Human Leukemia Cell Line K562 Research Articles

Three new cyclopentenone derivatives from Actinoalloteichus nanshanensis NEAU 119

Three new cyclopentenone derivatives (1–3) were isolated from the rare actinomycete Actinoalloteichus nanshanensis NEAU 119. Their structures were elucidated by extensive spectroscopic analysis. Compound 1 showed moderate cytotoxic activity against human lung adenocarcinoma cell line A549, human leukemia cell line K562, and human renal carcinoma cell line ACHN with an IC50 of 14.67, 11.87, and 23.36 μg ml− 1, respectively.

Eupolyphaga sinensis Walker inhibits human chronic myeloid leukemia cell K562 growth by inducing G2–M phase cell cycle arrest and targeting EGFR signaling pathway and in S180 tumor-bearing mice

Eupolyphaga sinensis Walker is not only used as a food to enhance immunity, but is used as a traditional Chinese medicine and is known as the “preferred drug to regulate blood flow”. Previous studies have reported its potential biological activities including anticoagulation, antithrombotic, liver protective effect and antitumor effects. Our results indicated that E. sinensis Walker 70% ethanol extract exhibited anti-tumor effects on S180 (murine sarcoma cell line) cells implanted mice. It effectively inhibited K562 (human chronic myeloid leukemia cell line) cells proliferation and induced G2–M phase arrest accompanying through up-regulation of cyclin B1, cdc2 and down-regulation of cyclin D1, cyclin E1, cdc25c and p53. In addition, it inhibited EGF secretion and EGFR kinase activity. Western blotting analysis indicated that it also inhibited the phosphorylation EGFR and activation of its downstream signaling molecules AKT and ERK. These results suggested that the antitumor mechanism of E. sinensis Walker involved altering the cell cycle and inhibiting EGFR phosphorylation in the EGFR signaling pathway.

Resveratrol inhibits the phosphatidylinositide 3-kinase/protein kinase B/mammalian target of rapamycin signaling pathway in the human chronic myeloid leukemia K562 cell line.

Resveratrol inhibits the initiation, promotion and progression of tumors, however, the mechanism by which resveratrol inhibits the proliferation of the human chronic myeloid leukemia K562 cell line remains unclear. The present study was conducted to investigate the effect of resveratrol on the activation of the phosphatidylinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling cascade in K562 cells. Resveratrol showed significant cytotoxic effects and induced apoptosis in K562 cells in a dose- and time-dependent manner. In addition, resveratrol attenuated the phosphorylation of PI3K, Akt and mTOR in the K562 cells. Furthermore, the selected inhibitors of PI3K (LY294002), Akt (SH-6) and mTOR (rapamycin) enhanced the effects of resveratrol in K562 cells. In addition, cyclin D1 levels were found to decrease and the activation of caspase-3 was observed. Resveratrol was also found to significantly attenuate the phosphorylation of the downstream molecules, p70S6K and 4EBP1. These results suggested that the downregulation of the PI3K/Akt/mTOR signaling cascades may be a crucial mediator in the inhibition of proliferation and induction of apoptosis by resveratrol in K562 cells.

Evaluation and Structure-Activity Relationship Analysis of a New Series of Arylnaphthalene lignans as Potential Anti-Tumor Agents

Arylnaphthalene lignan lactones have attracted considerable interest because of their anti-tumor and anti-hyperlipidimic activities. However, to our knowledge, few studies have explored the effects of these compounds on human leukemia cell lines. In this study, five arylnaphthalene lignans including 6′-hydroxy justicidin A (HJA), 6′-hydroxy justicidin B (HJB), justicidin B (JB), chinensinaphthol methyl ether (CME) and Taiwanin E methyl ether (TEME) were isolated from Justicia procumbens and their effects on the proliferation and apoptosis of the human leukemia K562 cell line were investigated then used to assess structure-activity relationships. To achieve these aims, cytotoxicity was assayed using the MTT assay, while intracellular SOD activity was detected using the SOD Activity Assay kit. Apoptosis was measured by both the using a cycle TEST PLUS DNA reagent kit as well as the FITC Annexin V apoptosis detection kit in combination with flow cytometry. Activation of caspase-mediated apoptosis was evaluated using a FITC active Caspase-3 apoptosis kit and flow cytometry. The results indicated that HJB, HJA and JB significantly inhibited the growth of K562 cells by decreasing both proliferation and SOD activity and inducing apoptosis. The sequence of anti-proliferative activity induced by the five tested arylnaphthalenes by decreasing strength was HJB > HJA > JB > CME > TEME. HJB, HJA and JB also decreased SOD activity and induced apoptosis in a dose-dependent manner. Activation of caspase-3 further indicated that HJB, HJA and JB induced caspase-dependent intrinsic and/or extrinsic apoptosis pathways. Together, these assays suggest that arylnaphthalene lignans derived from Justicia procumbens induce apoptosis to varying degrees, through a caspase-dependent pathway in human leukemia K562 cells. Furthermore, analysis of structure-activity relationships suggest that hydroxyl substitution at C-1 and C-6′ significantly increased the antiproliferative activity of arylnaphthalene lignans while a methoxyl at C-1 significantly decreased the effect.

Hypoxia-inducible factor 1α regulates the expression of CD18 and ninjurin-1

Objective To investigate the effects of hypoxia-inducible factor 1α (HIF-1α) small interfere RNA construct pSUPERH1-siHIF1α on the expression of CD18 and ninjurin 1 by K562 (human chronic myelogenous leukemia cell line) cells cultured with serums from patients with early stage of diabetic retinopathy.Methods K562 cells were cultured in 4 groups as control group (group A),diabetic group (group B),diabetes and pSUPERH1-siHIF1α transfect group (group C) and diabetes and pSUPER-retro transfect group (group D).The cells in group A were cultured in human serum from age-matched healthy control,and in group B,C and D,the cells were cultured in serum from the subjects of early stage of diabetic retinopathy.Twenty-four hours before the cells were cultured by the serum from the subjects of early stage of diabetic retinopathy,the HIF-1α specific siRNA expression vector pSUPERH1-siHIF1α and empty vector pSUPER-retro were transfected into the cells of group C and D,respectively.The percentages of CD18 and ninjurin-1 positive cell on the surface of K562 cells were measured by Flow Cytometry.The adherent rate between K562 and RF/6A was measured by the rose Bengal staining test.Results The percentages of CD18 positive cell in the group A,B,C and D were significantly different (F=14.33,P=0.01).The percentage of group B was significantly higher than that in group A (P=0.001) ; the percentage of group C was significantly lower than that in group B (P=0.001) and group D (P=0.02) ; the difference between group C and A was not significant (95％CI=-14.89-2.13,P=0.12).The differences of the percentage of ninjurin-1 positive cell among the group A,B,C and D were significant (F=39.38,P=0.001).The percentage of group B was significantly higher than that in group A (P=0.00); the difference of the percentage between group C and B was not significant (P =0.06),that was also not significant between group C and D (P=0.49).The differences of the adherent rate between K562 and RF/6A (rhesus monkey retinal choroid blood vessel endothelial cell line) among the group A,B,C and D were significant (F=20.62,P=0.00).The adherent rate of group B was significantly higher than that in group A (P=0.00),the adherent rate in group C was significantly lower than that in group B (P=0.01),but it was still significantly higher than that in group A (P=0.002),the difference of adherent rate between group B and D was not significant (P=0.68).Conclusion Under the early stage of diabetic retinopathy,HIF-1α small interfere RNA pSUPERH1-siHIF1α may significantly suppress the expression of CD18 on the surface of K562 cells,but it may not significantly influence the expression of ninjurin-1 on the surface of K562 cells. Key words: Diabetic retinopathy/pathophsiology; Hypoxia-inducible factor 1, alpha subunit; RNA, small interfering

Resveratrol induces human K562 cell apoptosis, erythroid differentiation, and autophagy

Resveratrol (Res) is a naturally occurring phytoalexin with apoptotic and inducing-glob effects in leukemic cells, but the potential induction of erythroid differentiation in cells is not fully understood. Here, we investigated the effects of Res on human erythro-megakaryoblastic leukemia cell line K562. Among the treated cells, proliferation was inhibited and the occurrence of cell apoptosis and cell death were detected. Erythroid differentiation assay was explored, and we found that Res could increase the expression of glycophorin A (GPA), HBA1, HBB, and γ-globin genes and enforced the expression of GPA, CD71, and Band3 proteins. Res also induced K562 cell autophagy when the concentration of Res was increased up to 50 or 100 μM. Our findings suggested that Res possesses the potency not only inducing apoptosis but also inducing erythroid differentiation and autophagy in K562 cells. These results provide that Res may be a therapeutic candidate for chronic myelogenous leukemia treatment.

Discovery and identification of PIM-1 kinase inhibitors through a hybrid screening approach

PIM-1 kinase is an important therapeutic target in the treatment of cancer. Discovery and identification of PIM-1 Inhibitors with novel scaffolds are an effective way for developing potent therapeutic agents for the treatment of cancers. Here we proposed a hybrid screening approach which combines an optimal structure-based drug design strategy and a simple pharmacophore model to discover PIM-1 kinase inhibitors. With the proposed hybrid screening approach, the SPECS database containing 204,580 molecules was screened. In total, 89 hits were obtained. Forty three of them were purchased and tested in bioassays. Finally, 5 lead compounds with novel scaffolds were identified to exhibit promising antitumor activities against human leukemia cell line MV4-11, K-562 and human prostate cancer cell line PC-3 and DU145. Their IC(50) values range from 4.40 to 37.96 μM. Three hits with 3 different scaffolds were selected from these five hits for binding mode analysis. It was demonstrated that the subtle differences in the interactions of the representatives with PIM-1 kinase contribute to the different inhibitory activities. It was also demonstrated that the suggested hybrid screening approach is an effective method to discover PIM-1 inhibitors possessing different scaffolds. These leads have a strong likelihood to act as further starting points for us in the optimization and development of potent PIM-1 inhibitors.

TNFα-induced apoptosis in human myeloid cell lines HL-60 and K562 is dependent of intracellular ROS generation

The present study determines the role of reactive oxygen species (ROS) production and calcium signaling evoked by the tumor necrosis factor-alpha (TNFα) on apoptosis in the human leukemia HL-60 and K562 cell lines. The results show that treatment of both cell lines cells with 10 ng/mL TNFα resulted in a rise in the percentage of apoptotic cells after 6 h of treatment. It was also observed that the administration of 10 ng/mL TNFα increased intracellular ROS production, as well as a time-dependent increase in caspase-8, -3, and -9 activities. The present results also show that the pretreatment with well-known antioxidants such as trolox and N-acetyl cysteine partially reduced the caspase activation caused by the administration of TNFα. The findings suggest that TNFα-induced apoptosis is dependent on alterations in intracellular ROS generation in human leukemia HL-60 and K562 cells.

Structure and biological activities of a pectic polysaccharide from Mosla chinensis Maxim. cv. Jiangxiangru

A water-soluble pectic polysaccharide (MP-A40) was isolated and purified from Mosla chinensis Maxim. cv. Jiangxiangru for the first time, with a molecular weight of 32,600Da. MP-A40 was comprised of 68.63% galacturonic acid and 13.05% neutral sugar. In addition, arabinose, galactose, rhamnose, mannose and glucose composed the neutral sugar in a relative ratio of 4.94, 3.07, 2.13, 1.62 and 1.29% of the dry weight of MP-A40, respectively. Structural characterization of MP-A40 was investigated by methylation analysis and 1D/2D NMR spectroscopy. From the results, the structure of MP-A40 was revealed as follows: 1,4-linked α-d-GalpA and 1,4-linked α-d-GalpA6Me interspersed with rare t-Araf (0.60%), t-Rhap (1.67%) and t-GalpA (10.15%). Esterification assay showed that about 32% of the carboxylic groups in GalA residues existed as methyl ester. In addition, MP-A40 could inhibit the growth of human leukemic cell line K562 and stimulate nitric oxide production from RAW 264.7 macrophages both in dose-dependent manners.

In vitro anti-telomerase activity of novel lycopene-loaded nanospheres in the human leukemia cell line K562.

Background:Lycopene, a plant carotenoid, has potent effects against the various types of cancer cells. To date, the effect of lycopene in the free and encapsulated forms on the telomerase activity in human leukemia cell line K562 have not been investigated. The aim of the present study was to prepare a novel lycopene-loaded nanosphere and compare its anti-telomearse activity in K562 cell line with those of free lycopene.Materials and Methods:The lycopene-loaded nanospheres were prepared by nanoprecipitation method. The lycopene entrapment efficacy was measured by high-performance liquid chromatography (HPLC) method. The anti-proliferation effect of the lycopene in the free and encapsulated forms in the different times (0-72 h) and the different doses (0-100 μg/ml) on K562 cell line was studied using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The changes of telomerase activity, following treatment with the lycopene in the free and encapsulated forms, were detected using the telomeric repeat amplification protocol-enzyme-linked immunosorbent assay.Results:The entrapment efficacy of lycopene was 78.5% ± 2. Treatment of the K562 cell line with lycopene, in particular in encapsulated form, resulted in a significant inhibition of the cell growth and increasing of percentage of apoptotic cells. It has also been observed that the telomerase activity in the lycopene-loaded nanospheres-treated cells was significantly inhibited in a dose and time-dependent manner.Conclusion:Our data suggest a novel mechanism in the anti-cancer activity of the lycopene, in particular in encapsulated form, and could be provided a basis for the future development of anti-telomerase therapies.

Studies on the effects of cucurbit (n = 7) uril on the physicohemical properties and anticancer activity of 10-hydroxycamptothecin

The changes in the physicochemical properties and the anticancer activity of 10-Hydroxycamptothecin (HCPT) in the presence of Cucurbit (n = 7) uril(Q[7]) were studied by using UV absorption spectroscopy, fluorescence spectroscopy and phase solubility method. The results revealed that in the acid solution (pH 2), the absorption band of the guest HCPT exhibited a progressively lower absorbance at λ265 nm, while a progressively higher absorbance at λ425 nm as the ratio of N(Q[7])/N(HCPT) was increased. A sharp isosbestic point at λ387 nm was consistent with a simple interaction between Q[7] and HCPT. The emission spectra of the guest HCPT also exhibited a progressive decrease in fluorescence intensity at λ558 nm with a violet shift when the ratio of N(Q[7])/N(HCPT) was increased. The measured data from both absorption spectrophotometric and fluorescence spectroscopy analysis fitted to a 2 : 1 host:guest complexation, yielded a calculated stable constants (β) of 1.549 x 10(13) L2 x mol(-2) and 0.907 x 10(13) L2 x mol(-2) respectively through Reactlab EQULLIBRIA software. The effect of Q[7] on the solubility of HCPT was investigated by using phase solubility method. Upon the addition of Q[7] concentration, the solubility of HCPT was enhanced step by step at the value of pH 2 and an about 250 times advance was attained when the concentration of Q[7] was 1 x 10(-3) mol x L(-1), while the Q[7]-HCPT inclusion complex made by total evaporation method can increase about 1 170 times compared to the pure HCPT. At the end, the mixtures of Q[7] and HCPT were tested in terms of cytotoxicity on the human lung cancer cell line A549 and human leukemia cell line K562 to compare their reactivity with the free HCPT and the comparative cytotoxic activity was got for A549 and K562.

Synthesis and anticancer evaluation of benzyloxyurea derivatives.

A series of novel benzyloxyurea derivatives was designed, synthesized by substituting different benzyls or phenyls on N,N'-positions of the hydroxyurea (HU). These target compounds were evaluated for their anticancer activity in vitro against human leukemia cell line K562 and murine leukemia cell line L1210 in comparison with HU by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Some of the compounds showed promising anticancer activity against the cells. Molecular docking experiments with Saccharomyces cerevisiae R1 domain indicated that 4a and 4f' have stronger affinity than 4m and 4n. Flow cytometry study showed that compound 4g exerted greater apoptotic activity against K562 cells line than HU.

Antioxidant enzyme inhibitor role of phosphine metal complexes in lung and leukemia cell lines.

Phosphine metal complexes have been recently evaluated in the field of cancer therapy. In this research, the cytotoxic effects of some metal phosphines {[PdCl2((CH2OH)2PCH2)2NCH3] (C1), [RuCl2(((CH2OH)2PCH2)2NCH3)2] (C2), [PtCl2((Ph2PCH2)2NCH3)(timin)2] (C3)} on K562 (human myelogenous leukemia cell line) and A549 (adenocarcinomic human alveolar basal epithelial cells) cells were investigated using the MTT test. C1 and C2 are water-soluble metal complexes, which may have some advantages in in vitro and in vivo studies. The effects of the above-mentioned metal complexes on thioredoxin reductase (TrxR) (EC: 1.8.1.9), glutathione peroxidase (GPx) (EC: 1.11.1.9), and catalase (Cat) (EC: 1.11.1.6) enzymes were also tested. The results of this research showed that all three metal complexes indicated dose-dependent cytotoxicity on A549 and K562 cell lines and that the complexes inhibited different percentages of the TrxR, GPx, and Cat enzymes of these tumor cells.

Human interleukin‐23 receptor antagonists derived from an albumin‐binding domain scaffold inhibit IL‐23‐dependent ex vivo expansion of IL‐17‐producing T‐cells

Engineered combinatorial libraries derived from small protein scaffolds represent a powerful tool for generating novel binders with high affinity, required specificity and designed inhibitory function. This work was aimed to generate a collection of recombinant binders of human interleukin-23 receptor (IL-23R), which is a key element of proinflammatory IL-23-mediated signaling. A library of variants derived from the three-helix bundle scaffold of the albumin-binding domain (ABD) of streptococcal protein G and ribosome display were used to select for high-affinity binders of recombinant extracellular IL-23R. A collection of 34 IL-23R-binding proteins (called REX binders), corresponding to 18 different sequence variants, was used to identify a group of ligands that inhibited binding of the recombinant p19 subunit of IL-23, or the biologically active human IL-23 cytokine, to the recombinant IL-23R or soluble IL-23R-IgG chimera. The strongest competitors for IL-23R binding in ELISA were confirmed to recognize human IL-23R-IgG in surface plasmon resonance experiments, estimating the binding affinity in the sub- to nanomolar range. We further demonstrated that several REX variants bind to human leukemic cell lines K-562, THP-1 and Jurkat, and this binding correlated with IL-23R cell-surface expression. The REX125, REX009 and REX128 variants competed with the p19 protein for binding to THP-1 cells. Moreover, the presence of REX125, REX009 and REX115 variants significantly inhibited the IL-23-driven expansion of IL-17-producing primary human CD4+ T-cells. Thus, we conclude that unique IL-23R antagonists derived from the ABD scaffold were generated that might be useful in designing novel anti-inflammatory biologicals. Proteins 2014; 82:975–989.

HO-1 Regulated Autophagy and Apoptosis In CML Cells Through Mitochondrial-beclin1 Pathway

AimTo explore the mechanisms underlying the HO-1-regulated apoptosis and autophagy in human chronic myeloid leukemia (CML) K562 cell line in vitro. MethodsK562 cells were devided into three groupsFK562(untreated),K562-H(infected by lenti-GFP-HO-1) and K562-siHO-1(infected by lenti-GFP-siHO-1). The protein expression of HO-1 were determined by Western blot, and its mRNA levels were obtained by real-time PCR. Cell viability was assessed by MTT assay. Morphological changes of apoptosis and autophagy were observed by the application of transmission electron microscope. Annexin V-FITC/PI dual staining assays were used to measure apoptosis. Autophagy was analyzed by Western blot and immunofluorescence staining with an anti-LC3 antibody. DCFH-DA was used as the cell-permeate indicator for intracellular ROS measurement. Protein levels of active caspase 3, Beclin1,cyto-C and Bcl2 were determined by Western blot. ResultsThe Expression of HO-1 on mRNA and protein level increased in K562-H group while reduced in K562-siHO-1, the differences showed statistically significance compared to K562-untreated group(p<0.05). Imatinib (0.20-1.60μmol/L) exerted inhibitory effects on cell proliferation in a concentration-dependent manner with IC(50) value of 1.0μmol/L, 0.7μmol/L and 0.4μmol/L, while viability of 63.41±1.46%, 41.24±2.01%, 29.49±1.87% in K562-H, K562 and K562-siHO-1(p<0.05), respectively. Exposure to imatinib (0.40 μmol/L) simultaneously induced mitochondrial-mediated apoptosis and Beclin1-dependent autophagy in all groups. Autophagy flux was found in K562-siHO-1 group while autophagosome formation decreased in K562-H group, which was significantly different from K562 group. The expressions of LC3 II and cyto-C were higher in K562-siHO-1 group than other two groups (p<0.05). Both apoptosis and autophagy caused cell death in a time-dependent manner. However, apoptosis played a more effective role in cell death. After exposure to imatinib (0.40 μmol/L), both intracellular ROS generation and the protein expression of Beclin1 and caspase9 were increased, while Bcl2 protein level were reduced in a time-dependent manner. The combination of NAC (2.50 mmol/L) with imatinib (0.40 μmol/L) contributed to the blockade of ROS generation which abrogated apoptosis, autophagy and Beclin1 expression. 3-MA (3.0mmol/L) and HO-1 silencing resulting in the inhibition of autophagy enhanced the effect of apoptosis which was induced by imatinib (respectively 70.65±2.71% and 69.27±3.13%), while this phenomenon was accompanied by increased ROS generation (1.3 and 1.6 folds higher than K562 group). ConclusionsImatinib and HO-1 siRNA simultaneously induced apoptosis and autophagy in K562 cells, which was associated with the increasing of intracellular ROS generation, cyto-C and Beclin1 protein expression. Beclin1, an autophagy related protein induced by HO-1 silencing, could further induce apoptosis in K562 cells through the inhibition of Bcl2. The responsive cytoprotection to chemotherapeutics caused by autophagy decreased while sustaining up-regulation of HO-1 in K562 cells. However, the underlying mechanisms remained to be well elucidated. Disclosures:No relevant conflicts of interest to declare.

Exosomes Derived from Hypoxic Leukemia Cells Enhance Tube Formation in Endothelial Cells

Hypoxia plays an important role during the evolution of cancer cells and their microenvironment. Emerging evidence suggests communication between cancer cells and their microenvironment occurs via exosomes. This study aimed to clarify whether hypoxia affects angiogenic function through exosomes secreted from leukemia cells. We used the human leukemia cell line K562 for exosome-generating cells and human umbilical vein endothelial cells (HUVECs) for exosome target cells. Exosomes derived from K562 cells cultured under normoxic (20%) or hypoxic (1%) conditions for 24 h were isolated and quantitated by nanoparticle tracking analysis. These exosomes were then cocultured with HUVECs to evaluate angiogenic activity. The exosomes secreted from K562 cells in hypoxic conditions significantly enhanced tube formation by HUVECs compared with exosomes produced in normoxic conditions. Using a TaqMan low-density miRNA array, we found a subset of miRNAs, including miR-210, were significantly increased in exosomes secreted from hypoxic K562 cells. We demonstrated that cancer cells and their exosomes have altered miRNA profiles under hypoxic conditions. Although exosomes contain various molecular constituents such as proteins and mRNAs, altered exosomal compartments under hypoxic conditions, including miR-210, affected the behavior of endothelial cells. Our results suggest that exosomal miRNA derived from cancer cells under hypoxic conditions may partly affect angiogenic activity in endothelial cells.

Genetically engineered fixed K562 cells: potent “off-the-shelf” antigen-presenting cells for generating virus-specific T cells

Background aimsThe human leukemia cell line K562 represents an attractive platform for creating artificial antigen-presenting cells (aAPC). It is readily expandable, does not express human leukocyte antigen (HLA) class I and II and can be stably transduced with various genes. MethodsIn order to generate cytomegalovirus (CMV) antigen-specific T cells for adoptive immunotherapy, we transduced K562 with HLA-A∗0201 in combination with co-stimulatory molecules. ResultsIn preliminary experiments, irradiated K562 expressing HLA-A∗0201 and 4-1BBL pulsed with CMV pp65 and IE-1 peptide libraries failed to elicit antigen-specific CD8+ T cells in HLA-A∗0201+ peripheral blood mononuclear cells (PBMC) or isolated T cells. Both wild-type K562 and aAPC strongly inhibited T cell proliferation to the bacterial superantigen staphylococcal enterotoxin B (SEB) and OKT3 and in mixed lymphocyte reaction (MLR). Transwell experiments suggested that suppression was mediated by a soluble factor; however, MLR inhibition was not reversed using transforming growth factor-β blocking antibody or prostaglandin E2 inhibitors. Full abrogation of the suppressive activity of K562 on MLR, SEB and OKT3 stimulation was only achieved by brief fixation with 0.1% formaldehyde. Fixed, pp65 and IE-1 peptide-loaded aAPC induced robust expansion of CMV-specific T cells. ConclusionsFixed gene-modified K562 can serve as effective aAPC to expand CMV-specific cytotoxic T lymphocytes for therapeutic use in patients after stem cell transplantation. Our findings have implications for broader understanding of the immune evasion mechanisms used by leukemia and other tumors.

Austdiol, fulvic acid and citromycetin derivatives from an endolichenic fungus, Myxotrichum sp.

A new austdiol analog myxodiol A (1), three novel fulvic acid derivatives myxotrichin A–C (2–4), and a new citromycetin analog myxotrichin D (5), were isolated from an endolichenic fungus Myxotrichum sp. inhabiting the lichen Cetraria islandica (L.) Ach. The structures of these compounds were elucidated unequivocally on the basis of comprehensive analysis of MS and NMR data. Compounds 2 and 5 displayed very weak cytotoxicity against human leukemia cell line K562, and compounds 1 showed very weak antifungal activity against Candida albicans (sc5314).

Human leukemic cell lines synthesize hyaluronan to avoid senescence and resist chemotherapy

Hyaluronan (HA) is one of the major components of the extracellular matrix. Several solid tumors produce high levels of HA, which promotes survival and multidrug resistance (MDR). HA oligomers (oHAs) can block HA effects. However, little is known about the role of HA in hematological malignancies. The aim of this work was to determine whether HA or its oligomers can modulate the proliferation of leukemia cells as well as their effect on MDR. Receptors and signaling pathways involved were also analyzed. For this purpose, the human leukemic cell lines K562 and Kv562, which are sensitive and resistant to Vincristine (VCR), respectively, were used. We demonstrated that HA induced cell proliferation in both cell lines. On K562 cells, this effect was mediated by cluster differentiation 44 (CD44) and activation of both phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) and mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) pathways, whereas on Kv562 cells, the effect was mediated by receptor for hyaluronan-mediated motility (RHAMM) and PI3K/Akt activation. The inhibition of HA synthesis by 4-methylumbelliferone (4MU) decreased cell line proliferation and sensitized Kv562 to the effect of VCR through P-glycoprotein (Pgp) inhibition, in both cases with senescence induction. Moreover, oHAs inhibited K562 proliferation mediated by CD44 as well as Akt and ERK down-regulation. Furthermore, oHAs sensitized Kv562 cells to VCR by Pgp inhibition inducing senescence. We postulate that the synthesis of HA would promote leukemia progression mediated by the triggering of the above-mentioned proliferative signals. These findings highlight the potential use of oHAs and 4MU as coadjuvant for drug-resistant leukemia.

Efficacy of combined therapy with paclitaxel and low-level ultrasound in human chronic myelogenous leukemia cell line K562

Background: Sonochemotherapy, which applies ultrasound in cancer chemotherapy, has been proven to be a promising therapeutic modality by some previous researches.Purpose: To investigate the interaction between an antineoplastic agent – paclitaxel (PTX) and low-level ultrasound in human chronic myelogenous leukemia cell line K562, their combined efficacy, and the potential mechanisms in sonochemotherapy.Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) analysis and Guava Viacount assay were adopted to examine cell viability. Apoptosis was analyzed using Annexin V-PE/7-amino-actinomycin D staining. Changes in plasma membrane permeability were monitored by FD500-uptake and lactate dehydrogenase (LDH) release assays. Additionally, the ultrastructure changes were evaluated under scanning electron microscope (SEM).Results: When 5 ng/ml PTX was combined with sonication at Load Power (LP) = 2 W, the expected synergistic effects on cell viability loss (p < 0.05) and apoptosis enhancement could be significantly detected, the plasma membrane permeability showed the best response, and relatively serious cell damages were observed under SEM.Conclusion: The interaction between these two therapies depended on specific parameter settings, such as PTX dose and ultrasound intensity. Under synergistic conditions, ultrasound significantly potentiated the efficacies of PTX, including cytotoxicity, apoptosis and cell damage induction, which may be due to the enhanced membrane permeability and the increased intracellular PTX level.