Human Promyelocytic Leukemia HL-60 Research Articles

The Pyridone-Annelated Isoindigo (5‘-Cl) Induces Apoptosis, Dysregulation of Mitochondria and Formation of ROS in Leukemic HL-60 Cells

Background/Aims: In our quest to develop an isoindigo with improved efficacy and bioavailability, we recently synthesized a series of novel substituted pyridone-annelated isoindigo and evaluated their antiproliferative effects. We identified the compound [(E)-1-(5'-Chloro-2'-oxoindolin-3'-ylidene)-6-ethyl-2,3,6,9-tetrahydro-2,9-dioxo-1H-pyrrolo[3,2-f] quinoline-8-carboxylic acid], abbreviated as 5'-Cl, which shows selective antiproliferative activities against various cancer cell lines mediated through apoptosis. Here we have investigated the molecular mechanisms underlying the apoptotic activity of 5'-Cl in the human promyelocytic leukemia HL-60 cells. Methods: We employed different methods to determine the apoptotic pathways triggered by 5'-Cl in HL-60 cells, using flow cytometry, nuclear staining, caspases activation, mitochondria functioning, generation of reactive oxygen species (ROS) and Western blotting techniques. Results: Low concentrations (1-8 µM) of 5'-Cl inhibited the growth of HL-60 cells in a dose and time-dependent manner. Cytotoxicity of this compound is found to be mediated by a caspase-dependent apoptosis. Also, there were indications of caspase independent apoptosis as z-VAD-FMK failed to fully rescue the cells from 5‘-Cl-induced apoptosis. In addition, the compound triggered generation of Reactive Oxygen Species (ROS), caused depolarization of the mitochondrial inner membrane, decreased the level of cellular ATP, modulated the expression and phosphorylation of Bcl-2 leading to loss of its association with Bax and increased the release of cytochrome c to the cytosol of treated cells. The effects of 5‘-Cl on mitochondria and apoptosis were substantially blocked in the presence of a combination between z-VAD-FMK and either of the ROS scavenger N-acetyl-L-cysteine (NAC) or pyrrolidine dithiocarbamate (PDTC). Conclusion: We demonstrated that the growth inhibitory effects of 5'-Cl in HL-60 cells involve multiple pathways of apoptosis and dysregulation of mitochondrial functions.

The Disulfide Analogues of Isophosphoramide Mustard for Anticancer Therapy

Novel disulfide analogues of isophosphoramide mustard (iPAM) were designed and synthesised. All compounds were hydrolytically stable and underwent reduction by L-glutathione with different kinetic parameters. Based on the HPLC-MS analysis a mechanism of activation by glutathione obtained disulfide analogues of iPAM was proposed. The compounds were tested for cytotoxic activity against human promyelocytic leukaemia HL-60, human lymphoblastic leukaemia MOLT-4, human lymphoblastic leukaemia CCRF/CEM, human bladder cancer HCV29T, murine melanoma B16-F0 and murine fibroblasts Balb3T3 cell lines. The most promising anticancer activity was exhibited by compound 4, which proved to be more active than the reference cisplatin against all cancer cell lines. Keywords: Anticancer activity, disulfide, glutathione, isophosphoramide mustard.

Nardostachys chinensis induces granulocytic differentiation with the suppression of cell growth through p27Kip1 protein-related G0/G1 phase arrest in human promyelocytic leukemic cells

Context: Nardostachys chinensis Batalin (Valerianaceae) has been used in Korean traditional medicine to elicit stomachic and sedative effects. However, the anti-leukemic activities of N. chinensis have not been well examined.Objective: To investigate the effect of N. chinensis on differentiation and proliferation in the human promyelocytic leukemic HL-60 cells.Materials and methods: The dried roots and stems of N. chiensis are extracted using hot water and then freeze-dried. The yield of extract was 12.82% (w/w). The HL-60 cells were treated with 25–200 μg/ml of N. chinensis for 72 h or 100 μg/ml of N. chinensis for 24–72 h.Results: Nardostachys chinensis significantly inhibited cell viability dose dependently with an IC50 of 100 μg/ml in HL-60 cells. Nardostachys chinensis induced differentiation of the cells as measured by reduction activity of NBT and expression of CD11b but not of CD14 as analyzed by flow cytometry, which indicates a differentiation toward the granulocytic lineage. Nardostachys chinensis also induced growth inhibition through G0/G1 phase arrest in the cell cycle of HL-60 cells. Among the G0/G1 phase in the cell cycle-related protein, the expression of cyclin-dependent kinase (CDK) inhibitor p27Kip1 was increased in N. chinensis-treated HL-60 cells, whereas the expression levels of CDK2, CDK4, CDK6, cyclin D1, cyclin D3, cyclin E, and cyclin A were decreased. Interestingly, N. chinensis markedly enhanced the binding of p27Kip1 with CDK2 and CDK6.Discussion and conclusion: This study demonstrated that N. chinensis is capable of inducing cellular differentiation and growth inhibition through p27Kip1 protein-related G0/G1 phase arrest in HL-60 cells.

Programmed cell death induced by (-)-8,9-dehydroneopeltolide in human promyelocytic leukemia HL-60 cells under energy stress conditions.

(+)-Neopeltolide is a marine macrolide natural product that exhibits potent antiproliferative activity against several human cancer cell lines. Previous study has established that this natural product primarily targets the complex III of the mitochondrial electron transport chain. However, the biochemical mode-of-actions of neopeltolide have not been investigated in detail. Here we report that (−)-8,9-dehydroneopeltolide (8,9-DNP), a more accessible synthetic analogue, shows potent cytotoxicity against human promyelocytic leukemia HL-60 cells preferentially under energy stress conditions. Nuclear morphology analysis, as well as DNA ladder assay, indicated that 8,9-DNP induced significant nuclear condensation/fragmentation and DNA fragmentation, and these events could be suppressed by preincubating the cells with a pan-caspase inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (zVAD). Immunoblot analysis demonstrated the release of cytochrome c from the mitochondria and the cleavage of full-length caspase-3 and poly(ADP-ribose) polymerase (PARP). These results indicated that 8,9-DNP induced caspase-dependent apoptotic programmed cell death under energy stress conditions. It was also found that 8,9-DNP induced non-apoptotic cell death in the presence/absence of zVAD under energy stress conditions. Immunoblot analysis showed the intracytosolic release of apoptosis-inducing factor (AIF), although it did not further translocate to the nucleus. It appears most likely that, in the presence of zVAD, 8,9-DNP triggered necrotic cell death as a result of severe intracellular ATP depletion.

Isolation and total syntheses of cytotoxic cryptolactones A1, A2, B1, and B2: α,β-unsaturated δ-lactones from a Cryptomyzus sp. aphid.

The cryptolactones A1, A2, B1, and B2, which are α,β-unsaturated δ-lactones, were isolated from a Cryptomyzus sp. aphid. The structures were established by 1-D and 2-D NMR spectra and CI-HRMS. Their absolute configurations were determined with the Kusumi-Mosher method, combined with asymmetric total syntheses. The syntheses were accomplished with the Mukaiyama aldol reaction and olefin metathesis, which utilized the second-generation Grubbs catalyst for the key steps. These compounds exhibited cytotoxic activity against human promyelocytic leukemia HL-60 cells with IC50 values of 0.97-5.3 μM.

3,6-Epidioxy-1,10-bisaboladiene inhibits G1 -specific transcription through Swi4/Swi6 and Mbp1/Swi6 via the Hog1 stress pathway in yeast.

•Swi4 physically interacts with Swi6 by anti tag coimmunoprecipitation (View interaction).

Annexin A3 plays a role in cytoplasmic calcium oscillation by extracellular calcium in the human promyelocytic leukemia HL-60 cells differentiated by phorbol-12-myristate-13-acetate

The roles of annexin A3 (ANXA3) in macrophages are not fully understood. In contrast to C5a, we have demonstrated that C-terminal ribosomal protein S19 (RP S19)-tagged S-tagged C5a (S-tagged C5a/RP S19) raises an alternative cytoplasmic calcium oscillation by extracellular calcium during macrophage migration into apoptotic cells. We here differentiated human promyelocytic leukemia HL-60 cells bearing with either control sense RNA and shRNA for ANXA3 mRNA or a vector cDNA with or without ANXA3 cDNA into macrophage-like cells by phorbol-12-myristate-13-acetate and found that a fluorescence ratio (340nm/380nm) upon the S-tagged C5a/RP S19-induced alternative cytoplasmic calcium oscillation by extracellular calcium was an equilateral association with a dose of ANXA3. Moreover, the ANXA3-dependent modification was partially reflected upon the S-tagged C5a-induced classical cytoplasmic calcium oscillation by both intracellular calcium and extracellular calcium. ANXA3 seems to extend the C5aR-mediated cytoplasmic calcium oscillation by extracellular calcium at least in the HL-60 macrophage-like cells.

Bioactive Steroids from a Marine-Derived Fungus Penicillium sp. from the South China Sea

Marine organisms have been found to be a source of steroids, particularly in terms of unique side-chain structures and unusual functionalization [1]. Marine steroids are often found in highly oxygenated forms, and such steroids sometimes show a variety of biological and pharmacological activities [2]. Despite the number of marine-derived steroids that have been found to date, marine-derived fungi appear to be an infertile source of novel bioactive steroids [3]. In the course of our ongoing investigation on bioactive secondary metabolites from marine fungi in the South China Sea, a sponge-derived fungus Penicillium sp. attracted our attention because of the fact that the EtOAc extract of the fungal culture showed pronounced cytotoxic activity against K562 chronic leukemia cell line. Investigation of the active extract of this fungus led to the isolation of 12 steroids (1–12). The structures of these steroids were determined on the basis of their 1H NMR, 13C NMR, and ESI-MS spectroscopic data, and by comparison with those previously reported in the literature, as six steroids with ketone carbonyl groups, dankasterone A (1) [4], dankasterone B (2) [4], 3 ,15 -dihydroxyl-(22E,24R)-ergosta-5,8(14),22-trien-7-one (3) [5], 3 ,15 -dihydroxyl-(22E,24R)-ergosta-5,8(14),22-trien-7-one (4) [5], 6 -hydroxy-ergosta-4,7,22-trien-3-one (5) [6], and 6 -hydroxy-stigmast-4-en-3-one (6) [7]; two epoxy-ergosterols, 5 ,6 -epoxy-(22E,24R)-ergosta-8(14),22-diene-3 ,7 -diol (7) [8] and 5 ,6 -epoxy-(22E,24R)-ergosta-8,22-diene-3 ,7 -diol (8) [8]; two epidioxy-ergosterols, 5 ,8 -epidioxy-ergosta6,22E-dien-3 -ol (9) [9] and 5 ,8 -epidioxy-ergosta-6,9,22E-trien-3 -ol (10) [9]; and two polyoxygenated sterols, 3 ,5 -dihydroxy-6 -methoxyergosta-7,22-diene (11) [10], and 3 ,5 ,6 ,9 -tetrahydroxyergosta-7,22-diene (12) [10]. The cytotoxic activity of all isolated compounds was evaluated against human promyelocytic leukemia HL-60, human lung carcinoma A-549, chronic leukemia K562, and human cervical carcinoma Hela cell lines. Compound 1 showed pronounced cytotoxic activity against HL-60, Hela, and K562 with IC50 values of 0.78, 4.11, and 7.57 M, respectively, especially to HL-60, which was stronger than that of the positive control adriamycin (IC50 0.98 M). Compound 2 also exhibited strong growth inhibition against HL-60, Hela, and K562 with IC50 values of 3.25, 4.74, and 7.89 M. Compounds 3 and 4 showed significant cytotoxic activity against K562 with IC50 values of 4.38 and 5.54 M, respectively. These results suggested that ketone carbonyl groups in compounds 1–4 should play an important role for enhancement of cancer cell growth inhibition. The antibacterial activity of all isolated compounds was also evaluated with a panel of pathogenic bacteria, including Gram-positive Staphylococcus aureus, S. albus, Bacillus cereus, Micrococcus tetragenus, and Kocuria rhizophila, and Gram-negative Escherichia coli, Vibrio parahaemolyticus, V. anguillarum, and Pseudomonas putida (Table 1). The results indicated that the isolated steroids (1–5, 7–11) exhibited widely significant antibacterial effects against most of pathogenic bacteria except for S. albus and V. parahaemolyticus. Among these steroids, compound 1 was found to be the most active and showed a broad spectrum of antibacterial activity, while compounds 2–5 and 7–11 exhibited selective antibacterial activity. In particular, compounds 1, 4, 7, and 8 showed strong antibacterial activity against V. anguillarum with MIC values of 0.39, 0.78, 0.78, and 0.39 M, respectively, which were stronger than or equivalent to that of the positive control ciprofloxacin. It is noteworthy that the activity of 1 was stronger than that of 2, and the only difference between their structures was the absence of double bond at C-4/C-5 in 2, indicating that the double bond at C-4/C-5 in 13(14 8)abeo-8-ergostane had a significant effect on the antibacterial activity. The results of antibacterial activity also suggested that the ketone carbonyl groups, epoxy rings, and peroxy groups in steroids played apparent role on the antibacterial activity. Moreover, compound 12 with four hydroxy groups was less active than 11, suggesting that the simultaneous presence of several hydroxy groups caused loss of activity.

Seed Dormancy Breaking Diterpenoids from the Liverwort Plagiochila sciophila and their Differentiation Inducing Activity in Human Promyelocytic Leukemia HL-60 Cells

To obtain the structural diversity of bioactive compounds similar to cotylenins and fusicoccins that modulate 14-3-3 protein-protein interactions in eukaryotes, screening tests were carried out using the lettuce seed dormancy breaking-assay. An acetone extract of the liverwort Plagiochila sciophila exhibited significant activity against the seeds in the presence of the plant hormone abscisic acid. Activity-guided fractionation of the extract afforded the isolation of seven novel fusicoccane-type diterpenoids, named fusicosciophins A-E (1-5), 8-deacetyl (6) and 9-deacetyl fusicosciophin E (7). Their structures were determined by spectroscopic methods and X-ray crystallographic analyses. All the pure isolated compounds (1-7) exhibited moderate lettuce seed dormancy breaking activity. In addition, the differentiation-inducing activity and cytotoxicity of these isolates, together with fusicoccin A (FC-A) and all-trans retinoic acid (ATRA), were evaluated in human promyelocytic leukemia HL-60 cells and human mouth epidermal carcinoma KB cells, respectively. Fusicosciophins (2 and 4) and FC-A exhibited moderate differentiation-inducing activity (EC50 31.2-59.1 microM) compared with ATRA (EC50 0.3 microM), while 2, 4 and ATRA exhibited higher selectivity indices (IC50/EC50 >3.38-667) than FC-A (IC50/EC50 1.05). This is the first report on the isolation of fusicoccane-type diterpenoids from liverworts having seed dormancy breaking activity and differentiation-inducing activity in mammal cells.

Euchromatic histone methyltransferase 2 inhibitor, BIX-01294, sensitizes human promyelocytic leukemia HL-60 and NB4 cells to growth inhibition and differentiation.

The involvement of histone lysine methyltransferases (HMT) in carcinogenesis is not well understood. Here, we describe a dose-dependent growth and survival inhibitory effects of BIX-01294, a specific inhibitor of euchromatic HMT2, in promyelocytic leukemia HL-60 and NB4 cells. BIX-01294 combined with all-trans retinoic acid or together with histone deacetylase and DNA methyltransferase inhibitors enhanced cell differentiation to granulocytes and induced cell line-specific changes in the expression of cell cycle-, survival- and differentiation regulating genes and proteins in association with histone modification state. Our results suggest that targeting EHMT2 may be of therapeutical benefits in myeloid leukemia.

Tempol, an Intracellular Antioxidant, Inhibits Tissue Factor Expression, Attenuates Dendritic Cell Function, and Is Partially Protective in a Murine Model of Cerebral Malaria

BackgroundThe role of intracellular radical oxygen species (ROS) in pathogenesis of cerebral malaria (CM) remains incompletely understood.Methods and FindingsWe undertook testing Tempol—a superoxide dismutase (SOD) mimetic and pleiotropic intracellular antioxidant—in cells relevant to malaria pathogenesis in the context of coagulation and inflammation. Tempol was also tested in a murine model of CM induced by Plasmodium berghei Anka infection. Tempol was found to prevent transcription and functional expression of procoagulant tissue factor in endothelial cells (ECs) stimulated by lipopolysaccharide (LPS). This effect was accompanied by inhibition of IL-6, IL-8, and monocyte chemoattractant protein (MCP-1) production. Tempol also attenuated platelet aggregation and human promyelocytic leukemia HL60 cells oxidative burst. In dendritic cells, Tempol inhibited LPS-induced production of TNF-α, IL-6, and IL-12p70, downregulated expression of co-stimulatory molecules, and prevented antigen-dependent lymphocyte proliferation. Notably, Tempol (20 mg/kg) partially increased the survival of mice with CM. Mechanistically, treated mice had lowered plasma levels of MCP-1, suggesting that Tempol downmodulates EC function and vascular inflammation. Tempol also diminished blood brain barrier permeability associated with CM when started at day 4 post infection but not at day 1, suggesting that ROS production is tightly regulated. Other antioxidants—such as α-phenyl N-tertiary-butyl nitrone (PBN; a spin trap), MnTe-2-PyP and MnTBAP (Mn-phorphyrin), Mitoquinone (MitoQ) and Mitotempo (mitochondrial antioxidants), M30 (an iron chelator), and epigallocatechin gallate (EGCG; polyphenol from green tea) did not improve survival. By contrast, these compounds (except PBN) inhibited Plasmodium falciparum growth in culture with different IC50s. Knockout mice for SOD1 or phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (gp91phox–/–) or mice treated with inhibitors of SOD (diethyldithiocarbamate) or NADPH oxidase (diphenyleneiodonium) did not show protection or exacerbation for CM.ConclusionResults with Tempol suggest that intracellular ROS contribute, in part, to CM pathogenesis. Therapeutic targeting of intracellular ROS in CM is discussed.

Shikonin targets cytosolic thioredoxin reductase to induce ROS-mediated apoptosis in human promyelocytic leukemia HL-60 cells

Shikonin, a major active component of the Chinese herbal plant Lithospermum erythrorhizon, has been applied for centuries in traditional Chinese medicine. Although shikonin demonstrates potent anticancer efficacy in numerous types of human cancer cells, the cellular targets of shikonin have not been fully defined. We report here that shikonin may interact with the cytosolic thioredoxin reductase (TrxR1), an important selenocysteine (Sec)-containing antioxidant enzyme with a C-terminal -Gly-Cys-Sec-Gly active site, to induce reactive oxygen species (ROS)-mediated apoptosis in human promyelocytic leukemia HL-60 cells. Shikonin primarily targets the Sec residue in TrxR1 to inhibit its physiological function, but further shifts the enzyme to an NADPH oxidase to generate superoxide anions, which leads to accumulation of ROS and collapse of the intracellular redox balance. Importantly, overexpression of functional TrxR1 attenuates the cytotoxicity of shikonin, whereas knockdown of TrxR1 sensitizes cells to shikonin treatment. Targeting TrxR1 with shikonin thus discloses a previously unrecognized mechanism underlying the biological activity of shikonin and provides an in-depth insight into the action of shikonin in the treatment of cancer.

Ascorbic Acid Synergistically Potentiates Phloxine B‐induced Photocytotoxicity in Human Acute Promyelocytic Leukemia Cells

Ascorbic acid (AsA) is known as an antioxidant but concomitantly possesses a pro-oxidant property. Because the impact of AsA on photodynamic therapy response is unclear, we investigated the effect of AsA on photocytotoxicity induced by phloxine B in human acute promyelocytic leukemia HL-60 cells. AsA synergistically enhanced phloxine B-induced photocytotoxic effects, including inhibition of cell proliferation, DNA ladder formation, and caspase-3 activation, whereas AsA itself showed no photocytotoxicity. AsA also enhanced the consumption of the reduced glutathione level compared with the cells treated with phloxine B alone under the light condition. Combination of AsA with phloxine B under the light condition enhanced the phosphorylation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase (MAPK). These effects were completely cancelled by catalase. These results suggest that AsA synergistically enhances phloxine B-induced photocytotoxicity, possibly through the extracellular oxidative stress-dependent MAPK pathway activation.

Study on human promyelocytic leukemia HL-60 cells apoptosis induced by fucosterol

In this study, we investigated the effect of fucosterol on HL-60 and the molecular mechanism. HL-60 Cells were treated with fucosterol, and 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) method was used to study fucosterol anti-tumor activity. Morphology of HL-60 cells was observed. Flow cytometry (FCM) was employed to detect the cell cycle. Laser scanning confocal microscope (LSCM) was used to analyze mitochondrial membrane potential (MMP) and the expressions of Fas, FasL, Fadd and Caspase-8. Western blot was performed to analyze the expressions of Cyt-C, Pro-Caspase-9 and Pro-Caspase-3. Caspase activity kits were used to determine the activity of Caspase-9, Caspase-8 and Caspase-3. The results showed fucosterol could inhibit the growth of HL-60 cells, and the cell cycle was arrested at G2/M phase. HL-60 cells showed obvious apoptosis morphology. After being treated with fucosterol for 24 h, HL-60 cells decreased MMP, induced Cyt-C release and Caspase-9, Caspase-3 activation. Fucosterol also increased the protein expression of Fas, FasL, Fadd and Caspase-8. Moreover, the activity of Caspase-9, Caspase-8 and Caspase-3 was increased significantly. In conclusion, Fucosterol can induce HL-60 cells apoptosis, suggesting that it may be a potent agent for cancer prevention and treatment.

Antioxidant activities of 5-hydroxyoxindole and its 3-hydroxy-3-phenacyl derivatives: The suppression of lipid peroxidation and intracellular oxidative stress

The antioxidant activities of 5-hydroxyoxindole (1) and newly synthesized 3,5-dihydroxy-3-phenacyl-2-oxindole derivatives against rat liver microsome/tert-butylhydroperoxide system-induced lipid peroxidation and hydrogen peroxide-induced intracellular oxidative stress were investigated. Compound 1 and its derivatives showed significant suppression of lipid peroxidation and an intracellular oxidative stress. The effects of the more lipophilic derivatives tended to be greater than that of the original compound 1. The cytotoxicity of all of the oxindole derivatives on human promyelocytic leukemia HL60 cells was lower than that of 2,6-di(tert-butyl)-4-hydroxytoluene (BHT), a widely used phenolic antioxidant. These results show that compound 1 and its 3-substituted derivatives could be good lead candidates for future novel antioxidant therapeutics.

Aza-cycloisodityrosine analogue of RA-VII, an antitumor bicyclic hexapeptide

An aza-cycloisodityrosine analogue of RA-VII, 3, was designed and synthesized. The key aza-cycloisodityrosine unit was prepared by copper(II)-acetate-mediated intramolecular phenylamine/arylboronic acid coupling of dipeptide followed by connection with the tetrapeptide segment to afford a hexapeptide. Subsequent macrocyclization of the hexapeptide with EDC·HCl and HOOBt under dilute conditions gave 3. Analogue 3 showed significant cytotoxic activity against human promyelocytic leukemia HL-60 cells and human colon carcinoma HCT-116 cells, but its activity was weaker than that of parent peptide RA-VII (1).

Bioreductive activation of mitoxantrone by NADPH cytochrome P450 reductase does not change its apoptotic stimuli properties in regard to sensitive and multidrug resistant leukaemia HL60 cells

The objective of this study was to examine the effect of bioreductive activation of antitumour drug, mitoxantrone (MX), by liver NADPH cytochrome P450 reductase (CPR) on inducing apoptosis of human promyelocytic sensitive leukaemia HL60 cell line and its multidrug resistance (MDR) sublines exhibiting two different phenotypes of MDR related to the overexpression of P-glycoprotein (HL60/VINC) or MRP1 (HL60/DOX). It was found that non-activated as well as CPR-activated form of MX used at IC90 were able to influence cell cycle of sensitive HL60 as well as resistant cells and induce apoptosis. Interestingly, it was evidenced that HL60/VINC cells were more susceptible to undergo caspase-3/caspase-8-dependent apoptosis induced by both studied forms of MX compared to HL60 and HL60/DOX cells. However, the examined agent did not change the expression of Fas receptors on the surface of HL60 sensitive as well as resistant cells regardless of its form used in the study. Obtained results suggest that CPR-dependent reductive activation of MX does not change its apoptotic stimuli properties in regard to sensitive HL60 and multidrug resistant (HL60/VINC and HL60/DOX) leukaemia cells. Nevertheless, taking into account that side toxic effects observed in course of patient treatment with antitumour drugs are dose-dependent, it seems that the reported increase in antiproliferative activity and ability to induce apoptosis of MX after its reductive activation by exogenous CPR against the MDR cells overexpressing both P-glycoprotein and MRP1 at much more lower concentrations of this drug could be of clinical importance for the treatment of tumours resistant to classical chemotherapy.

Hexane soluble extract of Mallotus philippensis(Lam.) Muell. Arg. root possesses anti-leukaemic activity

BackgroundMallotus philippensis (Lam.) Muell. Arg. is a well known medicinal plant of Asia and Australia. Various compounds from different aerial parts of the plant have been reported possessing potent pharmacological, antiviral, antibacterial and cytotoxic activities. We were interested to determine the effects of some root extracts from M. philippensis on human promyelocytic leukemia HL-60 cell proliferation, cell cycle regulators and apoptosis in order to investigate its anti-leukemic potential.ResultsRoot extract of M. philippensis was initially extracted in organic solvents, hexane, ethyl acetate, and n-butanol. The hexane extract showed highest toxicity against p53-deficient HL-60 cells (IC50 1.5 mg dry roots equivalent/ml medium) after 72 h and interestingly, inhibition of cell proliferation was preceded by the upregulation of the proto-oncogenes Cdc25A and cyclin D1 within 24 h. The hexane extract induced 18% apoptosis after 48 h of treatment. Chemical composition of the hexane extract was analyzed by GC-MS and the 90% fragments were matched with polyphenolic compounds.ConclusionsThe present study confirms that the hexane fraction of M. philippensis root extract possesses anti-leukemic activity in HL-60 cells. The polyphenols were the main compounds of the hexane extract that inhibited proliferation and induced apoptosis.

Induction of mitochondrial dependent apoptosis and cell cycle arrest in human promyelocytic leukemia HL-60 cells by an extract from Dorstenia psilurus: a spice from Cameroon

BackgroundThe use of edible plants is an integral part of dietary behavior in the West region of Cameroon. Dorstenia psilurus (Moraceae) is widely used as spice and as medicinal plant for the treatment of several diseases in Cameroon. The aim of this study is to investigate the cytotoxic and apoptotic potential of methanol extract of D. psilurus in human promyelocytic leukemia (HL-60) cells and prostate cancer (PC-3) cells.MethodsCytotoxicity of D. psilurus extract was tested in HL-60 and PC-3 cells using 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay and flow cytometric methodsResultsThe methanol extract of D. psilurus have significant in vitro cytotoxic activity in HL-60 cells and PC-3 cells with IC50 value of 12 ±1.54 μg/ml and 18 ± 0.45 μg/ml respectively after 48 h. The mechanism of antiproliferative activity showed that after 24 h, D. psilurus extract induces apoptosis on HL-60 cells by the generation of reactive oxygen species (ROS) along with concurrent loss of mitochondrial membrane potential, modification in the DNA distribution and enhance of G2/M phase cell cycle.ConclusionThe extract induces apoptosis of HL-60 cells associated with ROS production, loss of mitochondrial membrane potential and apoptotic DNA fragmentation.

Cheliensisin A Inhibits EGF-Induced Cell Transformation with Stabilization of p53 Protein Via a Hydrogen Peroxide/Chk1-Dependent Axis

Cheliensisin A (Chel A), a novel styryl-lactone isolated from Goniothalamus cheliensis Hu, has been shown to induce apoptosis in human promyelocytic leukemia HL-60 cells with Bcl-2 downregulation. Yet, the potential chemopreventive effect of Chel A has not been explored. Here, we showed that Chel A treatment with various concentrations (0.5, 1.0, 2.0, and 4.0 μmol/L) for 3 weeks could dramatically inhibit EGF-induced cell transformation in Cl41 cells (IC50 ∼2.0 μmol/L). Also, coincubation of Cl41 cells with Chel A (2.0 and 4.0 μmol/L) for 48 hours could induce cell apoptosis in a caspase-3-dependent manner. Mechanically, Chel A treatment could result in increased p53 phosphorylation at Ser15 and elevated p53 total protein expression. Moreover, we found that p53 induction by Chel A was regulated at the protein degradation level, but not at either the transcription or the mRNA level. Further studies showed that p53 stabilization by Chel A was mediated via induction of phosphorylation and activation of Chk1 protein at Ser345. This notion was substantiated by the results that transfection of dominant negative mutant of Chk1 (GFP-Chk1 D130A) significantly attenuated the p53 protein expression, cell apoptosis, and inhibition of cell transformation by Chel A. Finally, increased hydrogen peroxide was found to mediate Chk1 phosphorylation at Ser345, p53 protein induction, cell apoptotic induction, and transformation inhibition following Chel A treatment. Taken together, our studies identify Chel A as a chemopreventive agent with the understanding of the molecular mechanisms involved.