Oxygen Species In Cancer Cells Research Articles

New pyranoquinoline derivatives as vascular-disrupting anticancer agents

A series of new 4-aryl-pyranoquinoline derivatives with a focus on meta-nitro and meta-halophenyl derivatives were prepared and investigated for their structure-dependent antiproliferative effects on a panel of six human cancer cell lines. The compounds were highly active with nanomolar IC50 values. Some of them even exceeded the activities of known analogs such as LY290181 in vitro while not affecting non-malignant fibroblasts. These most active derivatives led to an increase of reactive oxygen species in cancer cells and to a disruption of their microtubular cytoskeleton by inhibiting tubulin polymerization. They also displayed vascular-disrupting activity in ovo as assessed by chorioallantoic membrane assays.

Discovery of a new class of MTH1 inhibitor by X-ray crystallographic screening

MutT homologue 1 (MTH1) protects the nucleotide pool from oxidative stress by hydrolyzing oxidized nucleoside triphosphates and prevents their incorporation into DNA. Cancer cells are dependent on the MTH1 activity for survival due to the high-level of reactive oxygen species in cancer cells; therefore, MTH1 is considered to be a novel target for treatment of various cancers. Here, we show by X-ray crystallographic screening using an in-house cocktail library that α-mangostin, a natural xanthone from mangosteen pericarp, binds to the active site of MTH1. A subsequent inhibition assay revealed that 3-isomangostin, a cyclized derivative of α-mangostin, was the most potent MTH1 inhibitor, with an IC50 value of 0.052 μM. Detailed structural analyses of the MTH1-3-isomangostin complex showed the novel binding mode of 3-isomangostin. Our results demonstrate that X-ray crystallographic screening is useful for the lead discovery for MTH1, and suggest that 3-isomangostin would be an attractive chemical tool for the development of anticancer agents.

Effect of High-dose Vitamin C Combined With Anti-cancer Treatment on Breast Cancer Cells.

The anti-cancer effect of high doses of intravenous vitamin C (high-dose vitamin C) remains controversial despite growing evidence that high-dose vitamin C exerts anti-tumorigenic activity by increasing the amount of reactive oxygen species in cancer cells without meaningful toxicities. Therefore, this study attempted to demonstrate the in vitro anti-cancer activity of high-dose vitamin C in combination with conventional treatment in breast cancer. The pro-apoptotic effects of high-dose vitamin C (1.25 to 20 mM) with or without anti-cancer agents (eribulin mesylate, tamoxifen, fulvestrant, or trastuzumab) were estimated using an MTT assay to measure the cell viability of a variety of breast cancer cell lines (MCF7, SK-BR3, and MDA-MB-231), as well as normal breast epithelial cells (MCF10A). High-dose vitamin C (≥10 mM) significantly decreased cell viability of all breast cancer cell lines, particularly of MCF-7 cells. The catalase activities of MCF7 and MDA-MD-231 cells were also lower than those of MCF10A cells. Moreover, cell viability of both MCF7 and MDA-MD-231 cells was decreased further when combining high-dose vitamin C and eribulin mesylate, and this was also true for MCF-7 cells when combining high-dose vitamin C with tamoxifen or fulvestrant and for SK-BR3 cells when combining high-dose vitamin C with trastuzumab in comparison with chemotherapy or endocrine therapy alone. Combining high-dose vitamin C with conventional anti-cancer drugs can have therapeutic advantages against breast cancer cells.

Synthesis of a Block Copolymer Exhibiting Cell-Responsive Phytochemical Release for Cancer Therapy.

Phytochemicals constitute a promising class of therapeutics for the treatment of various diseases, but their delivery poses significant challenges. In this work, a nanoscale polyactive emulsion was designed for smart, cell-responsive delivery of a curcumin prodrug (curcumin dicarboxylate, CDA) that was chemically conjugated to enzymatically labile oligo-peptides with polycaprolactone (PCL) as the carrier. Matrix metalloproteinase (MMP)-sensitive (PLGLYAL) or nonsensitive (GPYYPLG) peptides were used as spacers for conjugating CDA and PCL. This CDA nanoemulsion incorporating the MMP-sensitive sequence exhibited markedly higher anti-cancer activity, cell internalization, and generation of reactive oxygen species in cancer cells in vitro than the control with the nonsensitive oligopeptide. Moreover, the nanopolyactives induced minimal cytotoxicity in noncancerous cell line. This work presents a unique strategy to engineer smart nano-polyactives for efficient and targeted delivery of phytochemicals.

Transferrin-targeted, resveratrol-loaded liposomes for the treatment of glioblastoma

Glioblastomas (GBMs) are highly aggressive brain tumors with a very grim prognosis even after multi-modal therapeutic regimens. Conventional chemotherapeutic agents frequently lead to drug resistance and result in severe toxicities to non-cancerous tissues. Resveratrol (RES), a natural polyphenol with pleiotropic health benefits, has proven chemopreventive effects in all the stages of cancer including initiation, promotion and progression. However, the poor physico-chemical properties of RES severely limit its use as a free drug. In this study, RES was loaded into PEGylated liposomes (RES-L) to counter its drawbacks as a free drug. Since transferrin receptors (TfRs) are up-regulated in GBM, the liposome surface was modified with transferrin moieties (Tf-RES-L) to make them cancer cell-specific. The liposomal nanomedicines developed in this project were aimed at enhancing the physico-chemical properties of RES and exploiting the passive and active targeting capabilities of liposomes to effectively treat GBM.The RES-L were stable, had a good drug-loading capacity, prolonged drug-release in vitro and were easily scalable. Flow cytometry and confocal microscopy were used to study the association with, and internalization of, Tf-L into U-87 MG cells. The Tf-RES-Ls were significantly more cytotoxic and induced higher levels of apoptosis accompanied by activation of caspases 3/7 in GBM cells when compared to free RES or RES-L. The ability of RES to arrest cells in the S-phase of the cell cycle, and selectively induce production of reactive oxygen species in cancer cells were probably responsible for its cytotoxic effects. The therapeutic efficacy of RES formulations was evaluated in a subcutaneous xenograft mouse model of GBM. A tumor growth inhibition study and a modified survival study showed that Tf-RES-Ls were more effective than other treatments in their ability to inhibit tumor growth and improve survival in mice. Overall, the liposomal nanomedicines of RES developed in this project exhibited favorable in vitro and in vivo efficacies, which warrant their further investigation for the treatment of GBMs.

Antitumor effect of Japanese apricot extract (MK615) on human cancer cells in vitro and in vivo through a reactive oxygen species-dependent mechanism.

MK615 is produced from Japanese apricot and contains several cyclic triterpenes, such as oleanolic and ursolic acids. MK615 was shown to strongly suppress cutaneous in-transit metastasis in a patient with malignant melanoma. The present investigation was undertaken to clarify the antitumor effects of MK615 in vitro and in vivo. Several human cancer cell lines were exposed to MK615 for 7 days to examine its antiproliferative effects. The effect of MK615 on in vivo growth of human pancreatic cancer MIAPaCa-2 cells was also examined. MK615 inhibited the growth of several human cancer cell lines in a concentration-dependent way. Pancreatic cancer MIAPaCa-2 cells were highly sensitive to the growth-inhibiting effects of MK615. Treatment with MK615 preferentially induced cell death in human cancer cells while sparing normal cells such as human umbilical vein endothelial cells (HUVEC) and mouse bone marrow cells. When MIAPaCa-2 cells were incubated with MK615 in the presence of antioxidant, growth-inhibition was significantly reduced, and MK615 induced the accumulation of reactive oxygen species in cancer cells but not in HUVEC. MK615, in both the presence and absence of gemcitabine, significantly inhibited the growth of human pancreatic cancer cells as xenografts without apparent adverse effects. MK615, a supplement produced from Japanese apricot, may have therapeutic value in treating human cancers through a reactive oxygen species-dependent mechanism.

Structure of the human monomeric NEET protein MiNT and its role in regulating iron and reactive oxygen species in cancer cells

The NEET family is a relatively new class of three related [2Fe-2S] proteins (CISD1-3), important in human health and disease. While there has been growing interest in the homodimeric gene products of CISD1 (mitoNEET) and CISD2 (NAF-1), the importance of the inner mitochondrial CISD3 protein has only recently been recognized in cancer. The CISD3 gene encodes for a monomeric protein that contains two [2Fe-2S] CDGSH motifs, which we term mitochondrial inner NEET protein (MiNT). It folds with a pseudosymmetrical fold that provides a hydrophobic motif on one side and a relatively hydrophilic surface on the diametrically opposed surface. Interestingly, as shown by molecular dynamics simulation, the protein displays distinct asymmetrical backbone motions, unlike its homodimeric counterparts that face the cytosolic side of the outer mitochondrial membrane/endoplasmic reticulum (ER). However, like its counterparts, our biological studies indicate that knockdown of MiNT leads to increased accumulation of mitochondrial labile iron, as well as increased mitochondrial reactive oxygen production. Taken together, our study suggests that the MiNT protein functions in the same pathway as its homodimeric counterparts (mitoNEET and NAF-1), and could be a key player in this pathway within the mitochondria. As such, it represents a target for anticancer or antidiabetic drug development.

Exploiting mitochondrial and oxidative vulnerabilities with a synthetic analog of pancratistatin in combination with piperlongumine for cancer therapy.

Harsh adverse effects as a result of nonspecific targeting of chemotherapeutics currently pose obstacles in cancer therapy; thus, it would be invaluable to devise novel approaches to specifically target cancer cells. The natural compound pancratistatin (PST) has been shown to preferentially induce apoptosis in a variety of cancer cell types. Recently, several analogs of PST were shown to be efficacious in inducing apoptosis in a variety of aggressive cancer cell types via cancer cell mitochondrial targeting; it caused dissipation of mitochondrial membrane potential and decreased oxygen consumption, and with isolated mitochondria, it induced the release of apoptogenic factors. The natural compound piperlongumine has been shown to target the stress response to reactive oxygen species in cancer cells. We explored the combinatorial potential of two small molecules (SVTH-6 and piperlongumine) that target these vulnerabilities in cancer cells. Interestingly, when combined with the PST analog, SVTH-6, an increase in mitochondrial dysfunction was observed, leading to an enhanced cytotoxic effect against several human cancer cell types. Additionally, this combination treatment was effective in reducing cancer cell growth in physiologically more relevant 3-dimensional spheroid cell cultures. This enhanced effect was found to be dependent on reactive oxygen species generation because an antioxidant could rescue cancer cells from this combination treatment. Importantly, noncancerous cells were markedly less sensitive to this combination treatment. Thus, targeting mitochondrial and oxidative stress vulnerabilities of cancer cells could be an effective strategy for cancer therapy.-Ma, D., Gilbert, T., Pignanelli, C., Tarade, D., Noel, M., Mansour, F., Gupta, M., Ma, S., Ropat, J., Curran, C., Vshyvenko, S., Hudlicky, T., Pandey. S. Exploiting mitochondrial and oxidative vulnerabilities with a synthetic analog of pancratistatin in combination with piperlongumine for cancer therapy.

The quinone-based derivative, HMNQ induces apoptotic and autophagic cell death by modulating reactive oxygen species in cancer cells.

8-Hydroxy-2-methoxy-1,4-naphthoquinone (HMNQ), a natural compound isolated from the bark of Juglans sinensis Dode, displays cytotoxic activity against various human cancer cells. However, the molecular mechanism of the anticancer effect is unclear. In this study, we examined the cytotoxic mechanism of HMNQ at the molecular level in human cancer cells. Cells were treated with HMNQ in a dose- or time-dependent manner. HMNQ treatment inhibited cell viability, colony formation and cell migration, indicating that HMNQ induced cancer cell death. HMNQ-treated cells resulted in apoptotic cell death through PARP-1 cleavage, Bax upregulation and Bcl-2 downregulation. HMNQ was also observed to induce autophagy by upregulating Beclin-1 and LC3. Furthermore, HMNQ induced reactive oxygen species (ROS) production, which was attenuated by the ROS scavengers, NAC and GSH. Finally, HMNQ increased expression of JNK phosphorylation and the JNK inhibitor SP600125 rescued HMNQ-induced cell death, suggesting that the cytotoxicity of HMNQ is mediated by the JNK signaling pathway. Taken together, our findings show that HMNQ exhibits anticancer activity through induction of ROS-mediated apoptosis and autophagy in human cancer cells. These data suggest the potential value of HMNQ as a natural anticancer drug.

GSH-triggered size increase of porphyrin-containing nanosystems for enhanced retention and photodynamic activity.

Long retention of therapeutic agents in tumors is imperative for improving the therapeutic index. In this work, the GSH responsive porphyrin molecule (TPP 1) was synthesized, which is amphiphilic and linked by a disulfide bond. TPP 1 molecules could self-assemble into nanoparticles (TPP 1 NPs) with a size of about 100 nm in aqueous solution. The TPP 1 NPs exhibited high stability under different conditions and could form into large microparticles in the presence of glutathione (GSH). The TPP 1 NPs could be internalized by cancer cells, and they emitted enhanced red fluorescence compared to that of TPP 2 NPs (non-sensitive NPs) when cells were pretreated with GSH. In addition, in vitro MTT assays showed that TPP 1 NPs were biocompatible and could further be used as photosensitizers in nanoparticle formation. The cellular photodynamic activity of TPP 1 NPs was obviously higher than that of TPP 2 NPs, due to the increasing retention of TPP in cancer cells, which will generate more reactive oxygen species in cancer cells under light irradiation. These results highlight the potential of developing stimulus responsive nanoparticles for enhanced retention and improved therapeutic outcome.

Flow-cytometric analysis of reactive oxygen species in cancer cells under treatment with brassinosteroids

To explore the underlying mechanism of cancer cell growth inhibition by brassinosteroids (BS), reactive oxygen species (ROS) generation under treatment with 28-homocastasterone and its synthetic derivatives (22S,23S)-28-homocastasterone was measured in A549 human lung adenocarcinoma cells. BS induced ROS generation in A549 cells and their growth in a time and dose-dependent manner. The maximal effect was observed for (22S,23S)-28-homocastasterone which at 30μM concentration showed a 6-fold increase of ROS generation in comparison with the control.

Redox-Hypersensitive Organic Nanoparticles for Selective Treatment of Cancer Cells

The diselenide-containing fluorescent molecules (SeBDP) and antitumor drug paclitaxel (SePTX) were synthesized and used for constructing SeBDP nanoparticles (SeBDP NPs) and SePTX NPs in aqueous solution through nanoprecipitation method. Both SeBDP NPs and SePTX NPs exhibit high stability and excellent reduction-sensitivity. More interestingly, SeBDP and SePTX could coassemble into uniform and spherical nanoparticles (co-NPs) with dual functions of fluorescence imaging and antitumor activity. These organic NPs could be internalized by different cells as revealed by confocal laser microscopy. Importantly, the co-NPs exhibited selectivity of cytotoxicities between cancerous and normal cells. The cellular proliferation inhibition toward tumor cells (including HeLa and MCF-7 cells) was obviously higher than that toward normal cells (BEAS-2B and L929 cells), which might be attributed to the increasing reactive oxygen species in cancer cells treated by diselenide-containing NPs. These results highlight the poten...

One-pot synthesis of FePt/CNTs nanocomposites for efficient cellular imaging and cancer therapy

Here, we developed a facile route to synthesize carbon nanotube-based FePt nanocomposites (FePt/CNTs) as a potential theranostic platform in the cancer treatment. FePt/CNTs were firstly synthesized via one-pot polyol route, and then functionalized with 6-arm-polyethylene glycol-amine polymer. The average size of FePt nanoparticles (NPs) is 3–4 nm, which is dispersed on the CNT surface (ca.50–150 nm). The as-prepared FePt NPs display high cytotoxicity by highly reactive oxygen species in cancer cells. Folic acid and fluorescein isothiocyanate are assembled onto the surface of FePt/CNTs for effective targeting of folate receptor-positive cancer cells and simultaneously for the visualization of cellular uptake. Therefore, the FePt/CNTs NPs capability of simultaneously performing diagnosis, therapy, and targeting is, therefore, promising for future potential widespread application in biomedicine.

Voltage Dependent Anion Channels (VDAC) and Regulation of Mitochondrial Metabolism

Voltage dependent anion channels (VDAC) are responsible for permeability of mitochondrial outer membranes to hydrophilic metabolites like ATP, ADP and respiratory substrates. Although previously assumed to remain open, VDAC closure is emerging as an important mechanism for regulation of global mitochondrial function. Acetaldehyde formation from hepatic ethanol metabolism leads to VDAC closure, which suppresses exchange of mitochondrial metabolites and inhibits ureagenesis. In vivo, VDAC closure after ethanol occurs coordinately with mitochondrial depolarization. Together, VDAC closure and uncoupling foster selective and more rapid mitochondrial oxidation of toxic acetaldehyde formed by ethanol metabolism. Glycolysis and suppression of mitochondrial metabolism are metabolic characteristics of cancer cells that promote cell proliferation (Warburg phenomenon). High free αβ-tubulin dimers in cancer cells block VDAC and suppress mitochondrial function, and reversal of tubulin inhibition of VDAC with erastin and erastin-like small molecules has an anti-Warburg effect that hyperpolarizes mitochondria, enhances oxidative phosphorylation and decreases glycolysis. Erastin-like compounds also enhance mitochondria formation of reactive oxygen species in cancer cells, leading eventually to mitochondrial dysfunction and cell death. Thus, VDAC is a key regulator of mitochondrial function (DK073336, DK037034 and 14.Z50.31.0028).

Synthesis and evaluation of Tc-99m DTPA–glutathione as a non-invasive tumor imaging agent in a mouse colon cancer model

Glutathione (GSH) plays a critical role in detoxification reactions by reducing the levels of reactive oxygen species in cancer cells. This study aimed to develop technetium (Tc)-99m diethylenetriaminepentaacetic acid (DTPA)-GSH as a tumor imaging agent, and to evaluate the diagnostic performance of Tc-99m DTPA-GSH in terms of its ability to differentiate tumors from inflammatory lesions. DTPA-GSH was synthesized by reaction of GSH with DTPA anhydride under anhydrous conditions in a nitrogen atmosphere. DTPA-GSH was then reacted with Tc-99m sodium pertechnetate in a tin (II) chloride (SnCl2) solution. Gamma camera imaging was performed after intravenous injection of Tc-99m DTPA-GSH into a mouse CT-26 colon cancer model, or a mouse model of inflammation induced by the intramuscular injection of Freund's complete adjuvant. DTPA-GSH was successfully prepared via a straightforward synthetic procedure and radiolabeled with Tc-99m at a high labeling efficiency (>95%). Tc-99m DTPA-GSH was strongly internalized by tumors in colon cancer model mice, with the tumor-to-normal muscle ratio of the complex reaching 4.3±0.9 at 4 h. By contrast, Tc-99m DTPA-GSH showed relatively weak uptake in inflammatory lesions (target-to-non-target ratio=2.0±0.3 at 4 h). A competition study showed that the uptake of Tc-99m DTPA-GSH into tumors was blocked by co-injection with high concentrations of free GSH. The results of this work indicate that Tc-99m DTPA-GSH is a good candidate for development as a non-invasive tumor imaging agent. Furthermore, Tc-99m DTPA-GSH effectively distinguished between cancerous tissue and inflammatory lesions.

Mitochondrial reactive oxygen species accelerate the expression of heme carrier protein 1 and enhance photodynamic cancer therapy effect.

Photodynamic therapy using hematoporphyrin and its derivatives is clinically useful for cancer treatments. It has been reported that cancer cells incorporate hematoporphyrin and its derivatives via heme carrier protein 1, which is a proton-coupled folate transporter. However, the mechanism of this protein expression has not been elucidated. In general, the concentration of reactive oxygen species in cancer cells is higher than that in normal cells. We previously reported that reactive oxygen species from mitochondria involved in the expression of peptide transporter 1 and accelerate the uptake of 5-aminolevulinic acid, which is a precursor of protoporphyrin IX. We suggested mitochondrial reactive oxygen species also regulated the expression of heme carrier protein 1. In this study, we used a rat gastric mucosal cell line RGM1 and its cancer-like mutated cell line RGK1. We clarified the expression of heme carrier protein 1 increased in cancer cells and it decreased in manganese superoxide dismutase expressed cancer cells. In addition, the uptake level of hematoporphyrin and photodynamic therapeutic effect were also decreased in manganese superoxide dismutase expressed cancer cells in comparison with cancer cells. Thus, we concluded that mitochondrial reactive oxygen species regulated heme carrier protein 1 expression and photodynamic therapeutic effect.

Apoptotic signaling through reactive oxygen species in cancer cells

Apoptotic signaling through reactive oxygen species in cancer cells

Antitumor Effect of Japanese Apricot Extract (MK615) on Human Cancer Cells in Vitro and in Vivo through a Reactive Oxygen Species-Dependent Mechanism

MK615 is produced from Japanese apricot and contains several cyclic triterpenes, such as oleanolic and ursolic acids. MK615 was shown to strongly suppress cutaneous in-transit metastasis in a patient with malignant melanoma. The present investigation was undertaken to clarify the antitumor effects of MK615 in vitro and in vivo. Several human cancer cell lines were exposed to MK615 for 7 days to examine its antiproliferative effects. The effect of MK615 on in vivo growth of human pancreatic cancer MIAPaCa-2 cells was also examined. MK615 inhibited the growth of several human cancer cell lines in a concentration-dependent way. Pancreatic cancer MIAPaCa-2 cells were highly sensitive to the growth-inhibiting effects of MK615. Treatment with MK615 preferentially induced cell death in human cancer cells while sparing normal cells such as human umbilical vein endothelial cells (HUVEC) and mouse bone marrow cells. When MIAPaCa-2 cells were incubated with MK615 in the presence of antioxidant, growth-inhibition was significantly reduced, and MK615 induced the accumulation of reactive oxygen species in cancer cells but not in HUVEC. MK615, in both the presence and absence of gemcitabine, significantly inhibited the growth of human pancreatic cancer cells as xenografts without apparent adverse effects. MK615, a supplement produced from Japanese apricot, may have therapeutic value in treating human cancers through a reactive oxygen species-dependent mechanism.

ROS-activated Anticancer Prodrugs: a New Strategy for tumor-specific Damage

Targeting tumor cells is an important strategy to improve the selectivity of cancer therapies. With the advanced studies in cancer biology, we know that cancer cells are usually under increased oxidative stress. The high level of reactive oxygen species in cancer cells has been exploited for developing novel therapeutic strategies to preferentially kill cancer cells. Our group, amongst others, have used boronic acids/esters as triggers for developing ROS-activated anticancer prodrugs that target cancer cells. The selectivity was achieved by combining a specific reaction between boronates and H2O2, with the efficient masking of drug toxicity in the prodrug via boronates. Prodrugs activated via ferrocene-mediated oxidation have also been developed to improve the selectivity of anticancer drugs. We describe how the strategies of ROS-activation can be used for further development of new ROS-targeting prodrugs, eventually leading to novel approaches and/or combined technology for more efficient and selective treatment of cancers.

The Effect of Methoxyflavone on P-glycoprotein- Mediated Transport and Intracellular Reactive Oxygen Species in Cancer Cells

The Effect of Methoxyflavone on P-glycoprotein- Mediated Transport and Intracellular Reactive Oxygen Species in Cancer Cells