Younghyun Lee

Genotoxic effects of 3 T magnetic resonance imaging in cultured human lymphocytes.

The clinical and preclinical use of high-field intensity (HF, 3 T and above) magnetic resonance imaging (MRI) scanners have significantly increased in the past few years. However, potential health risks are implied in the MRI and especially HF MRI environment due to high-static magnetic fields, fast gradient magnetic fields, and strong radiofrequency electromagnetic fields. In this study, the genotoxic potential of 3 T clinical MRI scans in cultured human lymphocytes in vitro was investigated by analyzing chromosome aberrations (CA), micronuclei (MN), and single-cell gel electrophoresis. Human lymphocytes were exposed to electromagnetic fields generated during MRI scanning (clinical routine brain examination protocols: three-channel head coil) for 22, 45, 67, and 89 min. We observed a significant increase in the frequency of single-strand DNA breaks following exposure to a 3 T MRI. In addition, the frequency of both CAs and MN in exposed cells increased in a time-dependent manner. The frequencies of MN in lymphocytes exposed to complex electromagnetic fields for 0, 22, 45, 67, and 89 min were 9.67, 11.67, 14.67, 18.00, and 20.33 per 1000 cells, respectively. Similarly, the frequencies of CAs in lymphocytes exposed for 0, 45, 67, and 89 min were 1.33, 2.33, 3.67, and 4.67 per 200 cells, respectively. These results suggest that exposure to 3 T MRI induces genotoxic effects in human lymphocytes.

Enhanced cytotoxic and genotoxic effects of gadolinium following ELF-EMF irradiation in human lymphocytes

Abstract Gadolinium (Gd) and its chelated derivatives are widely utilized for various industrial and medical purposes, particularly as a contrast agent for magnetic resonance imaging (MRI). There are many studies of Gd nephrotoxicity and neurotoxicity, whereas research on cyto- and genotoxicity in normal human lymphocytes is scarce. It is important to investigate the effect of extremely low-frequency electromagnetic fields (ELF-EMF) on Gd toxicity, as patients are co-exposed to Gd and ELF-EMF generated by MRI scanners. We investigated the cytotoxicity and genotoixcity of Gd and the possible enhancing effect of ELF-EMF on Gd toxicity in cultured human lymphocytes by performing a micronuclei (MN) assay, trypan blue dye exclusion, single cell gel electrophoresis, and apoptosis analyses using flow cytometry. Isolated lymphocytes were exposed to 0.2–1.2 mM of Gd only or in combination with a 60-Hz ELF-EMF of 0.8-mT field strength. Exposing human lymphocytes to Gd resulted in a concentration- and time-dependent decrease in cell viability and an increase in MN frequency, single strand DNA breakage, apoptotic cell death, and ROS production. ELF-EMF (0.8 mT) exposure also increased cell death, MN frequency, olive tail moment, and apoptosis induced by Gd treatment alone. These results suggest that Gd induces DNA damage and apoptotic cell death in human lymphocytes and that ELF-EMF enhances the cytotoxicity and genotoxicity of Gd.

Cyto-/genotoxic effects of the ethanol extract of Chan Su, a traditional Chinese medicine, in human cancer cell lines.

ETHNOPHARMACOLOGICAL RELEVANCE Chan Su, an ethanolic extract from skin and parotid venom glands of the Bufo bufo gargarizans Cantor, is widely used as a traditional Chinese medicine for cancer therapy. Although the anti-cancer properties of Chan Su have been investigated, no information exists regarding whether Chan Su has genotoxic effects in cancer cells. The aim of the present study was to examine the cyto-/genotoxic effect of Chan Su in human breast carcinoma (MCF-7 cells), human lung carcinoma (A-549 cells), human T cell leukemia (Jurkat T cells), and normal human lymphocytes. MATERIALS AND METHODS Effects on the viability of MCF-7, A-549, Jurkat T cells, and normal lymphocytes were evaluated by Trypan blue exclusion assays. The DNA content in the sub-G1 region was detected by propidium iodide (PI) staining and flow cytometry. The genotoxicity of Chan Su was assessed by single-cell gel electrophoresis (comet assay) and the cytokinesis-block micronucleus assay (CBMN assay). RESULTS Chan Su significantly inhibited the viability of MCF-7, A-549, and Jurkat T cells dose dependently, but had no effect on normal human lymphocytes. Apoptotic death of the cancer cells was evident after treatment. Chan Su also induced genotoxicity in a dose-dependent manner, as indicated by the comet and cytokinesis-block micronucleus assays. CONCLUSIONS These findings suggest that Chan Su can induce apoptotic death of, and exert genotoxic effects on, MCF-7, A-549, and Jurkat T cells.

Nontoxic concentration of DNA-PK inhibitor NU7441 radio-sensitizes lung tumor cells with little effect on double strand break repair.

High‐linear energy transfer (LET) heavy ions have been increasingly employed as a useful alternative to conventional photon radiotherapy. As recent studies suggested that high LET radiation mainly affects the nonhomologous end‐joining (NHEJ) pathway of DNA double strand break (DSB) repair, we further investigated this concept by evaluating the combined effect of an NHEJ inhibitor (NU7441) at a non‐toxic concentration and carbon ions. NU7441‐treated non‐small cell lung cancer (NSCLC) A549 and H1299 cells were irradiated with X‐rays and carbon ions (290 MeV/n, 50 keV/μm). Cell survival was measured by clonogenic assay. DNA DSB repair, cell cycle distribution, DNA fragmentation and cellular senescence induction were studied using a flow cytometer. Senescence‐associated protein p21 was detected by western blotting. In the present study, 0.3 μM of NU7441, nontoxic to both normal and tumor cells, caused a significant radio‐sensitization in tumor cells exposed to X‐rays and carbon ions. This concentration did not seem to cause inhibition of DNA DSB repair but induced a significant G2/M arrest, which was particularly emphasized in p53‐null H1299 cells treated with NU7441 and carbon ions. In addition, the combined treatment induced more DNA fragmentation and a higher degree of senescence in H1299 cells than in A549 cells, indicating that DNA‐PK inhibitor contributes to various modes of cell death in a p53‐dependent manner. In summary, NSCLC cells irradiated with carbon ions were radio‐sensitized by a low concentration of DNA‐PK inhibitor NU7441 through a strong G2/M cell cycle arrest. Our findings may contribute to further effective radiotherapy using heavy ions.

Enhancement of cisplatin cytotoxicity by benzyl isothiocyanate in HL-60 cells.

Cis-diamminedichloroplatinum (II) (cisplatin) is one of the most widely used chemotherapeutic drugs, but its effectiveness is limited by tumor cell resistance and the severe side effects it causes. One strategy for overcoming this problem is the concomitant use of natural dietary compounds as therapeutic agents. Benzyl isothiocyanate (BITC) is a promising chemopreventive agent found in cruciferous vegetables and papaya fruits. The aim of this study was to investigate the effects of BITC on cisplatin-induced cytotoxicity in human promyelocytic leukemia cells and normal human lymphocytes. The combined treatment of HL-60 cells with BITC followed by cisplatin (BITC/cisplatin) caused a significant decrease in cell viability. BITC also increased apoptotic cell death compared to cisplatin treatment alone. In normal human lymphocytes, BITC did not enhance the cytotoxic effects of cisplatin. Cellular exposure to BITC/cisplatin increased reactive oxygen species (ROS) generation but decreased the total glutathione (GSH) level in HL-60 cells. Pretreatment of HL-60 cells with N-acetylcysteine or glutathione monoethyl ester effectively decreased BITC/cisplatin-induced cell death. The addition of the extracellular signal-regulated kinase (ERK) inhibitor PD98059 abolished BITC/cisplatin-induced apoptosis. Taken together, our results suggest that BITC enhances cisplatin-induced cytotoxicity through the generation of ROS, depletion of GSH, and ERK signaling in HL-60 cells.

Bakuchiol suppresses proliferation of skin cancer cells by directly targeting Hck, Blk, and p38 MAP kinase

Bakuchiol is a meroterpene present in the medicinal plant Psoralea corylifolia, which has been traditionally used in China, India, Japan and Korea for the treatment of premature ejaculation, knee pain, alopecia spermatorrhea, enuresis, backache, pollakiuria, vitiligo, callus, and psoriasis. Here, we report the chemopreventive properties of bakuchiol, which acts by inhibiting epidermal growth factor (EGF)-induced neoplastic cell transformation. Bakuchiol also decreased viability and inhibited anchorage-independent growth of A431 human epithelial carcinoma cells. Bakuchiol reduced A431 xenograft tumor growth in an in vivo mouse model. Using kinase profiling, we identified Hck, Blk and p38 mitogen activated protein kinase (MAPK) as targets of bakuchiol, which directly bound to each kinase in an ATP-competitive manner. Bakuchiol also inhibited EGF-induced signaling pathways downstream of Hck, Blk and p38 MAPK, including the MEK/ERKs, p38 MAPK/MSK1 and AKT/p70S6K pathways. This report is the first mechanistic study identifying molecular targets for the anticancer activity of bakuchiol and our findings indicate that bakuchiol exhibits potent anticancer activity by targeting Hck, Blk and p38 MAPK.

Radiation-induced changes in DNA methylation and their relationship to chromosome aberrations in nuclear power plant workers

Abstract Purpose: We investigated the association between occupational radiation exposure and DNA methylation changes in nuclear power plant workers. We also evaluated whether radiation- induced DNA methylation alterations are associated with chromosome aberrations. Materials and methods: The study population included 170 radiation-exposed workers and 30 controls. We measured global, long interspersed nuclear element-1 (LINE-1), and satellite 2 methylation levels in blood leukocyte DNA. The analysis of chromosome aberrations was performed on peripheral lymphocytes. Results: Global DNA methylation levels were lower in radiation-exposed workers than in controls. The methylation levels were negatively associated with the recent 1.5-year radiation dose in a multiple linear regression model (β = − 0.0088, p ≤ 0.001); the levels increased proportionally with the total cumulative dose in radiation-exposed workers. LINE-1 methylation levels were higher in radiation-exposed workers than in controls and were significantly associated with the total cumulative radiation dose in a multiple linear regression model (β = − 0.031, p = 0.035). Global DNA methylation levels were also correlated with chromosome aberrations among workers. Workers with low global methylation levels had a higher frequency of chromosome aberrations than did subjects with high global methylation levels. Conclusion: Occupational exposure to low-dose radiation could affect DNA methylation levels, and the radiation-induced DNA methylation alterations may be associated with chromosome aberrations.

TAS-116, a Novel Hsp90 Inhibitor, Selectively Enhances Radiosensitivity of Human Cancer Cells to X-rays and Carbon Ion Radiation.

Hsp90 inhibitors have been investigated as cancer therapeutics in monotherapy and to augment radiotherapy; however, serious adverse effects of early-generation Hsp90 inhibitors limited their development. TAS-116 is a novel Hsp90 inhibitor with lower adverse effects than other Hsp90 inhibitors, and here, we investigated the radiosensitizing effects of TAS-116 in low linear energy transfer (LET) X-ray and high LET carbon ion–irradiated human cancer cells and mouse tumor xenografts. TAS-116 decreased cell survival of both X-ray and carbon ion–irradiated human cancer cell lines (HeLa and H1299 cells), and similar to other Hsp90 inhibitors, it did not affect radiosensitivity of noncancerous human fibroblasts. TAS-116 increased the number of radiation-induced γ-H2AX foci and delayed the repair of DNA double-strand breaks (DSB). TAS-116 reduced the expression of proteins that mediate repair of DSBs by homologous recombination (RAD51) and nonhomologous end joining (Ku, DNA-PKcs), and suppressed formation of RAD51 foci and phosphorylation/activation of DNA-PKcs. TAS-116 also decreased expression of the cdc25 cell-cycle progression marker, markedly increasing G2–M arrest. Combined treatment of mouse tumor xenografts with carbon ions and TAS-116 showed promising delay in tumor growth compared with either individual treatment. These results demonstrate that TAS-116 radiosensitizes human cancer cells to both X-rays and carbon ions by inhibiting the two major DSB repair pathways, and these effects were accompanied by marked cell-cycle arrest. The promising results of combination TAS-116 + carbon ion radiotherapy of tumor xenografts justify further exploration of TAS-116 as an adjunct to radiotherapy using low or high LET radiation. Mol Cancer Ther; 16(1); 16–24. ©2016 AACR.

The purine scaffold Hsp90 inhibitor PU-H71 sensitizes cancer cells to heavy ion radiation by inhibiting DNA repair by homologous recombination and non-homologous end joining

BACKGROUND AND PURPOSE PU-H71 is a purine-scaffold Hsp90 inhibitor developed to overcome limitations of conventional Hsp90 inhibitors. This study was designed to investigate the combined effect of PU-H71 and heavy ion irradiation on human tumor and normal cells. MATERIALS AND METHODS The effects of PU-H71 were determined by monitoring cell survival by colony formation, and DNA double-strand break (DSB) repair by γ-H2AX foci and immuno-blotting DSB repair proteins. The mode of cell death was evaluated by sub-G1 DNA content (as an indicator for apoptosis), and mitotic catastrophe. RESULTS PU-H71 enhanced heavy ion irradiation-induced cell death in three human cancer cell lines, but the drug did not radiosensitize normal human fibroblasts. In irradiated tumor cells, PU-H71 increased the persistence of γ-H2AX foci, and it reduced RAD51 foci and phosphorylated DNA-PKcs, key DSB repair proteins involved in homologous recombination (HR) and non-homologous end joining (NHEJ). In some tumor cell lines, PU-H71 altered the sub-G1 cell fraction and mitotic catastrophe following carbon ion irradiation. CONCLUSION Our results demonstrate that PU-H71 sensitizes human cancer cells to heavy ion irradiation by inhibiting both HR and NHEJ DSB repair pathways. PU-H71 holds promise as a radiosensitizer for enhancing the efficacy of heavy ion radiotherapy.

Selective Effects of Curcumin on CdSe/ZnS Quantum-dot-induced Phototoxicity Using UVA Irradiation in Normal Human Lymphocytes and Leukemia Cells

Quantum dots (QDs) have received considerable attention due to their potential role in photosensitization during photodynamic therapy. Although QDS are attractive nanomaterials due to their novel and unique physicochemical properties, concerns about their toxicity remain. We suggest a combination strategy, CdSe/ZnS QDs together with curcumin, a natural yellow pigment from turmeric, to reduce QD-induced cytotoxicity. The aim of this study was to explore a potentially effective cancer treatment: co-exposure of HL-60 cells and human normal lymphocytes to CdSe/ZnS QDs and curcumin. Cell viability, apoptosis, reactive oxygen species (ROS) generation, and DNA damage induced by QDs and/or curcumin with or without ultraviolet A (UVA) irradiation were evaluated in both HL-60 cells and normal lymphocytes. In HL-60 cells, cell death, apoptosis, ROS generation, and single/double DNA strand breaks induced by QDs were enhanced by treatment with curcumin and UVA irradiation. The protective effects of curcumin on cell viability, apoptosis, and ROS generation were observed in normal lymphocytes, but not leukemia cells. These results demonstrated that treatment with QD combined with curcumin increased cell death in HL-60 cells, which was mediated by ROS generation. However, curcumin acted as an antioxidant in cultured human normal lymphocytes.