miR-582-3p-FREM1 were identified as the most potential ceRNA networks involved in drug resistance to CRC. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of two ceRNA regulatory networks showed that the TNF signaling pathway may be crucial in the process of CRC drug resistance.Conclusions: A large number of mRNAs and ncRNAs in chemo-resistant cell lines were different expressed, which may play pivotal roles in development of drug resistance through the ceRNA regulatory network. This study may improve our understanding of the underlying mechanisms and provide a promising therapeutic strategy for CRC chemo-resistance.

scholarly journals Importance of Cullin4 Ubiquitin Ligase in Malignant Pleural Mesothelioma

Cancers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3460
Author(s):  
Mayura Meerang ◽  
Jessica Kreienbühl ◽  
Vanessa Orlowski ◽  
Seraina L. C. Müller ◽  
Michaela B. Kirschner ◽  
...  
Keyword(s):  
Cell Lines ◽  
Malignant Pleural Mesothelioma ◽  
Ubiquitin Ligase ◽  
Tumor Model ◽  
Progression Free Survival ◽  
Neurofibromatosis Type ◽  
Resistant Cell ◽  
Pleural Mesothelioma ◽  
G2 Cell Cycle Arrest

Neurofibromatosis type 2 (NF2), the tumor suppressor frequently lost in malignant pleural mesothelioma (MPM), suppresses tumorigenesis in part by inhibiting the Cullin4 ubiquitin ligase (CUL4) complex in the nucleus. Here, we evaluated the importance of CUL4 in MPM progression and tested the efficacy of cullin inhibition by pevonedistat, a small molecule inhibiting cullin neddylation. CUL4 paralogs (CUL4A and CUL4B) were upregulated in MPM tumor specimens compared to nonmalignant pleural tissues. High gene and protein expressions of CUL4B was associated with a worse progression-free survival of MPM patients. Among 13 MPM cell lines tested, five (38%) were highly sensitive to pevonedistat (half maximal inhibitory concentration of cell survival IC50 < 0.5 µM). This remained true in a 3D spheroid culture. Pevonedistat treatment caused the accumulation of CDT1 and p21 in both sensitive and resistant cell lines. However, the treatment induced S/G2 cell cycle arrest and DNA rereplication predominantly in the sensitive cell lines. In an in vivo mouse model, the pevonedistat treatment significantly prolonged the survival of mice bearing both sensitive and resistant MPM tumors. Pevonedistat treatment reduced growth in sensitive tumors but increased apoptosis in resistant tumors. The mechanism in the resistant tumor model may be mediated by reduced macrophage infiltration, resulting from the suppression of macrophage chemotactic cytokines, C-C motif chemokine ligand 2 (CCL2), expression in tumor cells.

scholarly journals SORT1/LAMP2-mediated Extracellular Vesicle Secretion and Cell Adhesion Are Linked to Lenalidomide Resistance in Multiple Myeloma

2022 ◽  
Author(s):  
Tomofumi Yamamoto ◽  
Jun Nakayama ◽  
Yusuke Yamamoto ◽  
Masahiko Kuroda ◽  
Yutaka Hattori ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Adhesion ◽  
Cell Lines ◽  
Extracellular Vesicle ◽  
Parental Cell ◽  
Resistant Cell ◽  
Cellular Mechanisms ◽  
Hematopoietic Malignancy ◽  
Patient Prognosis ◽  
Resistant Cells

Multiple myeloma (MM) is a hematopoietic malignancy whose prognosis has improved with the development of new agents such as lenalidomide over the last decade. However, long-term exposure to drugs induces the acquisition of resistance by MM cells and leads to treatment failure and poor prognosis. Here, we show the molecular and cellular mechanisms of lenalidomide resistance in MM. In a comparison between lenalidomide-resistant cell lines and the parental cell lines, the EV (Extracellular versicles) secretion and adherence abilities were significantly elevated in the resistant cells. Whole-transcriptome analysis revealed that the SORT1 and LAMP2 genes were key regulators of EV secretion. Silencing of these genes caused decreased EV secretion and loss of cell adhesion in the resistant cells, resulting in increased sensitivity to lenalidomide. Analysis of publicly available transcriptome data confirmed the relationship between genes related to EV secretion and cell adhesion and patient prognosis. Together, our findings reveal a novel mechanism of lenalidomide resistance in MM mediated by EV secretion and cell adhesion via SORT1 and LAMP2.

A New Form Of Therapeutic Resistance: Drug Glucuronidation Regulated By The Sonic Hedgehog Factor Gli1

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 821-821
Author(s):  
Hiba A Zahreddine ◽  
Biljana Culjkovic-Kraljacic ◽  
Sarit Assouline ◽  
Abdellatif Amri ◽  
Patrick Gendron ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Cell Lines ◽  
Phase Ii ◽  
Resistant Cell ◽  
Type I ◽  
Type Ii ◽  
Protein Levels ◽  
Gli 1 ◽  
Type Ii Resistance ◽  
Resistant Cells

Abstract Despite many recent successes in the treatment of cancer, the development of chemoresistance in many of the initially responding patients, and primary resistance in others, remains a major impediment in therapy development. Our studies provide evidence for a novel mechanism underlying drug resistance: Gli1 dependent drug glucuronidation. While carrying out a Phase II clinical trial of targeting the eukaryotic translation initiation factor eIF4E with ribavirin in M4/M5 subtypes of AML, we observed that all responding patients eventually became clinically and molecularly resistant. To understand the cause of this resistance, we generated ribavirin resistant cell lines. In these models, ribavirin no longer targeted eIF4E activity or impaired growth, and importantly, the ability of ribavirin to bind eIF4E was severely impaired. However, the eIF4E gene was not mutated and its protein levels were not altered. The cell lines could be divided into two groups: type I with a defect in drug uptake and type II with a normal uptake. In type I resistant cells, we observed a substantial reduction in levels of Adenosine Kinase (ADK) an enzyme that catalyzes the rate limiting step in the metabolic activation of ribavirin allowing its retention in the cells. We used RNA Sequencing to examine the molecular underpinnings of type II resistance. Our data revealed a drastic increase in the levels of Gli1. In stably overexpressing cells, Gli1 was sufficient to produce the same resistance phenotype that we observed for type II cell models, both molecularly and at the level of cell growth. In addition, Gli1 overexpression correlated with the loss of drug-to-target interaction, as observed by our eIF4E immunoprecipitation studies using 3H-Ribavirin, similarly to the resistant cell lines. Conversely, Gli1 knockdown in type II cells or its pharmacological inhibition with the FDA approved Gli1 inhibitor GDC0449/Vismodegib, restored the eIF4E-ribavirin interaction and re-sensitized these cells to ribavirin. Our subsequent studies revealed a close correlation between Gli1 expression and the protein levels of the UGT1A glucuronosyl transferase enzymes involved in phase II drug metabolism whereby xenobiotics or metabolites are modified by the addition of a sugar, glucuronic acid. Given these findings, we examined whether the loss of the eIF4E interaction in resistant cells was due to the glucuronidation of ribavirin. Using 13C/12C ribavirin and mass spectrometry, we observed glucuronidated forms of ribavirin in resistant cells and cells overexpressing Gli1 but not in parental cells and that ribavirin is glucuronidated on its triazole ring which binds eIF4E. Treatment of cells with the Gli1 inhibitor GDC0449 reduces UGT1A levels, and correlates with reduced levels of ribavirin-glucuronides and the re-emergence of ribavirin-eIF4E complexes. We further hypothesized that the type II resistant cells could be resistant to other drugs. We observe that our ribavirin resistant cell lines are also resistant to the cornerstone of AML therapy, cytarabine. GDC0449/Vismodegib treatment reverts resistance to cytarabine in these cells. Preliminary studies indicate that these cells are also resistant to azacytidine and cisplatin. This is particularly striking as these cells were never exposed to these compounds. Thus, this could represent a novel form of multi-drug resistance. To establish the clinical relevance of our findings to patients in our AML ribavirin trial, we examined features of type I and type II resistance. Out of 10 patient samples available for evaluation, all six responding patient specimens showed elevated Gli-1 mRNA levels, up to 26 fold, upon relapse relative to levels during response. For most, the ratio of Gli1 during response relative to at relapse was about 2-4 fold with some patients up to 10 fold. For the two patients examined that did not respond, both had highly elevated Gli-1 levels prior to treatment relative to healthy individuals, and this was not lowered after 28 days of ribavirin treatment. We also noted elevated UGT1A protein levels upon relapse in our patient population. Type I resistance was observed in only two patients whereas Gli1 and UGT1A were dysregulated at relapse in all patients examined. In summary, we identified a novel form of drug resistance: Gli1 dependent drug glucuronidation. Treatment with Gli1 inhibitors appears to be a promising avenue for overcoming this form of drug resistance. Disclosures: No relevant conflicts of interest to declare.

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.

scholarly journals Molecular Alterations Associated with Acquired Drug Resistance during Combined Treatment with Encorafenib and Binimetinib in Melanoma Cell Lines

Cancers ◽  
2021 ◽  
Vol 13 (23) ◽  
pp. 6058
Author(s):  
Vikas Patel ◽  
István Szász ◽  
Viktória Koroknai ◽  
Tímea Kiss ◽  
Margit Balázs
Keyword(s):  
Drug Resistance ◽  
Cell Lines ◽  
Melanoma Cell ◽  
Acquired Resistance ◽  
Resistance Mechanisms ◽  
Resistant Cell ◽  
Differentially Expressed ◽  
Molecular Alterations ◽  
Melanoma Cell Lines ◽  
Resistant Cells

Combination treatment using BRAF/MEK inhibitors is a promising therapy for patients with advanced BRAFV600E/K mutant melanoma. However, acquired resistance largely limits the clinical efficacy of this drug combination. Identifying resistance mechanisms is essential to reach long-term, durable responses. During this study, we developed six melanoma cell lines with acquired resistance for BRAFi/MEKi treatment and defined the molecular alterations associated with drug resistance. We observed that the invasion of three resistant cell lines increased significantly compared to the sensitive cells. RNA-sequencing analysis revealed differentially expressed genes that were functionally linked to a variety of biological functions including epithelial-mesenchymal transition, the ROS pathway, and KRAS-signalling. Using proteome profiler array, several differentially expressed proteins were detected, which clustered into a unique pattern. Galectin showed increased expression in four resistant cell lines, being the highest in the WM1617E+BRes cells. We also observed that the resistant cells behaved differently after the withdrawal of the inhibitors, five were not drug addicted at all and did not exhibit significantly increased lethality; however, the viability of one resistant cell line (WM1617E+BRes) decreased significantly. We have selected three resistant cell lines to investigate the protein expression changes after drug withdrawal. The expression patterns of CapG, Enolase 2, and osteopontin were similar in the resistant cells after ten days of “drug holiday”, but the Snail protein was only expressed in the WM1617E+BRes cells, which showed a drug-dependent phenotype, and this might be associated with drug addiction. Our results highlight that melanoma cells use several types of resistance mechanisms involving the altered expression of different proteins to bypass drug treatment.

scholarly journals Understanding the Role of Extracellular Vesicles in Lenalidomide-Resistance Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1887-1887
Author(s):  
Tomofumi Yamamoto ◽  
Nobuyoshi Kosaka ◽  
Ochiya Takahiro ◽  
Yutaka Hattori
Keyword(s):  
Bone Marrow ◽  
Drug Resistance ◽  
Cell Line ◽  
Cell Lines ◽  
Parental Cell ◽  
Resistant Cell ◽  
Bone Marrow Microenvironment ◽  
Drug Resistant ◽  
Apoptosis Assay ◽  
Drug Resistant Cell

Abstract Multiple myeloma (MM) is a plasma cell malignancy that develops by the accumulation of various genetic abnormalities. In recent years, the prognosis of MM has improved by the development of novel drugs including immunomodulatory drugs (IMiDs), proteasome inhibitors, and antibody drugs. However, MM cells acquired drug resistance by long-term exposure to these therapeutic drugs. MM is a multiclonal disease, and various clone subtypes develop within the bone marrow microenvironment. It has been suggested that drug resistant phenotype could transmit from resistant clones to sensitive ones, and also to immune cells, or mesenchymal stem cells, resulting in the change of bone marrow microenvironment suitable for MM cell survival. It has been shown that extracellular vesicles (EVs) are one of the means of information transmission. EVs are secreted from almost all cells, and the amount of EV secretion is particular high from cancer cells. It was already known that cancer-derived EVs transmitted information associated with cancer progressions such as angiogenesis, metastasis, and drug resistance to the surrounding cells. Thus, EVs were proposed to play an important role in acquisition of drug resistance even though the mechanisms have not been fully understood in MM. In order to understand the mechanism of drug resistance in MM mediated by EVs, lenalidomide resistant cell lines were established by long-term exposure of lenalidomide. The amount of EVs was measured by ExoScreen, which is ultra-sensitive detection method of EVs by measuring surface protein of EVs, such as, CD9 and CD63, and by the nanoparticle tracking analysis. We found that lenalidomide resistant cell lines in KMS21R, KMS27R and KMS34R cell lines secreted about twice more EVs than their parental cell lines, and the amount of EV secretion was correlated with the drug sensitivity of lenalidomide. Suppression of EV secretion by knockdown of TSG101, which is known for EV secretion-associated protein, did not affect lenalidomide resistance. We could suppress the EV secretion to two-thirds, however cell proliferation and caspase activity were not change. From these results, we postulated the two possibilities; 1) EV secretion pathway other than TSG101 is associated with drug resistance via EVs; 2) EV derived from lenalidomide resistant cells can affect the cells exist in bone marrow microenvironment. From these hypotheses, we have done the following experiments. Firstly, to identify the genes which involved in EV secretion pathway associated with drug resistance, RNA sequence among the drug-resistant cell lines and their parental cell lines was performed. Drug resistant cell lines had some genetic abnormality, for instance immune system or angiogenesis. Now, we are detecting the EV secretion associated genes in drug resistant cell lines. Secondly, EV derived from the drug-resistant cell lines and EV derived from the parent cell lines were added to drug sensitive MM cell lines, then lenalidomide is added after 24hr. The cell proliferation and apoptosis assay were evaluated after 48hr. EV derived from the drug-resistant cell lines in KMS34R cell line significantly inhibited cell death measured by MTS assay and apoptosis assay compared with those from the drug sensitive KMS21 and KMS34 cell lines. EVs from KMS34R cell line, which is the most progressed cell line we established, could more transmit drug resistance than those from other cell lines. These results suggested that drug resistance was transmitted from drug-resistant cell lines to non-resistant cell lines via EVs. Now, we are analyzing the component of EV from drug-resistant MM cells by proteome analysis to identify the molecules associated with lenalidomide resistance in MM. In addition, we are investigating the molecules which associated with the secretion of EVs from drug-resistance MM. These results prompted us to hypothesize that attenuating the function of a molecule responsible for EV secretion could lead to the inhibition of cancer development such as drug resistance. It is expected that EVs will be novel therapeutic targets in refractory or relapsed MM. Disclosures Hattori: IDAC inc.: Research Funding; Takeda: Research Funding.

scholarly journals KIFC1 Inhibitor CW069 Induces Apoptosis and Reverses Resistance to Docetaxel in Prostate Cancer

2019 ◽  
Vol 8 (2) ◽  
pp. 225 ◽  
Author(s):  
Yohei Sekino ◽  
Naohide Oue ◽  
Yuki Koike ◽  
Yoshinori Shigematsu ◽  
Naoya Sakamoto ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Viability ◽  
Cell Lines ◽  
Cancer Progression ◽  
Parental Cell ◽  
Resistant Cell ◽  
Potential Strategy ◽  
Kinesin Family ◽  
Resistant Cells

Kinesin family member C1 (KIFC1) is a minus end-directed motor protein that plays an essential role in centrosome clustering. Previously, we reported that KIFC1 is involved in cancer progression in prostate cancer (PCa). We designed this study to assess the involvement of KIFC1 in docetaxel (DTX) resistance in PCa and examined the effect of KIFC1 on DTX resistance. We also analyzed the possible role of a KIFC1 inhibitor (CW069) in PCa. We used DTX-resistant PCa cell lines in DU145 and C4-2 cells to analyze the effect of KIFC1 on DTX resistance in PCa. Western blotting showed that KIFC1 expression was higher in the DTX-resistant cell lines than in the parental cell lines. Downregulation of KIFC1 re-sensitized the DTX-resistant cell lines to DTX treatment. CW069 treatment suppressed cell viability in both parental and DTX-resistant cell lines. DTX alone had little effect on cell viability in the DTX-resistant cells. However, the combination of DTX and CW069 significantly reduced cell viability in the DTX-resistant cells, indicating that CW069 re-sensitized the DTX-resistant cell lines to DTX treatment. These results suggest that a combination of CW069 and DTX could be a potential strategy to overcome DTX resistance.

Overcoming Resistance to 5-Azacytidine in Acute Myelogenous Leukemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1370-1370
Author(s):  
Piyanuch Sripayap ◽  
Tadashi Nagai ◽  
Mitsuyo Uesawa ◽  
Hiroyuki Kobayashi ◽  
Tomonori Tsukahara ◽  
...  
Keyword(s):  
Gene Expression ◽  
Drug Resistance ◽  
Cell Lines ◽  
Resistant Cell ◽  
Myelogenous Leukemia ◽  
P16 Gene ◽  
Resistant Lines ◽  
Acute Myelogenous ◽  
Dependent Mechanism ◽  
Resistant Cells

Abstract Abstract 1370 Background: The DNA methylation inhibitor 5-azacytidine (AZA), which is approved for treatment of myelodysplastic syndrome, is also a potential agent for treatment of leukemia; however, drug resistance is an ongoing problem, and mechanisms underlying developing resistance to AZA are poorly understood. Therefore, clarifying the resistance mechanisms is central to establish effective countermeasures. Methods: To probe the mechanisms of resistance to AZA and to develop an effective method for overcoming them, we first generated two AZA-resistant cell lines, THP-1/AR and HL60/AR, from the human acute myelogenous leukemia cell lines THP-1 and HL60. We then studied variations between the parental and resistant lines. Results: AZA increased the percentages of sub-G1 and G2/M-phase cells in the AZA-sensitive parental cell lines; whereas, it had no similar effect in the resistant lines. Consistent with these results, the AZA-induced increases in the levels of cleaved forms of caspase 3, caspase 7, caspase 9, and PARP seen in sensitive cells were diminished in resistant cells. Furthermore, AZA markedly elevated the level of phospho JNK/SAPK in sensitive cells, but not in resistant cells. These results suggest that AZA induced apoptosis as well as G2/M arrest due to activation of JNK/SAPK signaling, and that induction of these changes was prevented in resistant cells. We also found that the activity as well as protein levels of DNA methyltransferases (DNMTs), which are the main target molecules of AZA, were suppressed by AZA in sensitive cells. However, in resistant cells, this effect was abrogated; and accordingly, AZA-induced up-regulation of p16 gene expression was also negated. These findings thus suggest that resistance was acquired by a DNMT-dependent mechanism. There was no remarkable difference between resistant cells and sensitive cells in the levels of uridine-cytidine kinase 2 (UCK2), which is a key enzyme for conversion of AZA to active form. However, several point mutations were found restrictedly in exon 4 of the UCK2 gene in both resistant cells. These results raised the possibility that the AZA activation process was perturbed due to reduction of UCK activity; and consequently, AZA failed to suppress DNMT in resistant cells. In addition, by microarray analysis, we identified eleven genes that were expressed at significantly different levels in resistant cells versus sensitive cells. Finally, we showed that the histone deacetylase inhibitor romidepsin induced p16 gene expression and increased the levels of apoptosis-related molecules, while suppressing growth in both sensitive and resistant cell lines. An isobologram analysis demonstrated that simultaneous administration of AZA and romidepsin resulted in an additive inhibitory effect on both AZA-sensitive and AZA-resistant cell growth. These results suggest that romidepsin can overcome AZA resistance; therefore, the combination of AZA and romidepsin not only augments the anti-leukemia effect but also prevents acquisition of resistance to AZA. Conclusions: Newly established 5-azacytidine-resistant cell lines THP-1/AR and HL60/AR are good models to analyze the mechanisms of drug resistance to 5-azacytidine. Using these cell lines, we revealed that acquisition of resistance is primarily caused by a DNMT-dependent mechanism, which can be surmounted with addition of romidepsin. It is likely that the combination of AZA and romidepsin can prevent patients from acquiring resistance to AZA while augmenting its anti-leukemia therapeutic effect. Disclosures: No relevant conflicts of interest to declare.

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