The prospective application of a hypoxic radiosensitizer, doranidazole to rat intracranial glioblastoma with blood brain barrier disruption.

BACKGROUND: Glioblastoma is one of the intractable cancers and is highly resistant to ionizing radiation. This radioresistance is partly due to the presence of a hypoxic region which is widely found in advanced malignant gliomas. In the present study, we evaluated the effectiveness of the hypoxic cell sensitizer doranidazole (PR-350) using the C6 rat glioblastoma model, focusing on the status of blood brain barrier (BBB). METHODS: Reproductive cell death in the rat C6 glioma cell line was determined by means of clonogenic assay. An intracranial C6 glioma model was established for the in vivo experiments. To investigate the status of the BBB in C6 glioma bearing brain, we performed the Evans blue extravasation test. Autoradiography with [(14)C]-doranidazole was performed to examine the distribution of doranidazole in the glioma tumor. T2-weighted MRI was employed to examine the effects of X-irradiation and/or doranidazole on tumor growth. RESULTS: Doranidazole significantly enhanced radiation-induced reproductive cell death in vitro under hypoxia, but not under normoxia. The BBB in C6-bearing brain was completely disrupted and [(14)C]-doranidazole specifically penetrated the tumor regions. Combined treatment with X-irradiation and doranidazole significantly inhibited the growth of C6 gliomas. CONCLUSIONS: Our results revealed that BBB disruption in glioma enables BBB-impermeable radiosensitizers to penetrate and distribute in the target region. This study is the first to propose that in malignant glioma the administration of hydrophilic hypoxic radiosensitizers could be a potent strategy for improving the clinical outcome of radiotherapy without side effects.

Functional expression of choline transporter-like protein 1 (CTL1) in small cell lung carcinoma cells: a target molecule for lung cancer therapy.

Choline is essential for the synthesis of the major membrane phospholipid phosphatidylcholine and the neurotransmitter acetylcholine (ACh). Elevated levels of choline and up-regulated choline kinase activity have been detected in cancer cells. Thus, the intracellular accumulation of choline through choline transporters is the rate-limiting step in phospholipid metabolism and a prerequisite for cancer cell proliferation. However, the uptake system for choline and the functional expression of choline transporters in lung cancer cells are poorly understood. We examined the molecular and functional characterization of choline uptake in the small cell lung carcinoma cell line NCI-H69. Choline uptake was saturable and mediated by a single transport system. Interestingly, removal of Na(+) from the uptake buffer strongly enhanced choline uptake. This increase in choline uptake under the Na(+)-free conditions was inhibited by dimethylamiloride (DMA), a Na(+)/H(+) exchanger (NHE) inhibitor. Various organic cations and the choline analog hemicholinium-3 (HC-3) inhibited the choline uptake and cell viability. A correlation analysis of the potencies of organic cations for the inhibition of choline uptake and cell viability showed a strong correlation (R=0.8077). RT-PCR revealed that choline transporter-like protein 1 (CTL1) mRNA and NHE1 are mainly expressed. HC-3 and CTL1 siRNA inhibited choline uptake and cell viability, and increased caspase-3/7 activity. The conversion of choline to ACh was confirmed, and this conversion was enhanced under Na(+)-free conditions, which in turn was sensitive to HC-3. These results indicate that choline uptake through CTL1 is used for ACh synthesis. Both an acetylcholinesterase inhibitor (eserine) and a butyrylcholinesterase inhibitor (ethopropazine) increased cell proliferation, and these effects were inhibited by 4-DAMP, a mAChR3 antagonist. We conclude that NCI-H69 cells express the choline transporter CTL1 which uses a directed H(+) gradient as a driving force, and its transport functions in co-operation with NHE1. This system primarily supplies choline for the synthesis of ACh and secretes ACh to act as an autocrine/paracrine growth factor, and the functional inhibition of CTL1 could promote apoptotic cell death. Identification of this new CTL1-mediated choline transport system provides a potential new target for therapeutic intervention.

Animal model of dermatophytosis.

Dermatophytosis is superficial fungal infection caused by dermatophytes that invade the keratinized tissue of humans and animals. Lesions from dermatophytosis exhibit an inflammatory reaction induced to eliminate the invading fungi by using the host's normal immune function. Many scientists have attempted to establish an experimental animal model to elucidate the pathogenesis of human dermatophytosis and evaluate drug efficacy. However, current animal models have several issues. In the present paper, we surveyed reports about the methodology of the dermatophytosis animal model for tinea corporis, tinea pedis, and tinea unguium and discussed future prospects.

Establishment of a novel model of onychomycosis in rabbits for evaluation of antifungal agents.

We developed a novel model of onychomycosis in which we observed fungi in the deep layer of the nail, and we used the model to evaluate the efficacy of two topical antifungal drugs. To establish an experimental, in vivo model of onychomycosis, we applied Trichophyton mentagrophytes TIMM2789 to the nails of the hind limbs of rabbits that underwent steroid treatment. The nails were taken from the rabbits' feet at 0, 2, and 6 weeks after a 2-week infection. The localization of the fungi was evaluated histopathologically. Some fungi were seen to penetrate to the nail bed, and the infection rate in the sample at 0, 2, and 6 weeks after infection was 57, 87, and 93%, respectively. In addition, fungi proliferated and moved proximally into the nail plate in a manner that depended on the duration of infection. Second, using this model we evaluated antifungal efficacy both by the culture recovery method and histopathological examination. Two topical antifungal drugs, 8% ciclopirox nail lacquer and 5% amorolfine nail lacquer, were applied to the nail for 4 weeks in each group. On histopathological examination, two antifungal treatment groups showed no significant difference against the nontreated control group. However, there were a significantly low fungus-positive rate and intensity of the recovery of fungi on culture between antifungal treatment and nontreated control groups. We therefore suggest that we have established an in vivo model of onychomycosis that is useful for the evaluation of the efficacy of antifungal agents.

Ascorbic acid gives different protective effects in human cells exposed to X-rays and heavy ions.

We studied the effects and mechanisms of ascorbic acid as a radiation protector. Cell survival, repair of DNA double strand breaks (DSBs), and sister chromatid exchanges (SCEs) were examined in normal human fibroblasts irradiated with X-rays and heavy ions. Post-irradiation treatment with 5mM ascorbic acid for 24 h in plateau phase (non-cycling) cells enhanced cell survival and DNA double strand break repair, and reduced SCEs after X-rays irradiation. On the other hand, only reduced SCEs were observed after heavy ion exposure such as to carbon ions. Judging from our data, it is possible that the radioprotective action of ascorbic acid would be effective in non-complex type DNA damage such as induced by X-rays. These findings provide new insight into the mechanism of DNA damage and repair produced by heavy ion irradiation.

NBS1 regulates a novel apoptotic pathway through Bax activation.

DNA damage induced apoptosis, along with precise DNA damage repair, is a critical cellular function, and both of these functions are necessary for cancer prevention. The NBS1 protein is known to be a key regulator of DNA damage repair. It acts by forming a complex with Rad50/Mre11 and by activating ATM. We show here that NBS1 regulates a novel p53 independent apoptotic pathway in response to DNA damage. DNA damage induced apoptosis was significantly reduced in NBS1 deficient cells regardless of their p53 status. Experiments using a series of cell lines expressing mutant NBS1 proteins revealed that NBS1 is able to regulate the activation of Bax and Caspase-3 without the FHA, Mre11-binding, or the ATM-interacting domains, whereas the phosphorylation sites of NBS1 were essential for Bax activation. Expression of apoptosis-related transcription factors such as E2F1 and their downstream pro-apoptotic factors were not related to this apoptosis induction. Interestingly, NBS1 regulates a novel Bax activation pathway by disrupting the Ku70-Bax complex which is required for activation of the mitochondrial apoptotic pathway. This dissociation of the Ku70-Bax complex can be mediated by acetylation of Ku70, and NBS1 can function in this process through a protein-protein interaction with Ku70. Thus, NBS1 is a key protein involved in the prevention of carcinogenesis, not only through the precise repair of damaged DNA by homologous recombination (HR) but also by its role in the elimination of inappropriately repaired cells.

Chemopreventive agent sulforaphane enhances radiosensitivity in human tumor cells.

Sulforaphane (SFN), an isothiocyanate derived from broccoli and other cruciferous vegetables, is a positive regulator of phase II detoxification enzymes and is highly effective in protection against chemically induced cancers by inducing apoptosis and cell cycle arrest. Here, we report that SFN also enhances radiosensitivity in human tumor cells. Cell survival in HeLa human cervix carcinoma cells pretreated with SFN was significantly lower than in cells treated with radiation only. Constant-field gel electrophoresis and a gamma-H2AX foci assay showed marked inhibition of DSB repair in irradiated cells treated with SFN, while little inhibition was observed in cells with DMSO (control). In addition, immunofluorescence experiments revealed a significant delay in Rad51 (a key protein for homologous recombination repair) foci formation and disappearance in irradiated cells treated with SFN when compared to the cells with X-irradiation alone. The dephosphorylation of DNA-PKcs (a critical nonhomologous end joining protein) was also markedly delayed by SFN pretreatment in irradiated cells. These DSB repair inhibition data partially support the high apoptotic frequency of irradiated cells pretreated with SFN. Furthermore, the combined treatment of X-rays and SFN (i.p. 300 micromol/kg) in the xenograft model with HeLa cells showed efficient inhibition of in vivo tumor growth. To the best of our knowledge, our study is the first report showing SFN-enhanced radiosensitivity of tumor cells in vitro and in vivo, which opens the door for a multitude of clinical applications for chemoradiotherapy using SFN.

Sulforaphane induces DNA double strand breaks predominantly repaired by homologous recombination pathway in human cancer cells.

Cytotoxicity and DNA double strand breaks (DSBs) were studied in HeLa cells treated with sulforaphane (SFN), a well-known chemo-preventive agent. Cell survival was impaired by SFN in a concentration and treatment time-dependent manner. Both constant field gel electrophoresis (CFGE) and gamma-H2AX assay unambiguously indicated formation of DSBs by SFN, reflecting the cell survival data. These DSBs were predominantly processed by homologous recombination repair (HRR), judging from the SFN concentration-dependent manner of Rad51 foci formation. On the other hand, the phosphorylation of DNA-PKcs, a key non-homologous end joining (NHEJ) protein, was not observed by SFN treatment, suggesting that NHEJ may not be involved in DSBs induced by this chemical. G2/M arrest by SFN, a typical response for cells exposed to ionizing radiation was also observed. Our new data indicate the clear induction of DSBs by SFN and a useful anti-tumor aspect of SFN through the induction of DNA DSBs.

X irradiation combined with TNF alpha-related apoptosis-inducing ligand (TRAIL) reduces hypoxic regions of human gastric adenocarcinoma xenografts in SCID mice.

Our previous study showed that X irradiation induced the expression of death receptor DR5 on the cell surface in tumor cell lines under not only normoxia but also hypoxia. X irradiation combined with TNF alpha-related apoptosis-inducing ligand (TRAIL), which is the ligand of DR5, induced apoptosis in vitro (Takahashi et al., (2007) Journal of Radiation Research, 48: 461-468). In this report, we examined the in vivo antitumor efficacy of X irradiation combined with TRAIL treatment in tumor xenograft models derived from human gastric adenocarcinoma MKN45 and MKN28 cells in SCID mice. X irradiation combined with TRAIL synergistically suppressed the tumor growth rates in the xenograft models derived from MKN45 and MKN28 cells, which have wild type Tp53 and mutated Tp53, respectively, indicating that the antitumor effects occurred in a Tp53-independent manner. Histological analysis showed that the combination of X irradiation and TRAIL induced caspase-3-dependent apoptotic cell death. Moreover, the immunohistochemical detection of hypoxic regions using the hypoxic marker pimonidazole revealed that caspase-3-dependent apoptosis occurred in the hypoxic regions in the tumors. These results indicated that X irradiation combined with TRAIL may be a useful treatment to reduce tumor growth in not only normoxic but also hypoxic regions.

Preclinical study on hypoxic radiosensitizing effects of glycididazole in comparison with those of doranidazole in vitro and in vivo.

To overcome the radioresistance of hypoxic cells in solid tumor, numerous types of radiosensitizers specifically against them have been developed. Glycididazole has a chemical structure in which two metronidazole forms are combined, and is widely used as a hypoxic radiosensitizer in China. However, a detailed investigation of its radiosensitizing properties has not been performed. The present study reported a comparative assessment of glycididazole and doranidazole, another hypoxic radiosensitizer. All experiments were performed using the murine squamous cell carcinoma cell line SCCVII. Prior to X-irradiation, the cells were treated with the test drugs at concentrations of 10 mM and 200 mg/kg in vitro and in vivo, respectively. Uptake and their intratumor chemical forms were analyzed by high performance liquid chromatography (HPLC). Both drugs enhanced the reproductive cell death induced by X-irradiation under hypoxia. However, the growth delay assay of the transplanted tumor revealed the combination of X-irradiation and glycididazole showed a similar antitumor effect to that of X-irradiation alone, whereas doranidazole significantly sensitized the cells to X-irradiation. HPLC analysis revealed that incorporated glycididazole was decomposed to metronidazole and was therefore present at a lower concentration compared with that of doranidazole. The decomposition of glycididazole to metronidazole reduced its radiosensitizing efficiency in vivo. Elucidation of the kinetics of drugs containing metabolizable chemical forms is necessary for the optimization of clinical treatment.

Transport of D-[1-14C]-amino acids into Chinese hamster ovary (CHO-K1) cells: implications for use of labeled d-amino acids as molecular imaging agents.

INTRODUCTION: The fact that d-amino acids have been found in various tissues and are involved in various functions is a clue to how to develop new imaging agents. We examined d-amino acid transport mechanisms in Chinese hamster ovary (CHO-K1) cells because CHO-K1 cells are widely used in biomedical studies and are thought to be useful for expression of genes involved in metabolism of D-amino acids. METHODS: Uptake experiments were performed. CHO-K1 cells cultured in 60-mm plastic culture dishes under ordinary culture conditions were incubated with 18.5 kBq of radiolabeled amino acid in 2 ml of phosphate-buffered-saline-based uptake solution at 37 degrees C. The following radiolabeled amino acid tracers were used: D-[1-(14)C]-alanine, L-[1-(14)C]-alanine, D-[1-(14)C]-serine, L-[1-(14)C]-serine, D-[1-(14)C]-methionine, L-[1-(14)C]-methionine, D-[1-(14)C]-phenylalanine, L-[1-(14)C]-phenylalanine, D-[1-(14)C]-leucine, L-[1-(14)C]-leucine, D-[1-(14)C]-valine, L-[1-(14)C]-valine, D-[1-(14)C]-tyrosine, L-[1-(14)C]-tyrosine, D-[1-(14)C]-glutamic acid, L-[1-(14)C]-glutamic acid, D-[1-(14)C]-lysine, L-[1-(14)C]-lysine, D-[1-(14)C]-arginine and L-[L-(14)C]-arginine. We tested the inhibitory effects of the following compounds (1.0 mM) on transport: 2-(methylamino)isobutyric acid (a specific inhibitor of system A, in Na(+)-containing uptake solution) and 2-amino-bicyclo[2,2,1]heptane-2-carboxylic acid (a specific inhibitor of system L, in Na(+)-free uptake solution). RESULTS: D-[1-(14)C]-methionine, D-[1-(14)C]-phenylalanine and D-[1-(14)C]-tyrosine accumulated mainly via system L. D-[1-(14)C]-alanine and D-[1-(14)C]-serine accumulated primarily via system ASC. High uptake of D-[1-(14)C]-alanine, D-[1-(14)C]-methionine, D-[1-(14)C]-phenylalanine and D-[1-(14)C]-leucine was observed. The uptake of radiolabeled serine, valine, tyrosine, glutamic acid and arginine into CHO-K1 was highly stereoselective for l-isomers. CONCLUSIONS: We observed high uptake of D-[1-(14)C]-alanine via system ASC (most likely alanine-serine-cysteine-selective amino acid transporter-1) and high uptake of D-[1-(14)C]-methionine and D-[1-(14)C]-phenylalanine via system L (most likely L-type amino acid transporter-1).

Pharmacokinetics of 3-[125I]iodo-alpha-methyl-L-tyrosine, a tumor imaging agent, after probenecid loading in mice implanted with colon cancer DLD-1 cells.

INTRODUCTION: In order to improve tumor imaging, changes in the pharmacokinetics of 3-[123I]iodo-alpha-methyl-l-tyrosine ([123I]IMT), an artificial amino acid that exhibits high tumor accumulation, after probenecid (PBC) loading was studied in mice implanted with colon cancer DLD-1 cells using 125I-labeled IMT ([125I]IMT). METHODS: DLD-1-implanted KSN-slc nude male mice received 740 kBq of [125I]IMT via the tail vein at 5 min after 50 mg/kg body weight PBC loading, and autoradiography was performed at 5, 15 and 30 min after injection. Male ddY mice then received 670 kBq of [125I]IMT and 50 mg/kg 2-amino-bicyclo[2,2,1]heptane-2-carboxylic acid (BCH) or p-aminohippurate (PAH) via the tail vein, and kidney autoradiography was performed at 5 min after injection. In vitro inhibition study was then performed based on the accumulation mechanisms of [125I]IMT in DLD-1, using 1 mM l-tyrosine, BCH, alpha-(methylamino)-isobutyric acid, N-benzoyl-beta-alanine, PBC, PAH, 2,4-dinitrophenol and sodium azide. Both Na+-dependent and Na+-independent uptake were investigated. RESULTS: Higher tumor accumulation in PBC-loaded DLD-1-implanted mice was seen when compared to control mice. PAH and BCH, respectively, reduced renal accumulation in the tubule segment-2 (S2)-like and S1-like regions. We confirmed that [125I]IMT transport is predominantly mediated by l-type amino acid transporter-1 in DLD-1 cells. CONCLUSIONS: [125I]IMT uptake is mediated by organic anion and amino acid transporters in the kidney. Organic anion transporter inhibitors may yield better tumor images with good tumor/normal tissue radioactivity ratios if adequate administration plans are developed.

The chemopreventive flavonoid apigenin confers radiosensitizing effect in human tumor cells grown as monolayers and spheroids.

Apigenin, a common dietary flavonoid present in many fruits and vegetables, is a nonmutagenic chemopreventive agent. In the present study, we investigated the effect of apigenin on the radiosensitivity of SQ-5 cells, which are derived from a human lung carcinoma. Actively growing cells were incubated for 16 h at 37 degrees C in medium containing 40 muM apigenin. The cells were then irradiated with X-rays and incubated with apigenin for a further 8 h. Radiosensitivity was assessed using a clonogenic assay. Apoptosis and necrosis were assessed using acridine orange/ethidium bromide double staining. Cells incubated with apigenin exhibited significantly greater radiosensitivity and apoptosis levels than cells not incubated with apigenin. Protein levels were measured by Western blotting. Incubation with apigenin increased protein expression of WAF1/p21 and decreased protein expression of Bcl-2. Furthermore, apigenin sensitized SQ-5 spheroids (cell aggregates growing in a three-dimensional structure that simulate the growth and microenvironmental conditions of in vivo tumors) to radiation. Thus, apigenin appears to be a promising radiosensitizing agent for use against human carcinomas.

Enhancement of cell death by TNF alpha-related apoptosis-inducing ligand (TRAIL) in human lung carcinoma A549 cells exposed to x rays under hypoxia.

Our previous study showed that ionizing radiation induced the expression of death receptor DR5 on the cell surface in tumor cell lines and that the death receptor of the TNF alpha-related apoptosis-inducing ligand TRAIL enhanced the apoptotic pathway (Hamasu et al., (2005) Journal of Radiation Research, 46:103-110). The present experiments were performed to examine whether treatment with TRAIL enhanced the cell killing in tumor cells exposed to ionizing radiation under hypoxia, since the presence of radioresistant cells in hypoxic regions of solid tumors is a serious problem in radiation therapy for tumors. When human lung carcinoma A549 cells were irradiated under normoxia and hypoxia, respectively, radiation-induced enhancement of expression of DR5 was observed under both conditions. Incubation in the presence of TRAIL enhanced the caspase-dependent and chymotrypsin-like-protease-dependent apoptotic cell death in A549 cells exposed to X rays. Furthermore, it was shown that treatment with TRAIL enhanced apoptotic cell death and loss of clonogenic ability in A549 cells exposed to X rays not only under normoxia but also under hypoxia, suggesting that combination treatment with TRAIL and X irradiation is effective for hypoxic tumor cells.

Potentiality of syringetin for preferential radiosensitization to cancer cells.

PURPOSE: To examine the enhancing effects of syringetin on the radiosensitivity of normal and cancer cells, and the related mechanism. MATERIALS AND METHODS: We used normal human lung and mouse fibroblasts as well as human lung and mouse cancer cells derived from the above normal fibroblasts. Cell radiosensitivity was measured using a colony formation assay. Apoptosis was analyzed with DAPI staining and Western blots. DNA lesions were analyzed with γH2AX immunofluorescent staining. RESULTS: The colony formation assay showed that syringetin enhanced radiosensitivity more effectively in cancer cells (H1299 and C3H/MCA clone 15) compared with normal cells (HFL-III and C3H/10T1/2). The radiosensitizing effect of syringetin was observed in mutated p53 and wild-type p53-transfected H1299 cells regardless of p53 status. Apoptosis was more frequently observed in X-ray-irradiated H1299 cells combined with syringetin compared with X-ray-only-treated cells. Enhanced apoptosis by syringetin was not observed in HFL-III cells. Western blot analysis showed that X-ray-induced Caspase-3 activation was enhanced by syringetin in H1299 cells. The number of X-ray-induced DNA double-strand breaks (DSB) measured by quantitative analysis of γH2AX foci was the same for H1299 cells treated with X-rays with or without syringetin. CONCLUSIONS: This study supports the hypothesis that syringetin enhances radiosensitivity more effectively in cancer cells than in normal cells through enhancement of the Caspase-3-mediated apoptosis pathway. Syringetin could be useful in the development of novel efficacious radiosensitizers.

[Antifungal Activity of Luliconazole Nail Solution on in vitro and in vivo Onychomycosis Model].

We evaluated luliconazole nail solution, originally generated formulation, for the topical treatment of onychomycosis by two infection models. First, a suspension of Trichophyton mentagrophytes was dropped onto the ventral layer of human nail plate and these nails were set in Franz diffusion cells. After 9-day culture, luliconazole nail solutions (1, 3, and 5%) were applied to the dorsal surface of the nails once a day for 7 days. After application, fungal viability was assessed by measuring the ATP contents of the samples. The dose-dependent efficacy was confirmed, with 3% and 5% luliconazole nail solutions producing significantly lower ATP levels at 7-day treatment. When 3% and 5% luliconazole nail solutions were evaluated in a rabbit model of onychomycosis, both concentrations completely inhibited the recovery of fungi on culture after 4-week treatment. We therefore think these results indicate that 5% luliconazole nail solution is sufficiently potent for treatment of onychomycosis.

[Distribution of Luliconazole in Nail Plate by In Vitro Permeation and Efficacy by Zone of Inhibition Test after Treatment of Luliconazole Nail Solution].

To clarify the character of luliconazole nail solution we have developed, we investigated luliconazole distribution and antifungal activity in nail plate. An in vitro permeation study which measured luliconazole concentration of sliced nail in the transverse direction after treatment of luliconazole nail solution was conducted to investigate for concentration dependency and the influences of nail thickness and treatment duration. When 0.2, 1, 3, 5, and 7.5% luliconazole nail solutions were used, luliconazole was detected in the all the layers of nail and there was a concentration gradient from the dorsal side to deep nail layers. The luliconazole concentration was almost same after 14-day treatment with 5% luliconazole nail solution when using nails of different thicknesses. And we confirmed that concentration of luliconazole into the nail was increased depending on the treatment duration. In zone of inhibition test after 14-day treatment, 5% luliconazole nail solution showed statistically high formation rate of zones of inhibition compared to 8% ciclopirox nail lacquer. Above all, these data suggested that 5% luliconazole nail solution has the potential to show high therapeutic effect for onychomycosis.

Comparison of Characteristics of Two Topical Therapeutic Agents for Onychomycosis.

Two topical therapeutic agents were approved in Japan from 2015 to 2016, adding new options for onychomycosis therapy in the clinical field. In order to confirm the differences of formulation properties and nail pharmacokinetics between 5% luliconazole solution and 10% efinaconazole solution, drug concentration and antifungal activity in the nail were measured after topical treatment using human nail plates. In the in vitro permeation study, concentration of each drug was measured in the transversely sliced nail after single treatment with the two topical therapeutic agents. The results showed that concentration of luliconazole is higher than that of efinaconazole at all nail layers, differing by 1.7-8.4 times at each measurement point. Next, we examined antifungal activities of each drug in sliced nail after 14-day topical treatment. Mean rates of formation of inhibition zones for 5% luliconazole solution and 10% efinaconazole solution were 71.0% and 12.6%, respectively, and were statistically different. These results show that the two topical therapeutic agents have different properties, and suggest that 5% luliconazole solution has good nail permeation and retention characteristics. Moreover, luliconazole was found to retain enough antifungal activity in the nail plate against Trichophyton spp. after treatment with the topical agent.

Preferential sensitization of tumor cells to radiation by heat shock protein 90 inhibitor geldanamycin.

The purpose of this study was to investigate the radiosensitizing effect of geldanamycin (GA), an inhibitor of heat shock protein 90, on tumour cells and normal cells. We tested the effect of a combination of GA and radiation on cell survival, PI3K/Akt-related proteins and apoptosis induction. GA sensitized tumour cells to radiation in preference to normal cells. In addition, a combination of radiation and GA abolished Akt activities and strongly enhanced the induction of apoptosis in tumour cells which depend on Akt protein activities for cell survival. The present data support the hypothesis that GA sensitizes tumour cells by modulating the balance among mitogenic, antiproliferative and apoptotic pathways. Targeting Hsp90 in tumour cells may lead to the development of new radiosensitizing strategies in radiotherapy.

Effects of hypoxic cell radiosensitizer doranidazole (PR-350) on the radioresponse of murine and human tumor cells in vitro and in vivo.

We have investigated the radiosensitizing effect of doranidazole, a hypoxic cells radiosensitizer, using SCCVII tumor cells of C3H mice and CFPAC-1 and MIA PaCa-2 human pancreatic tumor cells. The radiosensitivity of hypoxic SCCVII cells in vitro increased with 1 mM doranidazole by a factor of 1.34 and 1.68, when determined by clonogenic survival and micronucleus (MN) formation, respectively. The radiation-induced growth delay of SCCVII tumors was significantly enhanced and the TCD(50/120) was reduced by a factor of 1.33 when 200 mg/kg doranidazole was injected, i.v., 20 min prior to tumor irradiation. The in vivo-in vitro excision assay showed that radiosensitivity of SCCVII cells in vivo increased by a factor of 1.47 with 200 mg/kg doranidazole. The radiation-induced growth delay of CFPAC-1 xenografts in nude mice was significantly enhanced and the TCD(50/90) was reduced by a factor of 1.30 by 200 mg/kg doranidazole. On the other hand, 200 mg/kg of doranidazole exerted no influence on the radiation-induced growth delay in MIA PaCa-2 xenografts. The tumor oxygenation status, as determined with an oxygen sensitive needle probe and the immunohistological study using pimonidazole, indicated that MIA PaCa-2 tumors are better oxygenated than CFPAC-1 tumors. The relatively well-oxygenated status in MIA PaCa-2 tumor may account for the lack of radiosensitization by doranidazole. It is concluded that the magnitude of radiosensitization of tumors by doranidazole is dependent on the oxygenation status of the tumors and that doranidazole may be useful in increasing the response of hypoxic human pancreatic tumor to IORT.