Shen-Jeu Won

Xanthone Derivatives as Potential Anti‐cancer Drugs

Xanthone derivatives have been shown to be potent inhibitors of tumour growth. Oxygenated xanthones and [3‐(dialkylamino)‐2‐hydroxypropoxy]xanthones have been prepared and tested for in‐vitro inhibition of human PLC/PRF/5, KB and 212 cells.

Effects of tramadol on T lymphocyte proliferation and natural killer cell activity in rats with sciatic constriction injury

&NA; We investigated the effects of acute and chronic tramadol treatment on T lymphocyte function and natural killer (NK) cell activity in rats receiving chronic constriction injury (CCI) of the sciatic nerve. T lymphocyte function was evaluated based on concanavalin‐A (ConA)‐ and phytohemagglutinin (PHA)‐induced splenocyte proliferation. NK cell activity was measured by lactic acid dehydrogenase release assay. The effects of tramadol on thermal hyperalgesia were also assessed by measuring paw withdrawal latency (PWL) in the rats. PWL was dose‐dependently reversed by tramadol after acute treatment (single subcutaneous injection) with 10, 20, and 30 mg/kg, respectively. There was no significant change among acute treatment groups in NK cell activity, whereas splenocyte proliferation induced by ConA and PHA was significantly suppressed starting from a dose of 20 mg/kg. The reversal of the thermal hyperalgesia persisted throughout a period of chronic tramadol treatment of 40 and 80 mg/kg per day, respectively, with continuous subcutaneous infusion for 7 days at a uniform rate via osmotic minipumps. No modulation of NK cell activity was found in either dose group. However, the activity of splenocyte proliferation was decreased in the 80 mg/kg per day group when compared with the saline and 40 mg/kg per day groups. These data suggest that tramadol treatment has an immunological profile different from pure &mgr;‐opioid agonists like morphine, which is known to suppress both NK cell activity and T lymphocyte proliferation at a subanalgesic dose in CCI rats. Considering analgesic and immunosuppressive effects, tramadol treatment may be a better choice than morphine for treatment of chronic neuropathic pain, particularly in patients with compromised immunity.

γ‐Pyrone Compounds as Potential Anti‐cancer Drugs

Abstract— The γ‐pyrones, artomunoxanthotrione epoxide, cyclocommunol, cyclomulberrin, and cyclocommunin exhibited potent inhibition of human PLC/PRF/5 and KB cells in‐vitro. Dihydroisocycloartomunin showed significant and potent inhibition of human PLC/PRF/5 and KB cells in‐vitro, respectively. Cyclomorusin, dihydrocycloartomunin and artomunoxanthone showed significant inhibition of KB cells in‐vitro. Based on the above finding and the reported antileukaemic activity of xanthone psorospermin, a series of natural γ‐pyrones was prepared and the inhibition of human PLC/PRF/5 and KB cells in‐vitro was measured. Structure‐activity analysis indicated the epoxide group substituted at 3‐hydroxyl and 2,6‐; 3,6‐; and 3,5‐dihydroxyl xanthone enhanced the anti‐tumour activity. The epoxide group substituted at the 6‐hydroxyl group of 1,6‐dihydroxyxanthone did not show anti‐tumour activity.

Morusin induces apoptosis and suppresses NF-κB activity in human colorectal cancer HT-29 cells

Morusin is a pure compound isolated from root bark of Morusaustralis (Moraceae). In this study, we demonstrated that morusin significantly inhibited the growth and clonogenicity of human colorectal cancer HT-29 cells. Apoptosis induced by morusin was characterized by accumulation of cells at the sub-G(1) phase, fragmentation of DNA, and condensation of chromatin. Morusin also inhibited the phosphorylation of IKK-alpha, IKK-beta and IkappaB-alpha, increased expression of IkappaB-alpha, and suppressed nuclear translocation of NF-kappaB and its DNA binding activity. Dephosphorylation of NF-kappaB upstream regulators PI3K, Akt and PDK1 was also displayed. In addition, activation of caspase-8, change of mitochondrial membrane potential, release of cytochrome c and Smac/DIABLO, and activation of caspase-9 and -3 were observed at the early time point. Downregulation in the expression of Ku70 and XIAP was exhibited afterward. Caspase-8 or wide-ranging caspase inhibitor suppressed morusin-induced apoptosis. Therefore, the antitumor mechanism of morusin in HT-29 cells may be via activation of caspases and inhibition of NF-kappaB.

Caspase-8 acts as a key upstream executor of mitochondria during justicidin A-induced apoptosis in human hepatoma cells

Justicia procumbens is a traditional Taiwanese herbal remedy used to treat fever, pain, and cancer. Justicidin A, isolated from Justicia procumbens, has been reported to suppress in vitro growth of several tumor cell lines as well as hepatoma cells. In this study, justicidin A activated caspase‐8 to increase tBid, disrupted mitochondrial membrane potential (Δψ m), and caused the release of cytochrome c and Smac/DIABLO in Hep 3B and Hep G2 cells. Justicidin A also reduced Bcl‐xL and increased Bax and Bak in mitochondria. Caspase‐8 inhibitor (Z‐IETD) attenuated the justicidin A‐induced disruption of Δψ m. Growth of Hep 3B implanted in NOD‐SCID mice was suppressed significantly by oral justicidin A (20 mg/kg/day). These results indicate that justicidin A‐induced apoptosis in these cells proceeds via caspase‐8 and is followed by mitochondrial disruption. Supplementary materials are available at http://myweb.ncku.edu.tw/~a725/.

Effects of morphine on immune response in rats with sciatic constriction injury

&NA; We investigated the effects of acute and of chronic morphine treatment on T‐lymphocyte function and natural killer (NK) cell activity in rats receiving chronic constriction injury (CCI) of the sciatic nerve. T‐Lymphocyte function was evaluated based on concanavalin‐A (ConA)‐ and phytohemagglutinin (PHA)‐induced splenocyte proliferation. The effects of morphine on thermal hyperalgesia were also assessed by measuring paw withdrawal latency (PWL) in rats. All of the rats that received CCI developed thermal hyperalgesia while sham‐operated rats did not. Thermal hyperalgesia was dose‐dependently reversed after acute (single injection) and after chronic (daily injection for 7 days) administration of morphine but persisted in saline‐treated CCI rats. There was no significant difference between sham and saline‐treated CCI groups in splenocyte proliferation and NK cell activity. NK cell activity and splenocyte proliferation induced by ConA and PHA were significantly suppressed by acute morphine treatment in a dose‐dependent manner. The reversal of the thermal hyperalgesia persisted throughout the period of chronic morphine treatment. No tolerance to the suppression of NK cell activity and splenocyte proliferation was observed after chronic morphine treatment. These data suggest that both acute and chronic morphine treatment can cause a dose‐dependent reversal of thermal hyperalgesia and inhibition of NK cell activity and splenocyte proliferation in rats with sciatic CCI, without concomitant development of tolerance. Opioid therapy for chronic neuropathic pain should be used cautiously, especially in immune‐compromised cases.

Synthesis and cytotoxic effect of 1,3-dihydroxy-9,10-anthraquinone derivatives

1,3-Dihydroxy-9,10-anthraquinone (4) was reacted with epichlorohydrin or 1,omega-dibromo-alkane to yield 1-hydroxy-3-(2,3-epoxypropoxy)-9,10-anthraquinone (5) and 1-hydroxy-3-(3-chloro-2-hydroxypropoxy)-9,10-anthraquinone (6) or 1-hydroxy-3-(omega-bromoalkoxy)-9,10-anthraquinone. Ring-opening of the epoxide (5) or 1-hydroxy-3-(omega-bromoalkoxy)-9,10-anthraquinones with appropriate amines, afforded various 1-hydroxy-3-(3-alkylamino-2-hydroxypropoxy)-9,10-anthraquinones. The synthetic compounds were tested in vitro inhibition of human T-24, Hep 3B, Hep G2, SiHa, HT-3, PLC/PRF/5 and 212 cells. Almost all compounds showed significant inhibitory activity against several different cancer cell lines. Structure-activity analysis indicated epoxidation of the hydroxyanthraquinone increased cytotoxicity against tumour cells, but ring-opening of the epoxide group with amine did not enhance the cytotoxic activity. The phosphatidylserine (PS) externalization and DNA fragmentation in SiHa cells were significantly observed after 48 h incubation with selected compound 19. The results show that 19 cause cell death by apoptosis.

Formosanin C‐induced apoptosis requires activation of caspase‐2 and change of mitochondrial membrane potential

Formosanin C is a pure compound isolated from Paris formosana Hayata (Liliaceae). The antitumor efficacy of formosanin C has been observed in cultured cells and animal systems. However, the molecular mechanisms of formosanin C remain unknown. The results of the present study indicate that formosanin C induced apoptosis of HT‐29 cells characterized by exposure of phosphatidylserine, accumulation of cells at the sub‐G1 phase, fragmentation of DNA, and change of nuclear morphology in a time‐ and dose‐related manner. The apoptotic signaling cascades may proceed via proteolytic activation of caspase‐2, change of mitochondrial membrane potential (Δψm), release of cytochrome c and second mitochondria‐derived activator of caspase/direct IAP binding protein with low pI (Smac/DIABLO), activation of caspase‐9 and ‐3, and cleavage of poly(ADP‐ribose) polymerase (PARP). Increase in apoptosis‐inducing factor and endonuclease G expressions in nuclei, and increase in Bax and Bak expressions and decrease in Bcl‐XL expression on mitochondria were also observed in formosanin C‐treated HT‐29 cells. Attenuation of formosanin C‐induced change of Δψm by caspase‐2 inhibitor (Z‐VDVAC) implies that caspase‐2 acts upstream of the mitochondria. Blockage of formosanin C‐induced apoptotic process by using either permeability transition pore inhibitor (cyclosporine A) or caspase‐9 inhibitor (Z‐LEHD) demonstrates the necessity of mitochondria and caspase‐9 in formosanin C‐induced apoptosis of HT‐29 cells. Taken together, the apoptotic mechanism of formosanin C in human colorectal cancer HT‐29 cells involves activation of caspase‐2 and the dysfunction of mitochondria. (Cancer Sci 2009; 100: 503–513)

Mechanisms and sites of pyrogenic action exerted by staphylococcal enterotoxin A in rabbits.

The febrile responses induced by i.v. administrations of staphylococcal enterotoxin A (SEA) was mimicked by direct injection of SEA into the organum vasculosum laminae terminalis (OVLT) in unanesthetized rabbits. Compared with the febrile responses induced by i.v. injection of SEA, the OVLT route of injection required a much lower dose of SEA to produce a similar fever. Furthermore, the fever induced by intra-OVLT or i.v. injection of SEA was significantly attenuated by pretreatment with intra-OVLT injection of anisomycin (a protein synthesis inhibitor), indomethacin or diclofenac (inhibitors of cyclo-oxygenase (COX)), and aminoguanidine or dexamethasone (inhibitors of inducible nitric oxide synthase (iNOS)). These results suggest that COX or iNOS pathway in the OVLT mediate the SEA-induced fever in rabbits.

Staphylococcal enterotoxin A acts through nitric oxide synthase mechanisms in human peripheral blood mononuclear cells to stimulate synthesis of pyrogenic cytokines.

ABSTRACT The pyrogenic response to supernatant fluids obtained from human peripheral blood mononuclear cells (PBMC) stimulated with staphylococcal enterotoxin A (SEA) was characteristic of a response to an endogenous pyrogen in that it was brief and monophasic and was destroyed by heating supernatant fluids at 70°C for 30 min. The febrile responses were in parallel with the levels of interleukin-1 (IL-1), tumor necrosis factor (TNF), interferon-γ (IFN-γ), IL-2, and IL-6 in supernatant fluids obtained from PBMC treated with SEA. Both the pyrogenicity and the levels of IL-1, TNF, IFN-γ, IL-2, and IL-6 in supernatant fluids started to rise at 6 to 18 h and reached their peak levels at 24 to 96 h after SEA incubation. Both the fever and the increased levels of IL-1, TNF, IFN-γ, IL-2, and IL-6 in supernatant fluids obtained from the SEA-stimulated PBMC were decreased by incubating SEA-PBMC with anisomycin (a protein synthesis inhibitor), aminoguanidine (an inhibitor of inducible nitric oxide synthase [NOS]), or dexamethasone (an inhibitor of NOS). The febrile response to supernatant fluids obtained from the SEA-stimulated PBMC was attenuated by adding either anti-IL-1β, anti-TNF-α, or anti-IFN-γ monoclonal antibody (MAb) to supernatant fluids. The antipyretic effects exerted by anti-IL-1β MAb were greater than those exerted by anti-TNF-α or anti-IFN-γ MAb. The data suggest that SEA acts through the NOS mechanisms in PBMC to stimulate synthesis of pyrogenic cytokines (in particular, the IL-1β).