Inhibition of HIF-1α by the anticancer drug TAS106 enhances X-ray-induced apoptosis in vitro and in vivo

In a previous study, we showed that a novel anticancer drug, 1-(3-C-ethynyl-β-D-ribo-pentofuranosyl)cytosine (TAS106, ECyd) increased the antitumour efficacy of X-irradiation. However, its effects on hypoxic cells in tumours remain unclarified. Here, we show that TAS106 enhances the induction of apoptosis in X-irradiated human gastric adenocarcinoma MKN45 and MKN28 cells under hypoxia in vitro. At the same time, the accumulation of HIF-1α observed under hypoxia was shown to be decreased to the level of normoxia in the presence of 0.1 μM TAS106. To study the function of HIF-1α protein in apoptosis of hypoxic cells, we employed an HIF-1α reductive approach using its specific antisense oligodeoxynucleotide. The reduction of HIF-1α gene expression dramatically enhanced X-ray-induced apoptosis in hypoxic cells. In in vivo experiments in which MKN45 cells were transplanted into severe combined immunodeficient (SCID) mice, TAS106 (0.5 mg kg−1) suppressed HIF-1α expression and subsequently reduced the area of the hypoxic region in the tumour and enhanced the induction of apoptosis in the hypoxic region when combined with 2 Gy of X-irradiation. These results suggest the possibility that TAS106 acts as a potent radiosensitiser through the inhibition of HIF-1α expression and can be a useful agent against radiotherapy-resistant hypoxic cells in solid tumours

BAAV Transcytosis Requires an Interaction with β-1-4 Linked- Glucosamine and gp96

Cell surface carbohydrates play an important role in virus entry and intracellular trafficking. Bovine Adeno-Associated Virus (BAAV) uses plasma membrane gangliosides for transduction and infection. In addition, independent of the infectious pathway, BAAV also has the ability to pass through barrier epithelia and endothelia using a transcytosis pathway dependent upon the presence of cell surface carbohydrates. Thus, in order to better define the carbohydrate interactions that are necessary for BAAV infection or transcytosis, a glycan microarray composed of both natural and synthetic carbohydrates was probed with HA-tagged BAAV particles. This identified chitotriose, a trimer of β-1-4-linked N-acetyl glucosamine, as having an interaction with BAAV. Competition experiments showed that the BAAV interaction with this carbohydrate is not necessary for infection but is instead important in the transcytosis pathway. The β-1-4-linked N-acetyl glucosamine modification has been reported on gp96, a glycoprotein involved in the transcytosis of bacteria and toxins. Significantly, immunoprecipitation and competition experiments with an anti-gp96 antibody and a soluble form of gp96, respectively, showed this glycoprotein can also interact with BAAV to serve as a receptor for its transcytosis

DNA methylation status of REIC/Dkk-3 gene in human malignancies

The REIC (reduced expression in immortalized cells)/Dkk-3 is down-regulated in various cancers and considered to be a tumor suppressor gene. REIC/Dkk-3 mRNA has two isoforms (type-a,b). REIC type-a mRNA has shown to be a major transcript in various cancer cells, and its promoter activity was much stronger than that of type-b. In this study, we examined the methylation status of REIC/Dkk-3 type-a in a broad range of human malignancies. We examined REIC/Dkk-3 type-a methylation in breast cancers, non-small-cell lung cancers, gastric cancers, colorectal cancers, and malignant pleural mesotheliomas using a quantitative combined bisulfite restriction analysis assay and bisulfate sequencing. REIC/Dkk-3 type-a and type-b expression was examined using reverse transcriptional PCR. The relationships between the methylation and clinicopathological factors were analyzed. The rate of REIC/Dkk-3 type-a methylation ranged from 26.2 to 50.0% in the various primary tumors that were examined. REIC/Dkk-3 type-a methylation in breast cancer cells was significantly heavier than that in the other cell lines that we tested. REIC/Dkk-3 type-a methylation was inversely correlated with REIC/Dkk-3 type-a expression. There was a correlation between REIC/Dkk-3 type-a and type-b mRNA expression. REIC/Dkk-3 type-a expression was restored in MDA-MB-231 cells using 5-aza-2'-deoxycytidine treatment. We found that estrogen receptor-positive breast cancers were significantly more common among the methylated group than among the non-methylated group. REIC/Dkk-3 type-a methylation was frequently detected in a broad range of cancers and appeared to play a key role in silencing REIC/Dkk-3 type-a expression in these malignancies

Potent antitumor effects of combined therapy with a telomerase-specific, replication-competent adenovirus (OBP-301) and IL-2 in a mouse model of renal cell carcinoma

OBP-301 (a telomerase-specific, replication-competent adenovirus with hTERT promoter) was constructed in a previous study and it showed a strong anticancer effect by inducing cell lysis in human lung and prostate cancer cells. This study investigated the effectiveness of a combination therapy of OBP-301 and interleukin-2 (IL-2) in a mouse model of renal cell carcinoma (RCC). The cell-killing effect of OBP-301 was confirmed in vitro in the RENCA cancer cells. In in vivo experiment, luciferase-expressing RENCA cells were implanted in the left kidney and lung of BALB/c mice to prepare the RCC metastatic model. The animals were randomly divided into four treatment groups: PBS, IL-2 alone, OBP-301 alone and the combination. The analyses of orthotopic tumor weight, lung metastasis and luciferin-stained tumor images 14 days after each treatment showed significant tumor growth inhibition in the combination group in comparison with that in the OBP-301- or IL-2-treated groups. In addition, the percentage of regulatory T-cells (Tregs) in the combination group was significantly suppressed in comparison with that in the PBS and single-agent treatment groups. The outcomes of this study suggest that tumor-specific oncolytic immunovirotherapy may become an attractive strategy for the treatment of human RCC

The Response of the Prostate to Circulating Cholesterol: Activating Transcription Factor 3 (ATF3) as a Prominent Node in a Cholesterol-Sensing Network

Elevated circulating cholesterol is a systemic risk factor for cardiovascular disease and metabolic syndrome, however the manner in which the normal prostate responds to variations in cholesterol levels is poorly understood. In this study we addressed the molecular and cellular effects of elevated and suppressed levels of circulating cholesterol on the normal prostate. Integrated bioinformatic analysis was performed using DNA microarray data from two experimental formats: (1) ventral prostate from male mice with chronically elevated circulating cholesterol and (2) human prostate cells exposed acutely to cholesterol depletion. A cholesterol-sensitive gene expression network was constructed from these data and the transcription factor ATF3 was identified as a prominent node in the network. Validation experiments confirmed that elevated cholesterol reduced ATF3 expression and enhanced proliferation of prostate cells, while cholesterol depletion increased ATF3 levels and inhibited proliferation. Cholesterol reduction in vivo alleviated dense lymphomononuclear infiltrates in the periprostatic adipose tissue, which were closely associated with nerve tracts and blood vessels. These findings open new perspectives on the role of cholesterol in prostate health, and provide a novel role for ATF3, and associated proteins within a large signaling network, as a cholesterol-sensing mechanism

RNA interference approaches for treatment of HIV-1 infection

HIV/AIDS is a chronic and debilitating disease that cannot be cured with current antiretroviral drugs. While combinatorial antiretroviral therapy (cART) can potently suppress HIV-1 replication and delay the onset of AIDS, viral mutagenesis often leads to viral escape from multiple drugs. In addition to the pharmacological agents that comprise cART drug cocktails, new biological therapeutics are reaching the clinic. These include gene-based therapies that utilize RNA interference (RNAi) to silence the expression of viral or host mRNA targets that are required for HIV-1 infection and/or replication. RNAi allows sequence-specific design to compensate for viral mutants and natural variants, thereby drastically expanding the number of therapeutic targets beyond the capabilities of cART. Recent advances in clinical and preclinical studies have demonstrated the promise of RNAi therapeutics, reinforcing the concept that RNAi-based agents might offer a safe, effective, and more durable approach for the treatment of HIV/AIDS. Nevertheless, there are challenges that must be overcome in order for RNAi therapeutics to reach their clinical potential. These include the refinement of strategies for delivery and to reduce the risk of mutational escape. In this review, we provide an overview of RNAi-based therapies for HIV-1, examine a variety of combinatorial RNAi strategies, and discuss approaches for ex vivo delivery and in vivo delivery

Suppressive effects of liquid crystal compounds on the growth of U937 human leukemic monocyte lymphoma cells.

BACKGROUND: \nThe aim of this study was to evaluate the biological and pharmaceutical activities of 14 amphiphilic liquid-crystalline compounds (LCs), i.e, phenylpyrimidine derivatives possessing D-glucamine and cyanobiphenyl derivatives with a terminal hydroxyl unit.\nRESULTS: \nThe cytotoxic properties of the LCs on the cell growth, cell cycle distribution, and cell signaling pathway of U937 human leukemic monocyte lymphoma cells were assessed by flow cytometry and western blot analysis. Some LCs showed cytostatic effects, suppressing cell growth via S-phase arrest and without apoptosis in U937 cells. To investigate the mechanisms of the LC-induced S-phase arrest, proteins relevant to cell cycle regulation were investigated by western blot analysis. The rate of LC-induced S-phase arrest was congruent with the decreased expression of MCM2, cyclin A, cyclin B, CDK2, phospho-CDK1 and Cdc25C. Observed changes in cell cycle distribution by LC treated might be caused by insufficient preparation for G2/M transition. Considering the structure of the LCs, the rod-like molecules displaying cytotoxicity against U937 cells possessed flexible spacers with no bulky polar group attached via the flexible spacer.\nCONCLUSIONS: \nOur results revealed that some LCs showed cytotoxic properties against non-solid type tumor human leukemic cells via LC-induced S-phase arrest and decreasing expression of several cell cycle related proteins