Tumor cell growth inhibition by liposome-encapsulated aromatic polyamidines

Apart from its antiproteinase activity, the aromatic polyamidine TAPP-Br [the bromo derivative of 1,3-di-(p-amidinophenoxy)-2,2-bis-(p-amidinophenoxymethyl)propane (TAPP-H)] is able to inhibit the in vitro growth of a variety of tumor cell lines, including human melanoma, and breast and kidney carcinoma. We have now shown that TAPP-Br can efficiently be encapsulated into egg phosphatidylcholine vesicles. When incorporated into these liposomes, the inhibitory effect of TAPP-Br is significantly enhanced compared with that of the free drug. Based on these promising results, a proposal is made for the delivery of this antiproliferative agent to tumor cells by using liposomes as the vehicle.     

Multisite inhibition by phenylacetate of PC-3 cell growth

Phenylacetate (PA) is a reversible inhibitor of tumor cell growth and an inhibitor of mevalonate pyrophosphate decarboxylase (MPD). We hypothesized that MPD inhibition should lower rates of protein accumulation and accretion of cell number in all cell lines regardless of tumorigenic status or origin of the cell lines. PA treatment inhibited growth of MCF-7, NIH-3T3, Detroit 551, UT-2, NCTC-929, COS-1 and PC-3 cell lines. NCTC-929 cells lack cadherins and Cos-1 cells are deficient in PPARalpha and PPARgamma, proteins suggested to be central to the action of PA. Oxidative metabolism was not impeded by PA treatment. One-dimensional and two-dimensional FACS analysis of BrdU incorporation failed to demonstrate a redistribution of nuclei in the cell cycle or that the rate of cells entering S phase had changed. Time-lapse photo-microscopy studies reveal a process that left condensed nuclei with little or no cytoplasm. However, negative TUNEL assay results and failure to block cell loss with z-VAD-fmk suggest this type of cell death is not typical apoptosis, but cell death is responsible for the lower rates of cell and protein accumulation. Supplementation studies with mevalonate pathway intermediates during inhibition of the mevalonate pathway of cholesterol biosynthesis by lovastatin confirmed MPD as a site of PA inhibition of growth, but in the presence of lovastatin with or without farnesyl pyrophosphate plus geranylgeranyl pyrophosphate, additive inhibition by PA revealed additional site(s). The existence of site(s) in addition to MPD suggests effective PA-based agents might be developed that would not inhibit MPD.     

Thioredoxin reductase and cancer cell growth inhibition by organotellurium antioxidants

Thioredoxin (Trx) expression is increased in several human primary cancers and the Trx/Trx reductase (TrxR) system therefore provides an attractive target for cancer drug development. Novel organotellurium antioxidants, especially a primitive analog of vitamin E (compound 1d) and compounds 7, 9 and 10--all carrying highly functionalized 4-(dialkylamino)phenyltelluro groups to secure high antioxidative capacity--were found to inhibit TrxR with IC50 values in the low micromolar range. Whereas antioxidant 1d also inhibited the growth of MCF-7 human breast cancer cells in culture at a similar level (IC50 = 1.8 microM), the other TrxR inhibitors were inactive in concentrations below about 10 M.     

Synergistic inhibition of human leukemia cell growth by deoxyguanosine and 1-beta-D-arabinofuranosylcytosine

We studied the ability of 2'-deoxyguanosine (dGuo) to influence 1-beta-D-arabinofuranosylcytosine (ara-C) inhibition of soft agar cloning of the cultured human leukemia cell line K562. Ara-C alone inhibited cloning in concentrations of greater than 10 nM, with a steep drop in colony formation observed between 10 and 100 nM. dGuo and ara-C synergistically inhibited cloning; the combination of ineffective concentrations of dGuo (10-50 microM) and ara-C (less than or equal to nM) inhibited cloning by 40-70%. In K562 cells, dGuo is metabolized by both nucleoside kinase and purine nucleoside phosphorylase (PNP), resulting in augmentation of both the GTP pool (to more than 200% of control after a 3 hr incubation with 500 microM dGuo) and the dGTP pool (to more than 2700% of control after 3 hr with 500 microM dGuo). dGuo (50-500 microM) caused a decrease in the dCTP and dTTP pools and an increase in the dATP pool. Synergistic concentrations of dGuo plus 10 nM ara-C augmented the ara-CTP pool up to 800% of control after 3 hr to levels equivalent to those observed after incubation with 500 nM ara-C alone. Incorporation of 10 nM ara-CTP into DNA also increased in the presence of dGuo (up to a maximum of 300% of control), but only to a level that approximated the value observed with nM ara-C alone. The disparity between enlargement of the ara-CTP pool and augmentation of ara-C incorporation into DNA is consistent with the observation of Steinberg et al. [Cancer Res. 39, 4330 (1979)] that high concentrations of dGTP may inhibit DNA polymerase activity. Thus, synergy between dGuo and ara-C is multifactorial, possibly involving inhibition of DNA polymerase by elevated dGTP and ara-CTP pools and augmented incorporation of ara-C into DNA.     

Acute inhibition of human renal tubular cell growth by cyclosporin A

Human proximal tubular cells were isolated and grown in culture for three passages. The proliferation of these cells were inhibited by the immunosuppressive drug cyclosporin A in dose dependent manner in the range of 250 to 2000 ng/ml growth medium. The cultures with low cell density were more sensitive to cyclosporin A compared to the cultures with high density, measured by the incorporation of 3H-thymidine. The toxic effect of cyclosporin A on cells isolated from patients treated with cyclosporin A, did not differ from cells isolated from normal tissue. The calcium channel blocker, verapamil, reversed the inhibitory effect of low concentrations of cyclosporin A on cell proliferation. The electron microscopy showed that cells treated with cyclosporin A, had severe morphological alterations with rounded mitochondria and giant vesicles in the cytoplasma. The results support the hypothesis that the toxic effect of cyclosporin A may be mediated through an increased Ca++ influx into the proximal tubular cells.     

In vitro prostate cancer cell growth inhibition by Brazilian plant extracts

In the present study, 1220 plant extracts obtained from 352 plants belonging to 73 families that grow in the Amazon and Atlantic rain forests were screened for cytotoxicity against PC-3 prostate cancer cell lines. Extracts were tested in the single dose of 100 microg/mL. Activity was observed in 17 aqueous or organic extracts belonging to Annonaceae, Apocynaceae, Araceae, Capparaceae, Commelinaceae, Flacourtiaceae, Lecythidaceae, Leguminosae, Passifloraceae, Rutaceae, and Violaceae.     

Enhanced cell growth inhibition by thiacremonone in paclitaxel-treated lung cancer cells

Activation of nuclear factor kappa-B (NF-κB) is implicated in drug resistant of lung cancer cells. Our previous data showed that thiacremonone inhibited activation of NF-κB. In the present study, we investigated whether thiacremonone enhanced susceptibility of lung cancer cells to a common anti-cancer drug paclitaxel by further inhibition of NF-κB. Thus, we used the threefold lower doses of IC₅₀ values (50 μg/ml thiacremonone and 2.5 nM paclitaxel). We found that combination treatment with thiacremonone and paclitaxel was more susceptible (combination index; 0.40 in NCI-H460 cells and 0.46 in A549 cells) in cell growth inhibition of two types of lung cancer cell lines compared to a single agent treatment. Consistent with the combination effect on cancer cell growth inhibition, the combination treatment further induced apoptotic cell death and arrested the cancer cells in G2/M phase accompanied with a much lower expression of cdc2 and cyclin B1, and inhibited colony formation. Much more inactivation of NF-κB and greater expression of NF-κB target apoptosis regulated genes such as caspase-8 and PARPs were found by the combination treatment. Molecular model and pull down assay as well as MALDI-TOF analysis demonstrated that thiacremonone directly binds to p50. These data indicated that thiacremonone leads to increased apoptotic cell death in lung cancer cell lines through greater inhibition of NF-κB by the combination treatment with paclitaxel.     

Stimulation and inhibition of DNA synthesis and cell growth by paraquat in cultured cells

Paraquat, a popular herbicide, is known to be very toxic to man and animals. Using cultured mammalian cells (human embryo lung cell, R-66), the effect of paraquat has been studied. With autoradiography, stimulation of DNA synthesis was observed when the cells were exposed to 0.08 or 0.4 microM of paraquat; however, DNA synthesis was inhibited with 40 microM of paraquat. To determine cell growth rate, accumulation of mitotic cells was determined after addition of colcemid (0.2 microgram/ml) into the culture medium. The ratios of mitotic cells observed were proportional to the level of DNA synthesis. And the stimulation and inhibition of cell growth were observed even after washing off paraquat from media.     

Thioredoxin reductase and cancer cell growth inhibition by organogold(III) compounds

Thioredoxin (Trx) expression is increased in several human primary cancers associated with aggressive tumor growth and decreased patient survival, and the Trx/Trx reductase (TrxR) system therefore provides an attractive target for cancer drug development. Various gold(III) compounds with none, one, two or three carbon-gold bonds were evaluated for their capacity to inhibit TrxR and the growth of MCF-7 cancer cells in vitro. Compounds with up to two carbon-gold bonds were often potent inhibitors of TrxR with IC50 values as low as 2 nmol/l. In the presence of Trx and insulin the inhibiting capacity was much lower. However, the inhibitory concentrations of the compounds did not correlate with the ability to kill cells. Out of the organometallics tested, only compound 8 with two carbon-gold bonds was able to inhibit colony formation by MCF-7 breast cancer cells at low micromolar concentrations (IC50=1.6 micromol/l). Unfortunately, the compound did not show any anti-tumor activity against MCF-7 breast cancer and HT-29 colon cancer xenografts in scid mice.     

Adenosine uptake-dependent C6 cell growth inhibition

In C6 glioma cells, adenine nucleotides, especially AMP, and adenosine inhibited cell proliferation in time- and concentration-dependent manners. alpha,beta-methylene-ADP, an ecto-5'-nucleotidase inhibitor, suppressed the hydrolysis of AMP and reversed the inhibition of cell growth induced by AMP but not by adenosine. Adenosine deaminase eliminated both AMP- and adenosine-mediated growth inhibitions. 5'-N-ethylcarboxamidoadenosine, an adenosine receptor agonist, had little effect on the cell growth. Equilibrative nucleoside transporters, ENT-1 and ENT-2, were expressed in C6 cells by determining their mRNAs. ENT inhibitors, nitrobenzylthioinosine and dipyridamole, suppressed the uptake of [(3)H]adenosine into C6 cells, and attenuated AMP- or adenosine-mediated growth inhibition. Furthermore, an adenosine kinase inhibitor 5-iodotubercidin reversed the growth inhibition induced by AMP and adenosine. When uridine was added in the extracellular space, AMP- or adenosine-induced cell growth inhibition was completely reversed, suggesting that intracellular pyrimidine starvation would be involved in their cytostatic effects. These results indicate that extracellular adenine nucleotides inhibit C6 cell growth via adenosine, which is produced by ecto-nucleotidases including CD73 at the extracellular space and then incorporated into cells by ENT2. Intracellular AMP accumulation by adenosine kinase after adenosine uptake would induce C6 cell growth inhibition through pyrimidine starvation.     

Biomedical nanoparticles modulate specific CD4+ T cell stimulation by inhibition of antigen processing in dendritic cells

Abstract

Understanding how nanoparticles may affect immune responses is an essential prerequisite to developing novel clinical applications. To investigate nanoparticle-dependent outcomes on immune responses, dendritic cells (DCs) were treated with model biomedical poly(vinylalcohol)-coated super-paramagnetic iron oxide nanoparticles (PVA-SPIONs). PVA-SPIONs uptake by human monocyte-derived DCs (MDDCs) was analyzed by flow cytometry (FACS) and advanced imaging techniques. Viability, activation, function, and stimulatory capacity of MDDCs were assessed by FACS and an in vitro CD4⁺ T cell assay. PVA-SPION uptake was dose-dependent, decreased by lipopolysaccharide (LPS)-induced MDDC maturation at higher particle concentrations, and was inhibited by cytochalasin D pre-treatment. PVA-SPIONs did not alter surface marker expression (CD80, CD83, CD86, myeloid/plasmacytoid DC markers) or antigen-uptake, but decreased the capacity of MDDCs to process antigen, stimulate CD4⁺ T cells, and induce cytokines. The decreased antigen processing and CD4⁺ T cell stimulation capability of MDDCs following PVA-SPION treatment suggests that MDDCs may revert to a more functionally immature state following particle exposure.     

Biomedical nanoparticles modulate specific CD4+ T cell stimulation by inhibition of antigen processing in dendritic cells

Understanding how nanoparticles may affect immune responses is an essential prerequisite to developing novel clinical applications. To investigate nanoparticle-dependent outcomes on immune responses, dendritic cells (DCs) were treated with model biomedical poly(vinylalcohol)-coated super-paramagnetic iron oxide nanoparticles (PVA-SPIONs). PVA-SPIONs uptake by human monocyte-derived DCs (MDDCs) was analyzed by flow cytometry (FACS) and advanced imaging techniques. Viability, activation, function, and stimulatory capacity of MDDCs were assessed by FACS and an in vitro CD4+ T cell assay. PVA-SPION uptake was dose-dependent, decreased by lipopolysaccharide (LPS)-induced MDDC maturation at higher particle concentrations, and was inhibited by cytochalasin D pre-treatment. PVA-SPIONs did not alter surface marker expression (CD80, CD83, CD86, myeloid/plasmacytoid DC markers) or antigen-uptake, but decreased the capacity of MDDCs to process antigen, stimulate CD4+ T cells, and induce cytokines. The decreased antigen processing and CD4+ T cell stimulation capability of MDDCs following PVA-SPION treatment suggests that MDDCs may revert to a more functionally immature state following particle exposure.     

PEG-OCL micelles for quercetin solubilization and inhibition of cancer cell growth

In this study, quercetin (QCT), a flavonoid with high anticancer potential, was loaded into polymeric micelles of PEG-OCL (poly(ethylene glycol)-b-oligo(ε-caprolactone)) with naphthyl or benzyl end groups in order to increase its aqueous solubility. The cytostatic activity of the QCT-loaded micelles toward different human cancer cell lines and normal cells was investigated. The results showed that the solubility of QCT entrapped in mPEG750-b-OCL micelles was substantially increased up to 1 mg/ml, which is approximately 110 times higher than that of its solubility in water (9 μg/ml). The average particle size of QCT-loaded micelles ranged from 14 to 19 nm. The QCT loading capacity of the polymeric micelles with naphthyl groups was higher than that with benzyl groups (10% and 6%, respectively). QCT-loaded, benzyl- and naphthyl-modified micelles effectively inhibited the growth of both sensitive and resistance cancer cells (human erythromyelogenous leukemia cells (K562) and small lung carcinoma cells (GLC4)). However, the benzyl-modified micelles have a good cytocompatibility (in the concentration range investigated (up to 100 μg/ml), they are well tolerated by living cells), whereas their naphthyl counterparts showed some cytotoxicity at higher concentrations (60–100 μg/ml). Flow cytometry demonstrated that the mechanism underlying the growth inhibitory effect of QCT in its free form was inducing cell cycle arrest at the G2/M phase. Benzyl-modified micelles loaded with QCT also exhibited this cycle arresting the effect of cancer cells. In conclusion, this paper shows the enhancement of solubility and cell cycle arrest of QCT loaded into micelles composed of mPEG750-b-OCL modified with benzyl end groups. These micelles are therefore considered to be an attractive vehicle for the (targeted) delivery of QCT to tumors.     

In vitro inhibition of rat arterial smooth muscle cell growth by extractive sulfated mucopolysaccharides

The effect of a sulfated mucopolysaccharide mixture of known composition, extracted from pig duodenum, was studied on the proliferation of rat arterial smooth muscle cells cultured from rat aorta. Cell growth, stimulated by fetal calf serum, was monitored by direct cell count and by determination of the mitotic index. The extractive mixture was studied in comparison with commercial heparin, with heparin with different electrophoretic mobilities in barium acetate and with dermatan and heparan sulfates. Heparins and the extractive mucopolysaccharide mixture inhibited cell growth measured at various time intervals, and in their presence the proliferation of smooth muscle cells plateaued at lower cell densities. Dermatan and heparan sulfates were either inactive or significantly less effective than the other mucopolysaccharides. A short preincubation (3 h) of smooth muscle cells with the extractive mixture, followed by incubation with the growing medium with no mucopolysaccharides added, slowed the cell growing rate, suggesting an interaction of the mixture components with the cell surface.     

金粉蕨素拮抗氧化损伤所抑制的内皮细胞增殖的作用及其机制

目的　探讨金粉蕨素拮抗Menadione氧化损伤所抑制的内皮细胞增殖的作用及其机制。方法　以Menadione(O- 2 )损伤人脐静脉内皮细胞作为氧化损伤模型 ,采用MTT法和细胞计数法 ,观察不同浓度金粉蕨素对Menadione损伤内皮细胞生长抑制率的影响 ;利用硝酸还原酶法测定培养液中NO含量 ;以Westernblot检测细胞eNOS活性及磷酸化ERK1 / 2的表达。结果　金粉蕨素保护组与损伤组相比 ,内皮细胞的生长抑制率明显降低 ,培养液中NO含量增高 ,eNOS活性增强 ,磷酸化ERK1 / 2表达上调。结论　NO和ERK1 / 2通路可能介导了金粉蕨素拮抗Menadione氧化损伤所抑制内皮细胞增殖的保护作用     

Targeting cell death in tumors by activating caspases

Cytotoxic approaches to killing tumor cells, such as chemotherapeutic agents, gamma-irradiation, suicide genes or immunotherapy, have been shown to induce cell death through apoptosis. The intrinsic apoptotic pathway is activated following treatment with cytotoxic drugs, and these reactions ultimately lead to the activation of caspases, which promote cell death in tumor cells. In addition, activation of the extrinsic apoptotic pathway with death-inducing ligands leads to an increased sensitivity of tumor cells toward cytotoxic stimuli, illustrating the interplay between the two cell death pathways. In contrast, tumor resistance to cytotoxic stimuli may be due to defects in apoptotic signaling. As a result of their importance in killing cancer cells, a number of apoptotic molecules are implicated in cancer therapy. The knowledge gleaned from basic research into apoptotic pathways from cell biological, structural, biochemical, and biophysical approaches can be used in strategies to develop novel compounds that eradicate tumor cells. In addition to current drug targets, research into molecules that activate procaspase-3 directly may show the direct activation of the executioner caspase to be a powerful therapeutic strategy in the treatment of many cancers.     

Targeting platelet-derived endothelial cell growth factor/thymidine phosphorylase for cancer therapy

Thymidine phosphorylase (TP) is a key enzyme in the pyrimidine nucleoside salvage pathway, but it also recognizes and inactivates various anti-cancer chemotherapeutic agents. Moreover, TP is identical to platelet-derived endothelial cell growth factor (PD-ECGF), an angiogenic factor with anti-apoptotic properties. Increased expression of PD-ECGF/TP is found in many tumor and stromal cells, and elevated TP levels are associated with aggressive disease and/or poor prognosis. Thus, progression and metastasis of TP-expressing tumors might be abrogated by TP inhibitors that are used as single agents or in combination with (TP-sensitive) nucleoside analogues. On the other hand, increased TP activity in tumors may be exploited for the tumor-specific activation of fluoropyrimidine prodrugs, such as capecitabine. This review will focus on the different biological activities of PD-ECGF/TP and their implications for cancer progression and treatment.     

反义阻断hMMS2基因表达以抑制细胞生长

将新近发现的人泛素缀合酶样蛋白基因h MMS2的 c DNA特定反向克隆到本室改建的真核细胞表达载体 p MAMneo- amp-中 ,构建了表达h MMS2反义 RNA的重组质粒 .将此反义表达重组质粒 (p MAM- anti- h MMS2 )用改良的磷酸钙法转染人羊膜 FL细胞 ,建立 FL- h MMS2 -细胞系 .比较该细胞系及 FL细胞和转染了空载体 p MAMneo-amp-的 FL细胞 (FL- MAMneo细胞 )的生长速度 ,发现 FL- h MMS2 -细胞系在地塞米松诱导下h MMS2基因表达被反义抑制后导致细胞生长受阻 ,提示 h MMS2基因为细胞生长所必需 .     

c－k－ras反义寡核苷酸抑制HCe8693细胞生长及p21~（ras）表达的序列特异性

本文采用逆转录－聚合酶链式反应（ＲＴ－ＰＣＲ），并结合ＰＣＲ产物直接测序法，证明人大肠癌细胞系ＨＲ８３４８和ＨＣｅ８６９３的ｋ－ｒａｓｃＤＮＡ片段（第３～８３密码子）中存在第１２密码子ＧＧＴ→ＧＡＴ突变，未发现其它突变或ＲＮＡ剪接异常．用分别与野生型和上述突变型ｋ－ｒａｓ第９～１５密码子互补的１８聚硫代磷酸型寡核苷酸（两者仅存在一个碱基差别）及随机设计的对照寡核苷酸（终浓度为６μｍｏｌ·Ｌ－１），发现与突变型ｋ－ｒａｓ互补的寡核苷酸能抑制ＨＣｅ８６９３细胞生长并有ｐ２１ｒａｓ蛋白表达量的抑制；而与野生型ｋ－ｒａｓ互补的寡核苷酸及随机合成的对照寡核苷酸对之无明显抑制作用，说明ｃ－ｋ－ｒａｓ反义寡核苷酸对ＨＣｅ８６９３细胞生长和ｐ２１ｒａｓ蛋白合成的抑制作用存在明显的核苷酸序列特异性．