Human Chronic Myeloid Leukemia Cells Research Articles

Bioassay-guided isolation of anti-leukemic steroids from Aglaia abbreviata by inducing apoptosis

The strategy of bioactivity-guided isolation is widely used to obtain active compounds as quickly as possible. Thus, the inhibitory effects on human erythroleukemia cells (HEL) were applied to guide the isolation of the anti-leukemic compounds from Aglaia abbreviata. As a result, 19 compounds (16 steroids, two phenol derivatives, and a rare C12 chain nor-sesquiterpenoid), including 13 new compounds, were isolated and identified based on spectroscopic data analysis, single-crystal X-ray diffraction data, and electronic circular dichroism (ECD) calculations. Among them, 9 steroids exhibited good selective anti-leukemic activity against HEL and K562 (human chronic myeloid leukemia cells) cells with IC50 values between 2.29 ± 0.18 μM and 19.58 ± 0.13 μM. Notably, all the active compounds had relatively lower toxicity on the normal human liver cell line (HL-7702). Furthermore, five compounds (1, 4, 8, 10, and 19) displayed good anti-inflammatory effects, with IC50 values between 7.15 ± 0.16 and 27.1 ± 0.37 μM. An α,β-unsaturated ketone or a 5,6Δ double bond was crucial for improving anti-leukemic effect from the structure–activity relationship analysis. The compound with the most potential, 14 was selected for the preliminary mechanistic study. Compound 14 can induce apoptosis and cause cell cycle arrest. The expression of the marker proteins, such as PARP and caspase 3, were notably effected by this compound, thus inducing apoptosis. In conclusion, our investigation implied that compound 14 may serve as a potential anti-leukemia agent.

Anticancer activity features of imidazole-based ionic liquids and lysosomotropic detergents: in silico and in vitro studies.

Long-chain imidazole-based ionic liquids (compounds 2, 4, 9) and lysosomotropic detergents (compounds 7, 3, 8) with potent anticancer activity were synthesized. Their inhibitory activities against neuroblastoma and leukaemia cell lines were predicted by the new in silico QSAR models. The cytotoxic activities of the synthesized imidazole derivatives were investigated on the SK-N-DZ (human neuroblastoma) and K-562 (human chronic myeloid leukaemia) cell lines. Compounds 2 and 7 showed the highest in vitro cytotoxic effect on both cancer cell lines. The docking procedure of compounds 2 and 7 into the NAD+ coenzyme binding site of deacetylase Sirtuin-1 (SIRT-1) showed the formation of protein-ligand complexes with calculated binding energies of -8.0 and - 8.1kcal/mol, respectively. The interaction of SIRT1 with compounds 2, 7 and 9 and the interaction of Bromodomain-containing protein 4 (BRD4) with compounds 7 and 9 were also demonstrated by thermal shift assay. Compounds 2, 4, 7 and 9 inhibited SIRT1 deacetylase activity in the SIRT-Glo assay. Compounds 7 and 9 showed a moderate inhibitory activity against Aurora kinase A. In addition, compounds 3, 4, 8 and 9 inhibited the Janus kinase 2 activity. The results obtained showed that long-chain imidazole derivatives exhibited cytotoxic activities on K562 leukaemia and SK-N-DZ neuroblastoma cell lines. Furthermore, these compounds inhibited a panel of molecular targets involved in leukaemia and neuroblastoma tumorigenesis. All these results suggest that both long-chain imidazole-based ionic liquids and lysosomotropic detergents may be an effective alternative for the treatment of neuroblastoma and chronic myeloid leukemia and merit further investigation.

Cell-penetrating peptide and cationic liposomes mediated siRNA delivery to arrest growth of chronic myeloid leukemia cells in vitro

Gene silencing through RNA interference (RNAi) is a promising therapeutic approach for a wide range of disorders, including cancer. Non-viral gene therapy, using specific siRNAs against BCR-ABL1, can be a supportive or alternative measure to traditional chronic myeloid leukemia (CML) tyrosine kinase inhibitor (TKIs) therapies, given the prevalence of clinical TKI resistance. The main challenge for such approaches remains the development of the effective delivery system for siRNA tailored to the specific disease model.The purpose of this study was to examine and compare the efficiency of endosomolytic cell penetrating peptide (CPP) EB1 and PEG2000-decorated cationic liposomes composed of polycationic lipid 1,26-bis(cholest-5-en-3-yloxycarbonylamino)-7,11,16,20-tetraazahexacosane tetrahydrochloride (2Х3) and helper lipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) for anti-bcr-abl siRNA delivery into the K562 human CML cell line. We show that both EB1 and 2Х3-DOPE-DSPE-PEG2000 (0.62 % mol.) liposomes effectively deliver siRNA into K562 cells by endocytic mechanisms, and the use of liposomes leads to more effective inhibition of expression of the targeted gene (BCR-ABL1) and cancer cell proliferation. Taken together, these findings suggest that PEG-decorated cationic liposomes mediated siRNA delivery allows an effective antisense suppression of certain oncogenes, and represents a promising new class of therapies for CML.

Effects of Apigenin and Apigenin- Loaded Nanogel on Induction of Apoptosis in Human Chronic Myeloid Leukemia Cells

Background: Diet plays an important role in cancer prevention. Apigenin, a flavonoid with thechemical formula C15H10O5, is abundantly present in vegetables. Vegetarian foods containing flavonoids are rich sources of bioactive compounds. Flavonoids have been utilized in herbal treatment. Nanogels are drug delivery systems based on polymers and are used in tissue engineering and for drug delivery. This study was conducted to compare the effects of apigenin and a nanodrug on the viability of the K562 cell line of chronic myeloid leukemia at different durations under laboratory conditions. Materials and Methods: Chitosan was first dissolved in 1% acetic acid, and ethylene dichloride EDC and NHS were added to the solution. Then, the nanodrug was prepared by loading apigenin into stearate–chitosan nanogel (scs nanogel), and its physical and morphological characteristics were evaluated by TEM, DLS, and FTIR. Trypan blue staining, MTT assay, and flow cytometry were used to analyze the effects of various concentrations of apigenin and apigenin-loaded chitosan–stearate nanogel (APG–SCS) at 24, 48, and 72 h after they were applied to the K562 cell line. Results: The diameter of the nanodrug particles was measured using DLS and confirmed by TEM. The K562 cells treated with APG–SCS and with apigenin exhibited significant differences compared with the control (P < 0.05). Apoptosis was detected by flow cytometry. Conclusion: This study showed that the toxic effects of apigenin and the nanodrug improved with increasing concentrations and exposure durations compared to those in the control.The toxic effect of apigenin loaded into the stearate-chitosan nanogel was greater than apigenin, and the toxic effects of both materials were greater compared to the control under laboratory conditions.[GMJ.2018;7:e1008]

P-Terphenyl and Diphenyl Ether Derivatives from the Marine-Derived Fungus Aspergillus candidus HM5-4.

Two undescribed p-terphenyl derivatives, asperterphenylcins A-B (1-2), and two undescribed diphenyl ether derivatives, asperdiphenylcins A-B (3-4), together with three previously described p-terphenyl derivatives-4″-deoxyterprenin (5), terphenyllin (6), and 3″-hydroxyterphenyllin (7)-were obtained from the solid-rice culture of the marine-derived fungus Aspergillus candidus HM5-4, which was isolated from sponges from the South China Sea. Their structures were elucidated by HRESIMS data and NMR spectroscopic analysis. Compound 1 showed a strong inhibitory effect on Neoscytalidium dimidiatum, with an inhibition circle diameter of 31.67 ± 2.36 mm at a concentration of 10.0 µg/disc. Compounds 5 and 7 displayed cytotoxic activity against human chronic myeloid leukemia cells (K562), human liver cancer cells (BEL-7402), human gastric cancer cells (SGC-7901), human non-small cell lung cancer cells (A549) and human HeLa cervical cancer cells, with IC50 values ranging from 3.32 to 60.36 µM, respectively. Compounds 2, 6 and 7 showed potent inhibitory activity against α-glucosidase, with IC50 values of 1.26 ± 0.19, 2.16 ± 0.44 and 13.22 ± 0.55 µM, respectively.

Recognition of Phosphatidylserine Externalisation on Human Chronic Myeloid Leukaemia (K562) Cells Induced by Chrysanthemum morifolium Methanol Extract

Chrysanthemum morifolium is one of the common plants used in traditional Chinese herbal medicine and currently being discovered as an invaluable source for treatment of cancerous diseases. Conventional approach to leukaemia treatment such as chemotherapeutic agents results in good overall survival (OS) in patients but issues of multi-drug resistance (MDR) and disease heterogeneity can make chemotherapy less effective. Thus there is a constant need for alternative therapeutic strategy such as developing new anti-cancer drug from natural products. The present study is carried out mainly to detect phosphatidylserine externalisation as an apoptosis indicator on human chronic myeloid leukaemia (K562) cells induced by Chrysanthemum morifolium methanol extract. K562 cells were treated with indicated IC50 values of bud and flower methanol crude extracts for 24 hours. Phase contrast microscope was used to observe morphological changes in K562 cells post-treatment. Annexin V-FITC/PI apoptosis assay via flow cytometry was done to investigate apoptosis phenomena by detecting phosphatidylserine (PS) externalisation in treated K562 cells. Morphological observations using phase contrast microscopy showed distinct apoptotic characteristics in K562 cells treated with both bud and flower methanol extracts. Annexin V-FITC/PI assay demonstrated a significantly high apoptotic cell population and apoptotic rate (p <0.05) in treated K562 cells compared to control cells. The study outcomes presented promising anti-cancer ability for Chrysanthemum morifolium.

In-house glutamine based compound loaded poly-(D-L-lactide-coglycolide) nanoparticles as a possible candidate for the management of chronic myeloid leukemia

The development of targeted medicines such tyrosine kinase inhibitors (TKIs) have turned chronic myeloid leukemia (CML) from a potentially fatal illness into a manageable chronic condition for many individuals. But in case of TKIs, some serious drug induced adverse events as well as drug resistance were reported. In continuation of our earlier work, we have synthesized two p-nitrobenzenesulfonyl-D-glutamine derivatives which were screened against human CML cell line (K562). In comparison with earlier reported p-nitrobenzenesulfonyl-L-glutamine derivatives (Compound 1 and Compound 2), newly synthesized compounds showed more than 3 times better activity against K562 cell line. The best active compound (Compound 3) was encapsulated into Poly-(D-L-lactide-coglycolide) nanoparticles (NPs). The NPs were characterized and screened against K562 and human PBMC cell line. The IC50 value of NP of Compound 3 was found in the nanomolar range (522 nM) against K562 cell line which is more than 10 times better active than Compound 3. DNA deformation assay also indicates cell shrinkage and nuclear fragmentation which are associated with apoptosis-mediated cell death. The prepared NPs of Compound 3 (NP3) are very much less toxic against normal human PBMC (IC50 value > 500 µM). The current study implies that NP3 holds promise for managing chronic myeloid leukemia.

8-Hydroxydaidzein Induces Apoptosis and Inhibits AML-Associated Gene Expression in U-937 Cells: Potential Phytochemical for AML Treatment.

8-hydroxydaidzein (8-OHD) is a compound derived from daidzein, known for its anti-inflammatory and anti-proliferative properties in K562 human chronic myeloid leukemia (CML) cells. However, its effects on acute myeloid leukemia (AML) cells have not been fully understood. To investigate its potential anti-AML mechanism, we employed an integrated in vitro-in silico approach. Our findings demonstrate that 8-OHD suppresses the expression of CDK6 and CCND2 proteins and induces cell apoptosis in U-937 cells by activating Caspase-7 and cleaving PARP-1. Microarray analysis revealed that 8-OHD downregulates differentially expressed genes (DEGs) associated with rRNA processing and ribosome biogenesis pathways. Moreover, AML-target genes, including CCND2, MYC, NPM1, FLT3, and TERT, were downregulated by 8-OHD. Additionally, molecular docking software predicted that 8-OHD has the potential to interact with CDK6, FLT3, and TERT proteins, thereby reducing their activity and inhibiting cell proliferation. Notably, we discovered a synergic pharmacological interaction between 8-OHD and cytarabine (Ara-C). Overall, this study provides insights into the therapeutic applications of 8-OHD in treating AML and elucidates its underlying mechanisms of action.

A novel cationic arginine-modified Self-Nanoemulsifying drug delivery system (SNEDDS) for improved anticancer and antioxidant activities of Naringin

Naringin (NRG) is a flavonoid derived from citrus fruits. Despite its pharmacological significance, NRG's therapeutic effectiveness is constrained by its unusual physicochemical characteristics, such as its lower aqueous solubility. This study was intended to develop an alkylated arginine (AL-ARG) containing self-nanoemulsifying drug delivery system (SNEDDS) serving as a positively charged amphiphile for enhanced anticancer and antioxidant activities of NRG. The drug solubility along with emulsification investigations resulted in the development of a ternary phase diagram for selected suitable formulation components. Lipid (cinnamon oil), surfactant (labrasol), and co-surfactant (Transcutol) were selected as the main components of the formulation. FTIR, zetasizer, and atomic force microscope were employed to characterize optimized formulations, which were then tested for dilution and thermodynamic stability. It was found that the average droplet size and zeta potential of NRG-SNEDDS were 118.47 ± 21.16 nm and −21 ± 2.10 mV, respectively, while those of AL-ARG-incorporated NRG-SNEDDS were 162.17 ± 19.11 nm and −17 ± 1.83 mV. The developed SNEDDS were extremely stable and did not change the chemical nature of the drug. The developed SNEDDS were tested for their antioxidant activity, and the findings showed that the antioxidant potential of NRG was greatly improved by its incorporation into the AL-ARG-modified SNEDDS. When AL-ARG was incorporated into NRG-SNEDDS, its anticancer potential was greatly increased against the human chronic myeloid leukemia cell. It can be concluded that novel cationic arginine-modified SNEDDS have the potential to substantially enhance the antioxidant and anticancer properties of NRG.

Inactivation of TRPM7 Kinase Targets AKT Signaling and Cyclooxygenase-2 Expression in Human CML Cells.

Cyclooxygenase-2 (COX-2) is a key regulator of inflammation. High constitutive COX-2 expression enhances survival and proliferation of cancer cells, and adversely impacts antitumor immunity. The expression of COX-2 is modulated by various signaling pathways. Recently, we identified the melastatin-like transient-receptor-potential-7 (TRPM7) channel-kinase as modulator of immune homeostasis. TRPM7 protein is essential for leukocyte proliferation and differentiation, and upregulated in several cancers. It comprises of a cation channel and an atypical α-kinase, linked to inflammatory cell signals and associated with hallmarks of tumor progression. A role in leukemia has not been established, and signaling pathways are yet to be deciphered. We show that inhibiting TRPM7 channel-kinase in chronic myeloid leukemia (CML) cells results in reduced constitutive COX-2 expression. By utilizing a CML-derived cell line, HAP1, harboring CRISPR/Cas9-mediated TRPM7 knockout, or a point mutation inactivating TRPM7 kinase, we could link this to reduced activation of AKT serine/threonine kinase and mothers against decapentaplegic homolog 2 (SMAD2). We identified AKT as a direct in vitro substrate of TRPM7 kinase. Pharmacologic blockade of TRPM7 in wildtype HAP1 cells confirmed the effect on COX-2 via altered AKT signaling. Addition of an AKT activator on TRPM7 kinase-dead cells reconstituted the wildtype phenotype. Inhibition of TRPM7 resulted in reduced phosphorylation of AKT and diminished COX-2 expression in peripheral blood mononuclear cells derived from CML patients, and reduced proliferation in patient-derived CD34+ cells. These results highlight a role of TRPM7 kinase in AKT-driven COX-2 expression and suggest a beneficial potential of TRPM7 blockade in COX-2-related inflammation and malignancy.

Arginine dependency is a therapeutically exploitable vulnerability in chronic myeloid leukaemic stem cells.

To fuel accelerated proliferation, leukaemic cells undergo metabolic deregulation, which can result in specific nutrient dependencies. Here, we perform an amino acid drop-out screen and apply pre-clinical models of chronic phase chronic myeloid leukaemia (CML) to identify arginine as a nutrient essential for primary human CML cells. Analysis of the Microarray Innovations in Leukaemia (MILE) dataset uncovers reduced ASS1 levels in CML compared to most other leukaemia types. Stable isotope tracing reveals repressed activity of all urea cycle enzymes in patient-derived CML CD34+ cells, rendering them arginine auxotrophic. Thus, arginine deprivation completely blocks proliferation of CML CD34+ cells and induces significantly higher levels of apoptosis when compared to arginine-deprived cell lines. Similarly, primary CML cells, but not normal CD34+ samples, are particularly sensitive to treatment with the arginine-depleting enzyme, BCT-100, which induces apoptosis and reduces clonogenicity. Moreover, BCT-100 is highly efficacious in a patient-derived xenograft model, causing > 90% reduction in the number of human leukaemic stem cells (LSCs). These findings indicate arginine depletion to be a promising and novel strategy to eradicate therapy resistant LSCs.

Amsacrine downregulates BCL2L1 expression and triggers apoptosis in human chronic myeloid leukemia cells through the SIDT2/NOX4/ERK/HuR pathway

Accumulating evidence indicates that the anticancer activity of acridine derivatives is mediated through the regulation of anti-apoptotic and pro-apoptotic BCL2 protein expression. Therefore, we investigated whether the cytotoxicity of amsacrine with an acridine structural scaffold in human chronic myeloid leukemia (CML) K562 cells was mediated by BCL2 family proteins. Amsacrine induced apoptosis, mitochondrial depolarization, and BCL2L1 (also known as BCL-XL) downregulation in K562 cells. BCL2L1 overexpression inhibited amsacrine-induced cell death and mitochondrial depolarization. Amsacrine treatment triggered SIDT2-mediated miR-25 downregulation, leading to increased NOX4-mediated ROS production. ROS-mediated inactivation of ERK triggered miR-22 expression, leading to increased HuR mRNA decay. As HuR is involved in stabilizing BCL2L1 mRNA, downregulation of BCL2L1 was noted in K562 cells after amsacrine treatment. In contrast, amsacrine-induced BCL2L1 downregulation was alleviated by restoring ERK phosphorylation and HuR expression. Altogether, the results of this study suggest that amsacrine triggers apoptosis in K562 cells by inhibiting BCL2L1 expression through the SIDT2/NOX4/ERK-mediated downregulation of HuR. Furthermore, a similar pathway also explains the cytotoxicity of amsacrine in CML MEG-01 and KU812 cells.

KOENZİM Q0 İNSAN KRONİK MYELOİD LÖSEMİ K562 HÜCRELERİNİN PROLİFERASYONUNU ENGELLER VE MAPK VE AKT SİNYAL YOLAKLARINI MODÜLE EDER

Objective: This study evaluated the antiproliferative and pro-apoptotic effects of coenzyme Q0 (CoQ0) in human chronic myeloid leukemia K562 cell line. Material and Method: The cytotoxic effect of CoQ0 on human chronic myeloid leukemia cell line, K562 was determined by MTT test. The activity of caspase-3, expression of proteins involved in apoptosis, MAPK and AKT signaling pathways were determined with enzymatic assay and western blot analysis, respectively. Result and Discussion: Results showed that CoQ0 inhibited cell viability of K562 cells at 5 μM and higher concentrations and Bax protein expression was significantly decreased at 12.5 μM concentration of CoQ0. However, CoQ0 did not significantly affect caspase 3 activity and Bcl-2 protein expression. p-c-Raf (Ser259) protein expression was significantly decreased at 12.5 μM of CoQ0. Treatment with 10 μM of CoQ0 induced significantly phosphorylation of p38 MAPK and 12.5 μM CoQ0 caused a nonsignificant decrease in p-ERK1/2 protein expression in K562 cell line. Interestingly, in K562 cells, phosphorylation of Akt (Ser473) was diminished at 12.5 μM of CoQ0, with no change observed in p-Akt (Thr308) protein expression among groups. In conclusion, CoQ0 inhibited cell proliferation and suppressed phosphorylation of c-Raf (Ser259), Akt (Ser473), but not ERK1/2 in K562 cells. There is still a need for new insights into the anticancer mechanisms of CoQ0 and develop treatment strategies for chronic myeloid leukemia.

Utilization of Dairy By-Products as a Source of Functional and Health Compounds-The Role of Ovine Colostrum and Milk Whey on Chronic Myeloid Leukemia Cells.

Nowadays, the search for food products that promote consumers' health has gained interest, and dairy by-products, due to their biological quality, could have a prominent position among products with health benefits. However, little is known about their activity on cancer cells. This study aimed to provide evidence about the effect of ovine colostrum and milk whey on K562 cells, a model of the human chronic myeloid leukemia cell line. The exposure of K562 cells to a single administration of sheep by-products at different concentrations for three days and three treatments for three days was carried out. Using a flow cytometric approach, we found that CD235a expression remained stable in the cells exposed to ovine whey (milk and colostrum) at concentrations ranging from 1 ng/mL to 100 μg/mL, after three days from one or three administrations, respectively. A significant reduction in fluorescent cells was observed in the populations exposed to 1 mg/mL of both milk and colostrum at the same time points. In these conditions, the size and granularity of the leukemic cells also changed, with a substantial reduction in the number of actively dividing cells in the S phase of the cell cycle. This phenomenon was highlighted by the Annexin V/PI cytofluorimetric test, which is able to provide quantitative results regarding the population of cells in early or late apoptosis or necrotic cells after exposure to a single dose or three doses of colostrum or sheep whey for three days, respectively. This report showed that both colostrum and milk whey were able to modify the phenotypic profile and cell cycle of the K562 cell line, inducing apoptosis at the highest concentration.

Overcoming ABCB1-mediated multidrug resistance by transcription factor BHLHE40

Multidrug resistance (MDR) hinders treatment efficacy in cancer therapy. One typical mechanism contributing to MDR is the overexpression of permeability-glycoprotein (P-gp) encoded by ATP-binding cassette subfamily B member 1 (ABCB1). Basic helix-loop-helix family member e40 (BHLHE40) is a well-known transcription factor that has pleiotropic effects including the regulation of cancer-related processes. However, whether BHLHE40 regulates MDR is still unknown. Chromatin immunoprecipitation-seq study revealed BHLHE40 occupancy in the promoter of ABCB1 gene. Adriamycin (ADM)-resistant human chronic myeloid leukemia cells (K562/A) and human breast cancer cells (MCF-7/A) were established. BHLHE40 expression was downregulated in the ADM-resistant cell lines. Overexpression of BHLHE40 resensitized resistant cells to ADM, promoted cell apoptosis in vitro and suppressed tumor growth in vivo, whereas BHLHE40 knockdown induced resistance to ADM in parental cells. Moreover, we found that BHLHE40 regulated drug resistance by directly binding to the ABCB1 promoter (-1605 to -1597) and inactivating its transcription. In consistence, the expression of BHLHE40 was negatively correlated with ABCB1 in various cancer cells, while positively with cancer cell chemosensitivity and better prognosis of patients with breast cancer. The study reveals the role of BHLHE40 as a transcriptional suppressor on the expression of ABCB1, major ABC transporter in chemoresistance. The findings extend the function of BHLHE40 in tumor progression and provides a novel mechanism for the reversal of multidrug resistance.

Novel indole retinoid derivative induces apoptosis and cell cycle arrest and modulates AKT and ERK signaling in HL-60 cells.

Chemotherapy with targeted drugs is the first line therapy option for acute and chronic myeloid leukemia. However, hematopoietic stem cell transplantation may be used in high-risk patients or patients with failed responses to chemo drugs. Discovery and development of more effective new agents with lower side effects is the main aim of leukemia treatment. In this study, a novel retinoid compound with tetrahydronaphthalene ring was synthesized and evaluated for anticancer activity in human chronic and acute myeloid leukemia cell lines K562 and HL-60. Novel N-(1H-indol-1-yl)-5,5,8,8-tetramethyl-5,6,7,8-tetrahydronaphthalene-2-carboxamide was synthesized based on molecular hybridization of the two different bioactive structures retinoid head and indole. The effects of the synthesized carboxamide compound, which was referred to as compound 5, were determined in K562 chronic myeloid leukemia and HL-60 acute myeloid leukemia cell lines and L929 fibroblast cell line, which served as a control. Colorimetric MTT and caspase3 activity tests, flow cytometry, western blot, and microscopic examinations were used to evaluate biological activity. Compound 5 more effectively induced cell death in HL60 cells in comparison to K562 cells and L929 fibroblast cells. Therefore, further mechanism of cell death was investigated in HL60 cell line. It was found that compound 5 induced remarkable cytotoxicity, caspase3 activation, and PARP fragmentation in HL60 cells. Flow cytometric staining showed that the percentage of cells arrested in G0/G1 was also increased with compound 5 treatment. Important modulator proteins of cell proliferation p-ERK, p-AKT, and p-m-TOR were also found to be inhibited with compound 5 treatment. Collectively, our results reveal compound 5, which is a novel indole retinoid compound as a potential active agent for the treatment of acute promyelocytic leukemia.

Cepharanthine inhibits dengue virus production and cytokine secretion

Dengue virus (DENV) infection is a public health problem in tropical and subtropical regions. It can cause a spectrum of clinical manifestations ranging from mild dengue fever (DF) to severe dengue haemorrhagic fever (DHF) and potentially life-threatening disease including dengue shock syndrome (DSS). Severe DENV infection is caused by high viral load and cytokine storm in dengue-infected patients. Currently, there is no specific antiviral drug for DENV infection. An anti-DENV agent that demonstrates inhibitory effects on both DENV replication and cytokine secretion is urgently needed. In this study, cepharanthine (CEP), which is an anti-inflammatory, anti-HIV, and anti-tumor compound isolated from Stephania cepharantha Hayata, was tested for inhibition of DENV infection. We investigated the efficacy of CEP to inhibit DENV infection, replication, and cytokine production. The inhibitory effect of CEP treatment was studied in DENV-infected human chronic myeloid leukemia (K562) cells. The levels of DENV E protein and DENV production were determined by flow cytometry and FFU assay, respectively. CEP treatment significantly reduced viral E protein and viral production in all DENV-1, 2, 3, 4 serotypes. In addition, CEP treatment reduced the IL-6 proinflammatory cytokine production in DENV-infected A549 cells. Taken together, CEP has inhibitory effects on DENV infection specifically at the initial viral replication states and proinflammatory cytokine secretion, and is a promising candidate for further development as an anti-DENV treatment.

Naturally Occurring Bicoumarin Compound Daphnoretin Inhibits Growth and Induces Megakaryocytic Differentiation in Human Chronic Myeloid Leukemia Cells.

Daphnoretin extracted from the stem and roots of Wikstroemia indica (L.) C.A. Mey has been shown to possess antiviral and antitumor activities. Herein, we hypothesized that daphnoretin might induce megakaryocytic differentiation, thereby inhibiting the proliferation of cells and serving as a differentiation therapy agent for chronic myeloid leukemia (CML). Daphnoretin-treated K562 and HEL cells were examined for growth inhibition, cell morphology, and megakaryocyte-specific markers. Potential mechanisms of megakaryocytic differentiation of daphnoretin-treated K562 cells were evaluated. The results showed that daphnoretin inhibited the growth of K562 and HEL cells in a dose- and time-dependent manner. Flow cytometry analyses revealed that daphnoretin treatment slightly increased the proportion of sub-G1 and polyploid cells compared to that of dimethyl sulfoxide (DMSO)-treated control cells. Morphological examination showed that daphnoretin-treated K562 and HEL cells exhibited enlarged contours and multinucleation as megakaryocytic characteristics compared to DMSO-treated control cells. Daphnoretin treatment also dramatically enhanced the expression of megakaryocytic markers CD61 and CD41. Under optimal megakaryocytic differentiation conditions, daphnoretin increased the phosphorylation of STAT3 but not STAT5. In summary, daphnoretin inhibited cell growth and induced megakaryocytic differentiation in K562 and HEL cells. The efficacy of daphnoretin in vivo and in patients with CML may need further investigations for validation.

PHYTOCHEMICAL CONSTITUENTS AND ANTILEUKEMIC EFFECTS OF JUNIPERUS OXYCEDRUS EXTRACT

Aim: Many genetic and environmental factors can be effective in the process of cancerization. Preventing the progression of leukemia may be possible by controlling the pathways involving mechanisms such as apoptosis and autophagy. When the literature is examined, there are studies showing the effects of various types of juniper on various cancer cell lines, including human chronic myeloid leukemia cells, but the signal pathways in which they act are not fully known. In this study, the anticancer effects of Juniperus oxycedrus extract on K-562 human chronic myeloid leukemia cells were investigated. Method: After the cells were treated with the Juniperus oxycedrus extract, cytotoxicity and gene expression analyzes were performed. Changes in the expression of Akt, the member of the PI3K/Akt/mTOR signaling pathway; caspase 3, which is one of the main effective genes in the pathways regulating apoptosis; and the apoptosis suppressor BCL-2 gene, which is an oncogene, were investigated. Results: According to the MTT test results, Juniperus oxycedrus extract showed over approximately 50% cell viability in K-562 cells at all doses. The most appropriate dose of Juniperus oxycedrus fruit extract in this research was determined as 50 µg/ mL considering cell viability. After the gene expression analysis, it was observed that Akt expression increased 1.092 times, BCL-2 expression decreased approximately 0.3 times, and caspase 3 expression increased 1.2 times. Conclusions: Constituents of Juniperus oxycedrus plant may have apoptotic effects on chronic myeloid leukemia cells.

Ketoconazole Reverses Imatinib Resistance in Human Chronic Myelogenous Leukemia K562 Cells.

Chronic myeloid leukemia (CML) is a hematologic disorder characterized by the oncogene BCR-ABL1, which encodes an oncoprotein with tyrosine kinase activity. Imatinib, a BCR-ABL1 tyrosine kinase inhibitor, performs exceptionally well with minimal toxicity in CML chemotherapy. According to clinical trials, however, 20–30% of CML patients develop resistance to imatinib. Although the best studied resistance mechanisms are BCR-ABL1-dependent, P-glycoprotein (P-gp, a drug efflux transporter) may also contribute significantly. This study aimed to establish an imatinib-resistant human CML cell line, evaluate the role of P-gp in drug resistance, and assess the capacity of ketoconazole to reverse resistance by inhibiting P-gp. The following parameters were determined in both cell lines: cell viability (as the IC50) after exposure to imatinib and imatinib + ketoconazole, P-gp expression (by Western blot and immunofluorescence), the intracellular accumulation of a P-gp substrate (doxorubicin) by flow cytometry, and the percentage of apoptosis (by the Annexin method). In the highly resistant CML cell line obtained, P-gp was overexpressed, and the level of intracellular doxorubicin was low, representing high P-gp activity. Imatinib plus a non-toxic concentration of ketoconazole (10 μM) overcame drug resistance, inhibited P-gp overexpression and its efflux function, increased the intracellular accumulation of doxorubicin, and favored greater apoptosis of CML cells. P-gp contributes substantially to imatinib resistance in CML cells. Ketoconazole reversed CML cell resistance to imatinib by targeting P-gp-related pathways. The repurposing of ketoconazole for CML treatment will likely help patients resistant to imatinib.