UCN-01 (7-hydroxy-staurosporine) is a potent and selective inhibitor of protein kinase C (PKC), one of several protein kinases examined. UCN-01 itself was shown to exhibit antitumor activity in vitro and in vivo in oncogene-activated human and murine tumor cell lines. Since the mechanism(s) of action of UCN-01 is thought to be different from those of alkylating agents, including mitomycin C (MMC), we tested the combined effect of UCN-01 with MMC on human epidermoid carcinoma A431 cells. UCN-01 potentiated the antiproliferative activity of MMC and yet it did not affect the growth of the cells in vitro. However, other nonselective protein kinase inhibitors, such as staurosporine, K-252a, KT6124 (a derivative of K-252a) and H7, did not enhance the activity of MMC. Isobologram analysis revealed that the interaction of UCN-01 with MMC was synergistic in its antiproliferative activity. A DNA histogram of A431 cells treated with both UCN-01 and MMC showed a block in the cell cycle at the G1/S phase. However, a histogram of cells treated with UCN-01 or MMC alone showed a G1 or a G2M block, respectively. The combined effect of UCN-01 with MMC was further examined in vivo in xenografted A431 cells in nude mice. The combination of both drugs in a single i.v. injection exhibited greater antitumor activity than MMC and UCN-01 alone (P < 0.01). This synergistic antitumor effect was also confirmed in two other solid tumor cell lines, i.e. human xenografted colon carcinoma Co-3 and murine sarcoma 180. The same was observed in the i.v.-inoculated P388 leukemia model, in which we saw an increased lifespan of mice when UCN-01 was combined with MMC. These results suggests the feasibility of using UCN-01 in clinical oncology, especially in combination with alkylating agents such as MMC. In addition, this combination therapy might be a novel chemotherapeutic approach to MMC-insensitive tumors in clinical trials.

We examined the signaling pathway by which hepatocyte growth factor (HGF) induces cell motility, with special focus on the role of extracellular signal-regulated kinase (ERK) in the nucleus. We used Madin-Darby canine kidney cells overexpressing ERK2 because of their prominent motility response to HGF. HGF stimulation of the cells induces not only a rapid, marked, and sustained activation and rapid nuclear accumulation of ERK1/2, but also a prolonged nuclear retention of the activated ERK1/2. Interruption of the ERK1/2 activation by PD98059 treatment of the cells 30 min after HGF stimulation abolishes the HGF-induced cell motility. Enforced cytoplasmic retention of the activated ERK1/2 by the expression of an inactive form of MKP-3 cytoplasmic phosphatase inhibits the cell motility response. Although epidermal growth factor stimulation of the cells induces the activation and nuclear accumulation of ERK1/2, it does not induce the prolonged nuclear retention of the activated ERK1/2, and fails to induce cell motility. In the nucleus, activated ERK1/2 continuously phosphorylate Elk-1, leading to the prolonged expression of c-fos, which results in the expression of several genes such as matrix metalloproteinase (mmp)-9; MMP-9 activity is required for the induction of the cell motility response. Our results indicate that the sustained activity of ERK1/2 in the nucleus is required for the induction of HGF-induced cell motility.Hepatocyte growth factor (HGF), 1 also known as scatter factor, is a multifunctional cytokine capable of inducing a wide variety of biological effects in normal and neoplastic cells. These include proliferation of hepatocytes as well as many types of epithelial and endothelial cells, dissociation of epithelial cell colonies into individual cells and stimulation of their motility, induction of epithelial morphogenesis, induction of angiogenesis, and promotion of the invasion of extracellular matrices. In vivo, HGF has been shown to be involved in embryological development, tissue regeneration, tumorigenesis, and metastasis (reviewed in Refs. 1 and 2).HGF exerts its diverse effects through a high-affinity HGF receptor, which has been identified as the c-met proto-oncogene product (reviewed in Ref. of the ␤-subunit; these phosphorylated tyrosine residues function as docking sites for Src homology 2 domain-containing signal transducers such as PI 3-kinase, phospholipase-C␥, Grb2, Shc, pp60 c-src , and Gab1. As a result of these interactions, HGF induces the activation of various signaling molecules, including Ras, PI 3-kinase, phospholipase-C␥, and 41-/ 43-kDa mitogen-activated protein kinases (ERK2 and ERK1, respectively) (3, 4). However, the precise signaling pathways downstream of c-Met that mediate each of the diverse biological effects of HGF remain largely obscure.Cell motility is a fundamental process required during normal embryonic development, wound repair, inflammatory response, and tumor metastasis (5). Recently, the molecular mechanisms involved in the regulation of cell motility have be...

UCN-01 (7-hydroxy-staurosporine), a selective inhibitor of protein kinase C (PKC), was shown to exhibit antitumor activity in murine and human tumor cell lines in vitro and in vivo. On the other hand, staurosporine, a non-selective protein kinase inhibitor, was not shown to exert antitumor activity in vivo despite its potent antiproliferative activity in vitro. To compare the modes of action of UCN-01 and staurosporine in vitro, the effects of both drugs on the cell cycle progression of human epidermoid carcinoma A431 cells were examined by flow cytometry using propidium iodide (PI) staining. At 50% growth inhibitory concentrations, both UCN-01 and staurosporine induced G1 phase accumulation in the cell cycle. At 80% growth inhibitory concentrations, UCN-01 also induced preferential G1 phase accumulation, but staurosporine mostly induced G2M phase accumulation. Staurosporine also induced higher DNA ploidy when the cells were exposed to the drug for more than one generation time of A431 cells. An analysis of cell kinetics by 5-bromo-2-deoxyuridine incorporation versus DNA content confirmed that the G1 phase block by UCN-01 and the G1 and G2M phase block by staurosporine at the respective doses, as was the case for PI staining. Additionally, DNA synthesis of the cells, which was determined by the uptake of 3H-TdR, was not suppressed at least 8 h after the treatment with UCN-01. These results suggested that UCN-01 could affect the G1 phase of cell cycle in A431 cells in quite different manners from staurosporine. The G1 phase block induced by UCN-01 might be important for the growth inhibitory activity of UCN-01 against A431 cells in vitro and in vivo.

PurposeTo determine the effects of intravenous and perineural dexamethasone on the duration of interscalene brachial plexus block (ISB) with ropivacaine in patients undergoing arthroscopic shoulder surgery.Patients and methodsIn this prospective, randomized, placebo-controlled trial, patients presenting for arthroscopic shoulder surgery with an ISB were randomized to receive ropivacaine 0.75% (group C), ropivacaine 0.75% plus perineural dexamethasone 4 mg (group Dperi), or ropivacaine 0.75% plus intravenous dexamethasone 4 mg (group Div). The primary outcome was the duration of analgesia, defined as the time between performance of the block and the first request for analgesic.ResultsThirty-nine patients were randomized. The median times of sensory block in groups C, Dperi, and Div were 11.2 hours (interquartile range [IQR] 8.0–15.0 hours), 18.0 hours (IQR 14.5–19.0 hours), and 14.0 hours (IQR 12.7–15.1 hours), respectively. Significant differences were observed between groups Dperi and C (P=0.001). Kaplan–Meier analysis for the first analgesic request showed significant differences between groups Dperi and C (P=0.005) and between groups Dperi and Div (P=0.008), but not between groups C and Div.ConclusionPerineural but not intravenous administration of 4 mg of dexamethasone significantly prolongs the duration of effective postoperative analgesia resulting from a single-shot ISB with ropivacaine 0.75%.

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