Adenovirus as a Vector and Oncolytic Virus.

Adenoviral vectors, both oncolytic viruses and gene delivery vectors, are among the earliest approved and commercialised vectors for gene therapy. Adenoviruses have high cytotoxicity and immunogenicity. Therefore, lentiviruses or adeno-associated viruses as viral vectors and herpes simplex virus as an oncolytic virus have recently drawn attention. Thus, adenoviral vectors are often considered relatively obsolete. However, their high cargo limit and transduction efficiency are significant advantages over newer viral vectors. This review provides an overview of the new-generation adenoviral vectors. In addition, we describe the modification of the fiber knob region that enhances affinity of adenoviral vectors for cancer cells and the utilisation of cancer-cell-specific promoters to suppress expression of unwanted transgenes in non-malignant tissues.

Identification of HUHS190, a human naftopidil metabolite, as a novel anti-bladder cancer drug.

We carried out structure-activity relationship study on anti-cancer effects of naftopidil (1) and its metabolites, resulted in identification of 1-(4-hydroxy-2-methoxyphenyl)piperazin-1-yl)-3-(naphthalen-1-yloxy) propan-2-ol (2, HUHS190), a major human metabolite of 1, which exhibited the most selective toxicities between against normal and cancer cells (Table 1). 2 was more hydrophilic compared to 1, was enough to be prepared in high concentration solution of more than 100 µM in saline for an intravesical instillation drug. Moreover, serum concentration of 2 was comparable to that of 1, an oral preparation drug, after oral administration at 32 mg/kg (Fig. 3). Both of 1 and 2 showed broad-spectrum anti-cancer activities in vitro, for example, 1 and 2 showed inhibitory activity IC50 = 21.1 µM and 17.2 µM for DU145, human prostate cancer cells, respectively, and IC50 = 18.5 µM and 10.5 µM for T24 cells, human bladder cancer cells. In this study, we estimated anticancer effects of 2 in a bladder cancer model after intravesical administration similar to clinical cases. A single intravesical administration of 2 exhibited the most potent inhibitory activities among the clinical drugs for bladder cancers, BCG and Pirarubicin, without obvious side effects and toxicity (Fig. 4). Thus, HUHS190 (2) can be effective for patients after post-TURBT therapy of bladder cancer without side effects, unlike the currently available clinical drugs.

Adenosine deaminase inhibitor EHNA exhibits a potent anticancer effect against malignant pleural mesothelioma.

BACKGROUND/AIMS: Malignant pleural mesothelioma (MPM) is an aggressive malignant tumor and an effective therapy has been little provided as yet. The present study investigated the possibility for the adenosine deaminase (ADA) inhibitor EHNA as a target of MPM treatment. METHODS: MTT assay, TUNEL staining, monitoring of intracellular adenosine concentrations, and Western blotting were carried out in cultured human MPM cell lines without and with knocking-down ADA. The in vivo effect of EHNA was assessed in mice inoculated with NCI-H2052 MPM cells. RESULTS: EHNA induced apoptosis of human MPM cell lines in a concentration (0.01-1 mM)- and treatment time (24-48 h)-dependent manner, but such effect was not obtained with another ADA inhibitor pentostatin. EHNA increased intracellular adenosine concentrations in a treatment time (3-9 h)-dependent manner. EHNA-induced apoptosis of MPM cells was mimicked by knocking-down ADA, and the effect was neutralized by the adenosine kinase inhibitor ABT-702. EHNA clearly suppressed tumor growth in mice inoculated with NCI-H2052 MPM cells. CONCLUSION: The results of the present study show that EHNA induces apoptosis of MPM cells by increasing intracellular adenosine concentrations, to convert to AMP, and effectively prevents MPM cell proliferation. This suggests that EHNA may be useful for treatment of the tragic neoplasm MPM.

Newly synthesized anticancer drug HUHS1015 is effective on malignant pleural mesothelioma.

The newly synthesized naftopidil analogue HUHS1015 reduced cell viability in malignant pleural mesothelioma cell lines MSTO-211H, NCI-H28, NCI-H2052, and NCI-H2452, with the potential greater than that for the anticancer drugs paclitaxel or cisplatin at concentrations higher than 30 µM. HUHS1015 induced both necrosis and apoptosis of MSTO-211H and NCI-H2052 cells. HUHS1015 upregulated expression of mRNAs for Puma, Hrk, and Noxa in MSTO-211H and NCI-H2052 cells, suggesting HUHS1015-induced mitochondrial apoptosis. HUHS1015 clearly suppressed tumor growth in mice inoculated with NCI-H2052 cells. Taken together, the results of the present study indicate that HUHS1015 could be developed as an effective anticancer drug for treatment of malignant pleural mesothelioma.

Naftopidil is useful for the treatment of malignant pleural mesothelioma.

Naftopidil, an α1-adrenoceptor blocker, induced apoptosis of human malignant pleural mesothelioma NCI-H2052 cells. Naftopidil upregulated the expression of tumor necrosis factor-α (TNF-α) mRNA in these cells. Naftopidil, alternatively, increased FasL secretion from NCI-H2052 cells, without affecting the expression of FasL mRNA and protein, and activated caspase-3 and -8 in NCI-H2052 cells. Naftopidil drastically suppressed tumor growth in mice inoculated with these cells. The results of the present study indicate that naftopidil induces apoptosis of NCI-H2052 cells by upregulating the expression of TNF-α and stimulating the secretion of FasL, a ligand for the death receptor Fas, both to activate caspase-8 and the effector caspase-3, leading to the suppression of NCI-H2052 cell proliferation in vivo. This raises the possibility that naftopidil could be developed as an effective drug for the treatment of malignant pleural mesothelioma.

Crosstalk between PI3 kinase/PDK1/Akt/Rac1 and Ras/Raf/MEK/ERK pathways downstream PDGF receptor.

BACKGROUND/AIMS: Our earlier studies suggested crosstalk between IRS/PI3 kinase/PDK1/Akt/Rac1/ROCK and (Shc2/Grb2/SOS)/Ras/Raf/MEK/ERK pathways downstream PDGF-ßß receptor responsible for chemotaxis and proliferation of malignant mesothelioma cells. The present study was conducted to obtain evidence for this. METHODS: To assess activation of Akt, MEK, and ERK, Western blotting was carried out on MSTO-211H malignant mesothelioma cells using antibodies against phospho-Thr308-Akt, phopho-Ser473-Akt, Akt, phospho-MEK, MEK, phopho-ERK1/2, and ERK1/2. To knock-down Akt, PI3 kinase, PDK1, and Rac1, siRNAs silencing each-targeted gene were constructed and transfected into cells. To monitor Rac1 activity, FRET monitoring was carried out on living and fixed cells. RESULTS: ERK was activated under the basal conditions in MSTO-211H cells, and the activation was prevented by inhibitors for PI3 kinase, PDK1, Akt, and Rac1 or by knocking-down PI3 kinase, PDK1, Akt, and Rac1. Akt was also activated under the basal conditions, and the activation was suppressed by a MEK inhibitor and an ERK1/2 inhibitor. In the FRET analysis, Rac1 was activated under the basal conditions, and the activation was inhibited by a MEK inhibitor and an ERK1/2 inhibitor. CONCLUSION: The results of the present study show that ERK could be activated by PI3 kinase, PDK1, Akt, and Rac1 and that alternatively, Akt and Rac1 could be activated by MEK and ERK in MSTO-211H cells.

A3 adenosine receptor mediates apoptosis in in vitro RCC4-VHL human renal cancer cells by up-regulating AMID expression.

PURPOSE: Accumulating studies have shown that extracellular adenosine induces apoptosis in various cancer cells via diverse signaling pathways. We sought to understand adenosine induced apoptosis in human renal cancer cells and the underlying pathway. MATERIALS AND METHODS: RCC4-VHL (European Collection of Animal Cell Cultures, Salisbury, United Kingdom), ACHN (Cell Resource Center for Biomedical Research, Institute of Development, Aging and Cancer, Tohuku University, Aoba-ku, Sendai, Japan) and 786-O (ATCC®) human renal cancer cells were cultured. MTT assay, TUNEL staining, reverse transcriptase-polymerase chain reaction and Western blot were done in cells untransfected and transfected with siRNA silencing the A(3) adenosine receptor targeted gene or the AMID targeted gene. RESULTS: Adenosine induced apoptosis in all cell types used in a concentration (1 to 10 mM) dependent manner. A similar effect was obtained with the A(3) adenosine receptor agonist 2-Cl-IB-MECA. Adenosine induced RCC4-VHL cell death was inhibited by the A(3) adenosine receptor inhibitor MRS1191 or by knocking down A(3) adenosine receptor or AMID. Adenosine up-regulated the expression of AMID mRNA and protein in RCC4-VHL cells, which was suppressed by A(3) adenosine receptor knockdown. Moreover, adenosine promoted AMID translocation from cytosol to nucleus. CONCLUSIONS: Adenosine induces RCC4-VHL cell apoptosis by up-regulating AMID expression and accumulating AMID in the nucleus via A(3) adenosine receptor.

Potential tumor markers of renal cell carcinoma: α-enolase for postoperative follow up, and galectin-1 and galectin-3 for primary detection.

The diagnosis of renal cell carcinoma is currently based on imaging techniques, mainly because there is no blood marker available for its detection. Thus, there is still the need for the development of novel tumor markers. We examined plasma levels of eight proteins in 15 renal cell carcinoma patients before and after surgery, and in 51 healthy controls using enzyme-linked immunosorbent assay. Plasma levels of α-enolase, calnexin, galectin-1, galectin-3 and lectin mannose-binding 2 were significantly higher in renal cell carcinoma patients than in controls (P < 0.05). Among these proteins, the sensitivities for galectin-1 and galectin-3 were higher than those for calnexin and lectin mannose-binding 2 in the specificity range from 80% to 100%. A combined use of galectin-1 and galectin-3 showed 98% specificity and 47% sensitivity. In addition, the assays showed that plasma α-enolase levels decreased significantly 4 weeks after nephrectomy (P = 0.0034), and this tendency continued until 12 weeks after nephrectomy (P = 0.0156). These findings suggest that α-enolase could be used in the postoperative follow up of renal cell carcinoma patients, whereas the combined use of galectin-1 and galectin-3 might represent a useful tool for primary detection.

A(2a) adenosine receptor mediates HepG2 cell apoptosis by downregulating Bcl-X(L) expression and upregulating Bid expression.

Extracellular adenosine induced apoptosis in HepG2 cells, a human hepatoma cell line, by tuning apoptosis-mediator gene transcription. The present study aimed at identifying the responsible adenosine receptor and clarifying the signaling pathway underlying adenosine-induced HepG2 cell apoptosis. Adenosine and CGS21680, an A(2a) adenosine receptor agonist, induced HepG2 cell apoptosis, and the effect was inhibited by DMPX, an A(2a) adenosine receptor antagonist, or by knocking-down A(2a) adenosine receptors. Adenosine reduced expression of Bcl-X(L) mRNA and protein but otherwise increased expression of the Bid mRNA and protein in HepG2 cells, and those effects were also prevented by knocking-down A(2a) adenosine receptors. Adenosine caused disruption of mitochondrial membrane potentials and stimulated cytochrome c efflux from the mitochondria in HepG2 cells. Adenosine activated caspases-3 and -9 in HepG2 cells, which was significantly inhibited by knocking-down A(2a) adenosine receptors. The results of the present study indicate that extracellular adenosine downregulates Bcl-X(L) expression and upregulates Bid expression, thereby disrupting mitochondrial membrane potentials to allow cytochrome c efflux from the mitochondria, and then causing activation of caspase-9 and the effector caspase-3, as mediated via A(2a) adenosine receptors.

Sphingosine induces apoptosis in MKN-28 human gastric cancer cells in an SDK-dependent manner.

BACKGROUND/AIMS: Evidence has pointed to the role of sphingosine in cellular differentiation, cell growth, and apoptosis. The present study investigated sphingosine-induced apoptosis in human gastric cancer cells. METHODS: Well differentiated MKN-28 and poorly differentiated MKN-45 human gastric cancer cells were cultured. MTT assay, TUNEL staining, Western blotting, and assay of caspase-3, -8, and -9 activities were carried out in cells transfected with and without the siRNA to silence the protein kinase C (PKC)-δ-targeted gene. RESULTS: Sphingosine induced apoptosis in MKN-28 cells, with the potential much greater than for MKN-45 cells. Transfection with the siRNA to silence the PKC-δ-targeted gene (PKC-δ siRNA) into MKN-28 cells significantly reduced presence of sphingosine-dependent protein kinase (SDK) in association with reduced PKC-δ expression. Sphingosine-induced apoptosis in MKN-28 cells was prevented by transfecting with the PKC-δ siRNA. Sphingosine promoted SDK production from PKC-δ and increased phosphorylated 14-3-3 protein for MKN-28 cells, but such effects were not found with MKN-45 cells. Moreover, sphingosine perturbed mitochondrial membrane potentials and activated caspase-3 and caspase-9 in MKN-28 cells, which were also inhibited by transfecting with the PKC-δ siRNA. CONCLUSION: The results of the present study indicate that sphingosine induces apoptosis in well differentiated MKN-28 human gastric cancer cells by increasing SDK production from PKC-δ, to phosphorylate 14-3- 3 protein, thereby causing disruption of mitochondrial membrane potentials and activating caspase-9 followed by the effector caspase-3.

A(3) adenosine receptor mediates apoptosis in 5637 human bladder cancer cells by G(q) protein/PKC-dependent AIF upregulation.

BACKGROUND/AIMS: A(3) adenosine receptor mediates apoptosis in a variety of cancer cells via diverse signaling pathways. The present study was conducted to assess A(3) adenosine receptor-mediated apoptosis in human bladder cancer cell lines and to understand the underlying mechanism. METHODS: Human bladder cancer cell lines such as 253J, 5637, KK-47, TCCSUP, T24, and UMUC-3 cells were cultured. The siRNA to silence the A(3) adenosine receptor-targeted gene was constructed and transfected into cells. MTT assay, TUNEL staining, Western blotting, and real-time RT-PCR were carried out. RESULTS: For all the investigated cell types adenosine induced apoptosis in a concentration (0.01-10 mM)- and treatment time (24-48 h)-dependent manner. Adenosine-induced 5637 cell death was significantly inhibited by the A(3) adenosine receptor inhibitor MRS1191 or knocking-down A(3) adenosine receptor, and the A(3) adenosine receptor agonist 2-Cl-IB-MECA mimicked the adenosine effect. The adenosine effect was prevented by GF109203X, an inhibitor of protein kinase C (PKC), but it was not affected by forskolin, an activator of adenylate cyclase. Adenosine-induced 5637 cell death, alternatively, was not inhibited by the pan-caspase inhibitor Z-VAD. Adenosine upregulated expression of apoptosis-inducing factor (AIF), that is suppressed by knocking-down A(3) adenosine receptor, and accumulated AIF in the nucleus. CONCLUSION: The results of the present study show that adenosine induces 5637 cell apoptosis by upregulating AIF expression via an A(3) adenosine receptor-mediated G(q) protein/PKC pathway.

Adenosine induces apoptosis in SBC-3 human lung cancer cells through A(3) adenosine receptor-dependent AMID upregulation.

BACKGROUND/AIMS: We have shown that A(3) adenosine receptor mediates apoptosis in human lung cancer cells such as A549 cells, an epithelial adenocarcinoma cell line, and Lu-65 cells, a giant cell cancer cell line, via each different signaling pathway. AMID, a pro-apoptotic protein, induces caspase-independent apoptosis by accumulating in the nucleus. The present study investigated AMID-dependent apoptosis through A(3) adenosine receptor in SBC-3 cells, a human small cell lung cancer cell line. METHODS: MTT assay, TUNEL staining, flow cytometry using propidium iodide and annexin V-FITC, and Western blotting were carried out in SBC-3 cells transfected with and without the siRNA to silence the A(3) adenosine receptor-targeted gene or the AMID-targeted gene. RESULTS: Adenosine induced SBC-3 cell apoptosis in a concentration (0.01-10 mM) and treatment time (24-72 h)-dependent manner, and a similar effect was obtained with the A(3) adenosine receptor agonist 2-Cl-IB-MECA. Adenosine-induced SBC-3 cell death was inhibited by the A(3) adenosine receptor inhibitor MRS1191, knocking-down A(3) adenosine receptor, or knocking-down AMID. Adenosine upregulated expression of the AMID mRNA and protein in SBC-3 cells, that is suppressed by knocking-down A(3) adenosine receptor. In addition, adenosine increased nuclear AMID localization in concert with decreased cytosolic AMID localization. CONCLUSION: The results of the present study show that adenosine induces SBC-3 cell apoptosis by upregulating AMID expression and promoting AMID translocation into the nucleus via A(3) adenosine receptor.

PDGF-D/PDGF-ßß receptor-regulated chemotaxis of malignant mesothelioma cells.

BACKGROUND/AIMS: Our earlier study suggested that platelet-derived growth factor (PDGF)- ßß receptor regulates chemotaxis of human malignant mesothelioma cells such as MSTO-211H, NCIH-2052, NCIH-2452, and NCIH-28 cells, but not non-malignant Met5A cells. The present study was designed to gain further insight into the PDGF-ßß receptor signals underlying the chemotaxis. METHODS: PDGF-D secreted from cells, activation of Akt and ERK, and cell migration were monitored for cells with and without knocking-down PDGF-ßß receptor. RESULTS: FBS significantly stimulated PDGF-D secretion from malignant mesothelioma cells, but not Met5A cells. PDGF-D activated Akt and ERK in both the non-malignant and malignant cells. PDGF-D significantly facilitated migration of malignant mesothelioma cells, but not Met5A cells, with the extent varying among the cell types. The facilitatory action of PDGF-D was clearly prevented by knocking-down PDGF-ßß receptor or inhibitors of PI3 kinase, PDK1, Akt, Rac1, ROCK, and MEK. CONCLUSION: The results of the present study indicate that PDGF-D promotes malignant mesothelioma cell chemotaxis through PDGF-ßß receptor signaling pathways along a PI3 kinase/PDK1/Akt/Rac1/ROCK axis and relevant to ERK activation.

Mesothelioma cell proliferation through autocrine activation of PDGF-ßß receptor.

BACKGROUND/AIMS: Growth factors play a critical role in proliferation for a variety of cancer cells. The present study was conducted to understand the signaling cascades underlying PDGF-D/PDGF-ßß receptor-mediated proliferation of mesothelioma cells. METHODS: Cell growth and cell cycle were analyzed in human non-malignant Met5A cells and malignant mesothelioma cells such as MSTO-211H, NCI-H28, NCI-H2052, and NCI-H2452 cells. RESULTS: Growth of all the cells used here was not affected by PDGF-D, regardless of concentrations (1-30 ng/ml) or treatment time (48-72 h). Spontaneous growth of those cells was significantly inhibited by knocking-down PDGFD or PDGF-ßß receptor, without affecting cell cycling. The cell growth was significantly inhibited by the Akt inhibitor MK2206 and the ROCK inhibitor Y27632 for all the cell types, by the PDK1 inhibitor BX912 for NCI-H28 cells alone, and by the Rac1 inhibitor NSC23766 for NCI-H2052 cells alone, while the PI3 kinase inhibitor wortmannin had no effect. The cell growth, alternatively, was significantly attenuated by MAP kinase kinase inhibitor PD98059 or the ERK1/2 inhibitor FR180204 for all the cell types. CONCLUSION: The results of the present study show that PDGF-D promotes mesothelioma cell proliferation by targeting ROCK or MAP kinase through autocrine activation of PDGF-ßß receptor.

Apoptosis-related gene transcription in human A549 lung cancer cells via A(3) adenosine receptor.

BACKGROUND/AIMS: Extracellular adenosine induces apoptosis in a variety of cancer cells via diverse signaling pathways. The present study investigated the mechanism underlying adenosine-induced apoptosis in A549 human lung cancer cells. METHODS: MTT assay, TUNEL staining, flow cytometry using propidium iodide and annexin V-FITC, real-time RTPCR, Western blotting, monitoring of mitochondrial membrane potentials, and assay of caspase-3, -8, and -9 activities were carried out in A549 cells, and the siRNA to silence the A(3) adenosine receptor-targeted gene was constructed. RESULTS: Extracellular adenosine induces A549 cell apoptosis in a concentration (0.01-10 mM)-dependent manner, and the effect was inhibited by the A3 adenosine receptor inhibitor MRS1191 or knocking-down A(3) adenosine receptor. Like adenosine, the A(3) adenosine receptor agonist 2-Cl-IB-MECA also induced A549 cell apoptosis. Adenosine increased expression of mRNAs for Puma, Bax, and Bad, disrupted mitochondrial membrane potentials, and activated caspase-3 and -9 in A549 cells, and those adenosine effects were also suppressed by knocking-down A3 adenosine receptor. CONCLUSION: Adenosine induces A549 cell apoptosis by upregulating expression of Bax, Bad, and Puma, to disrupt mitochondrial membrane potentials and to activate caspase-9 followed by the effector caspase-3, via A(3) adenosine receptor.

Sphingosine suppresses mesothelioma cell proliferation by inhibiting PKC-δ and inducing cell cycle arrest at the G(0)/G(1) phase.

BACKGROUND/AIMS: Sphingosine regulates cellular differentiation, cell growth, and apoptosis. The present study aimed at understanding sphingosine-regulated mesothelioma cell proliferation. METHODS: Human malignant mesothelioma cells such as NCI-H28, NCI-H2052, NCI-H2452, and MSTO-211H cells were cultured. The siRNA to silence the protein kinase C (PKC)-δ-targeted gene was constructed and transfected into cells. MTT assay, cell cycle analysis using a flow cytometry, and cell-free PKC-δ assay were carried out. RESULTS: For all the cell types sphingosine inhibited cell growth in a concentration (1-100 µM)-dependent manner. The sphingosine effect was not prevented by rottlerin, an inhibitor of protein kinase C-δ (PKC-δ); conversely, rottlerin further enhanced the sphingosine effect or rottlerin suppressed mesothelioma cell growth without sphingosine. In the cell-free PKC assay, sphingosine attenuated PKC-δ activity. Knocking-down PKC-δ induced cell cycle arrest at the G0/G1 phase and inhibited cell growth. CONCLUSION: The results of the present study show that sphingosine suppressed mesothelioma cell proliferation by inhibiting PKC-δ, to induce cell cycle arrest at the G0/G1 phase.

AMP converted from intracellularly transported adenosine upregulates p53 expression to induce malignant pleural mesothelioma cell apoptosis.

BACKGROUND/AIMS: The present study investigated adenosine-induced apoptosis in human malignant pleural mesothelioma cells. METHODS: MTT assay, TUNEL staining, flow cytometry using propidium iodide and annexin V-FITC, real-time RT-PCR, Western blotting, and assay of caspase-3, -8, and -9 activities were carried out using malignant pleural mesothelioma cell lines such as NCI-H28, NCI-H2052, NCI-H2452, and MSTO-211H cells, and p53 or A(3) adenosine receptor was knocked-down by transfecting each siRNA into cells. RESULTS: Adenosine induced apoptosis in all the malignant pleural mesothelioma cells used here, independently of caspase activation. The adenosine effect was prevented by the adenosine transporter inhibitor dipyridamole, the adenosine kinase inhibitor ABT-702, or the A(3) adenosine receptor inhibitor MRS1191. Adenosine upregulated expression of the p53 mRNA and protein, that is abolished by ABT-702, but not by knocking-down A(3) adenosine receptor. Adenosine-induced apoptosis in NCI-H28 cells was significantly inhibited by knocking-down p53 and in part by knocking-down A(3) adenosine receptor. CONCLUSION: The results of the present study show that AMP converted from intracellularly transported adenosine upregulates p53 expression to induce caspase-independent apoptosis in malignant pleural mesothelioma cells and that A(3) adenosine receptor also participates partially in the apoptosis by the different mechanism.

A3 adenosine receptor-mediated p53-dependent apoptosis in Lu-65 human lung cancer cells.

BACKGROUND/AIMS: A(3) adenosine receptor mediates apoptosis in cancer cells via diverse signaling pathways. The present study examined A(3) adenosine receptor-mediated apoptosis in Lu-65 cells, a human giant cell lung carcinoma cell line. METHODS: MTT assay, TUNEL staining, real-time RT-PCR, Western blotting, and assay of caspase-3, -8, and -9 activities were carried out in Lu-65 cells, and A(3) adenosine receptor or p53 was knocked-down by transfecting each siRNA into cells. RESULTS: Extracellular adenosine induces Lu-65 cell apoptosis in a concentration (0.01-10 mM)-dependent manner, and the effect was inhibited by the A(3) adenosine receptor inhibitor MRS1191 or by knocking-down A(3) adenosine receptor or p53. Like adenosine, the A(3) adenosine receptor agonist 2-Cl-IB-MECA also induced Lu-65 cell apoptosis. Adenosine upregulated expression of p53 and Noxa mRNAs and activated caspase-3 and -9, but not caspase-8. Those adenosine effects were still inhibited by knocking-down A(3) adenosine receptor or p53. CONCLUSION: The results of the present study show that adenosine upregulates p53 expression via A(3) adenosine receptor, to promote p53-dependent Noxa gene transcription, causing activation of caspase-9 and the effector caspase-3 to induce Lu-65 cell apoptosis.

Antitumor action of α(1)-adrenoceptor blockers on human bladder, prostate and renal cancer cells.

The present study investigated the antitumor action of α(1)-adrenoceptor blockers on human bladder, prostate and renal cancer cells. For bladder cancer cell lines used here such as 253J, 5637, KK-47, T24 and UM-UC-3 cells, prazosin, a selective α(1)-adrenoceptor blocker, reduced cell viability at concentrations more than 30 µmol/l. Likewise, naftopidil, a blocker of α(1A)- and α(1D)-adrenoceptors, reduced cell viability for all the bladder cancer cells used here in a concentration (10-100 µmol/l)-dependent manner, with a much greater advantage than prazosin. Naftopidil also reduced cell viability for human prostate cancer cell lines such as DU145, LNCap and PC-3 cells and ACHN human renal cancer cells, with a much higher potential than prazosin. Thus, the results of the present study suggest that naftopidil could be a beneficial antitumor drug for the treatment of urological cancers.

Nicotinamide Mononucleotide Is Safely Metabolized and Significantly Reduces Blood Triglyceride Levels in Healthy Individuals.

An increase in nicotinamide adenine dinucleotide (NAD+) levels alleviates age-related disease progression and promotes healthy life expectancy. Several studies have demonstrated that NAD+ levels can be efficiently replenished via nicotinamide mononucleotide (NMN) intake; additionally, the safety of its oral ingestion has been confirmed in recent clinical trials. However, the efficacy and safety of intravenous NMN administration in humans remain unclear. Therefore, we verified its safety in 10 healthy volunteers. Intravenous administration of NMN did not affect electrocardiograms, pulse, and blood pressure, nor did it affect metabolic markers in the liver, heart, pancreas, and kidneys. These results indicate that intravenous NMN administration is safe and beneficial in humans. Furthermore, NMN administration significantly increased blood NAD+ levels without damaging blood cells and significantly reduced blood triglyceride (TG) levels. These findings imply that intravenous administration of NMN may lead to the prevention and treatment of diseases associated with increased TG levels, such as fatty liver and diabetes.