Anti-epidermal growth factor receptor antibody C225 inhibits angiogenesis in human transitional cell carcinoma growing orthotopically in nude mice.
(17/35913)
Epidermal growth factor receptor (EGFR) regulates the growth and progression of human transitional cell carcinoma (TCC) of the bladder. We have shown that therapy targeting EGFR inhibited the growth of human TCC established orthotopically in nude mice. The purpose of this study was to evaluate whether EGFR-directed therapy affects angiogenesis associated with the growth and metastasis of human TCC. We determined the cytostatic effect and the effect on production of angiogenic factors after in vitro treatment of the human TCC cell line 253J B-V with MAb C225, a chimerized monoclonal anti-EGFR antibody. The 253J B-V cells were implanted orthotopically into athymic nude mice, and established tumors (4 weeks) were treated with i.p. MAb C225. Expression of the angiogenic factors vascular endothelial growth factor (VEGF), interleukin-8 (IL-8), and basic fibroblast growth factor (bFGF) was evaluated by immunohistochemistry and in situ mRNA hybridization analyses and correlated with microvessel density evaluated after immunohistochemical staining with anti-CD31. In vitro treatment with MAb C225 inhibited mRNA and protein production of VEGF, IL-8, and bFGF by 253J B-V cells in a dose-dependent manner. MAb C225 therapy of nude mice with established TCCs growing orthotopically resulted in inhibition of growth and metastasis compared with controls (P <0.0005). VEGF, IL-8, and bFGF expression was significantly lower in treated tumors than in controls. The down-regulation of these angiogenic factors preceded the involution of blood vessels. These studies indicate that therapy with anti-EGFR MAb C225 has a significant antitumor effect mediated, in part, by inhibition of angiogenesis. (+info)
Persistent induction of apoptosis and suppression of mitosis as the basis for curative therapy with S-1, an oral 5-fluorouracil prodrug in a colorectal tumor model.
(18/35913)
In an effort to improve the therapeutic selectivity of 5-fluorouracil (FUra) against colorectal cancer, S-1, a combination agent including a prodrug of FUra with two modulators, was recently developed by Taiho Pharmaceuticals Co. S-1 is a combination of tegafur (FT), 5-chloro-2,4-hydroxypyridine, and potassium oxonate in the molar ratio of 1.0:0.4:1.0, with the latter two components as inhibitors of dihydropyrimidine dehydrogenase and phosphoribosylpyrophosphate transferase, respectively. In this study, the therapeutic selectivity and efficacy of S-1 (oral) was compared with FT (oral) and FUra (i.v. infusion) in rats bearing advanced colorectal cancer by using clinically relevant schedules. The maximum tolerated doses (MTDs) of S-1, FT, and FUra were 31.5, 200, and 25 mg/kg/d for 7 days and 22.5, 150, and 12.5 mg/kg/d for 28 days, respectively. The therapeutic index of S-1 was 4- to 5-fold higher than that of either FT or FUra. S-1 achieved 100% complete tumor regression (CR) at its MTD in both 7-day and 28-day schedules. Furthermore, the high incidences of stomatitis, alopecia, and diarrhea observed with FUra and FT, were not observed with S-1. In an attempt to understand the basis for the observed superior therapeutic selectivity with S-1, we studied pharmacokinetic analysis of FUra, drug-induced apoptosis, suppression of mitosis, and inhibition of thymidylate synthase (TS) after S-1, FUra, or FT administration. The peak plasma FUra concentrations derived from FUra or S-1 (FT) at comparable MTDs were similar, but the plasma level of FUra was higher with S-1 than with FUra. Induction of high and sustained apoptosis was achieved with S-1. Although the initial level of apoptosis induced by FUra was comparable to S-1, it was not sustained. The sustained level of apoptosis appears to correlate with tumor growth inhibition. Mitotic figures were more greatly suppressed with S-1 treatment than with FUra. Studies on TS inhibition indicated that, although both S-1 and FUra caused a 4- to 6-fold induction of total TS protein, single oral administration of S-1 was superior to 24-h infusion of FUra in suppressing free TS. The data are consistent with the observation that the therapeutic efficacy of S-1 (100% cure) over FUra is associated with high and sustained levels of drug-induced apoptosis, greater suppression of mitosis, and inhibition of free TS in tumor tissues. (+info)
Tyrosine kinase inhibitor emodin suppresses growth of HER-2/neu-overexpressing breast cancer cells in athymic mice and sensitizes these cells to the inhibitory effect of paclitaxel.
(19/35913)
Overexpression of the HER-2/neu proto-oncogene, which encodes the tyrosine kinase receptor p185neu, has been observed in tumors from breast cancer patients. We demonstrated previously that emodin, a tyrosine kinase inhibitor, suppresses tyrosine kinase activity in HER-2/neu-overexpressing breast cancer cells and preferentially represses transformation phenotypes of these cells in vitro. In the present study, we examined whether emodin can inhibit the growth of HER-2/neu-overexpressing tumors in mice and whether emodin can sensitize these tumors to paclitaxel, a commonly used chemotherapeutic agent for breast cancer patients. We found that emodin significantly inhibited tumor growth and prolonged survival in mice bearing HER-2/neu-overexpressing human breast cancer cells. Furthermore, the combination of emodin and paclitaxel synergistically inhibited the anchorage-dependent and -independent growth of HER-2/neu-overexpressing breast cancer cells in vitro and synergistically inhibited tumor growth and prolonged survival in athymic mice bearing s.c. xenografts of human tumor cells expressing high levels of p185neu. Both immunohistochemical staining and Western blot analysis showed that emodin decreases tyrosine phosphorylation of HER-2/neu in tumor tissue. Taken together, our results suggest that the tyrosine kinase activity of HER-2/neu is required for tumor growth and chemoresistance and that tyrosine kinase inhibitors such as emodin can inhibit the growth of HER-2/neu-overexpressing tumors in mice and also sensitize these tumors to paclitaxel. The results may have important implications in chemotherapy for HER-2/neu-overexpressing breast tumors. (+info)
A requirement for protein kinase C inhibition for calcium-triggered apoptosis in acute lymphoblastic leukemia cells.
(20/35913)
We have evaluated the cytotoxicities of the combinations of calcium mobilizers and PKC inhibitors against human acute lymphoblastic leukemia (ALL) cells. Here we report that calcium mobilizers alone or PKC inhibitors alone do not induce apoptosis in human ALL cells. However, the combinations of calcium mobilizers with potent inhibitors of PKC cause significant apoptosis in ALL cells. Our results provide experimental evidence that PKC blocks Ca2+-triggered apoptosis in human ALL cells. Thus, PKC inhibitors can be used to enhance the antileukemic activity of chemical or biological agents that trigger an apoptotic calcium signal in ALL cells. The exquisite sensitivity of ALL cells to calcium-dependent apoptosis in the presence of PKC inhibitors could provide the basis for new treatment programs against ALL. (+info)
Differential responses of normal, premalignant, and malignant human bronchial epithelial cells to receptor-selective retinoids.
(21/35913)
Using an in vitro lung carcinogenesis model consisting of normal, premalignant, and malignant human bronchial epithelial (HBE) cells, we analyzed the growth inhibitory effects of 26 novel synthetic retinoic acid receptor (RAR)- and retinoid X receptor (RXR)-selective retinoids. RAR-selective retinoids such as CD271, CD437, CD2325, and SR11364 showed potent activity in inhibiting the growth of either normal or premalignant and malignant HBE cells (IC50s mostly <1 microM) and were much more potent than RXR-selective retinoids. Nonetheless, the combination of RAR- and RXR-selective retinoids exhibited additive effects in HBE cells. As the HBE cells became progressively more malignant, they exhibited decreased or lost sensitivity to many retinoids. The activity of the RAR-selective retinoids, with the exception of the most potent retinoid, CD437, could be suppressed by an RAR panantagonist. These results suggest that: (a) RAR/RXR heterodimers play an important role in mediating the growth inhibitory effects of most retinoids in HBE cells; (b) CD437 may act through an RAR-independent pathway; (c) some of the RAR-selective retinoids may have the potential to be used in the clinic as chemopreventive and chemotherapeutic agents for lung cancer; and (d) early stages of lung carcinogenesis may be responsive targets for chemoprevention by retinoids, as opposed to later stages. (+info)
Increased sensitivity of hydroxyurea-resistant leukemic cells to gemcitabine.
(22/35913)
Tumor cell resistance to certain chemotherapeutic agents may result in cross-resistance to related antineoplastic agents. To study cross-resistance among inhibitors of ribonucleotide reductase, we developed hydroxyurea-resistant (HU-R) CCRF-CEM cells. These cells were 6-fold more resistant to hydroxyurea than the parent hydroxyurea-sensitive (HU-S) cell line and displayed an increase in the mRNA and protein of the R2 subunit of ribonucleotide reductase. We examined whether HU-R cells were cross-resistant to gemcitabine, a drug that blocks cell proliferation by inhibiting ribonucleotide reductase and incorporating itself into DNA. Contrary to our expectation, HU-R cells had an increased sensitivity to gemcitabine. The IC50 of gemcitabine was 0.061 +/- 0.03 microM for HU-R cells versus 0.16 +/- 0.02 microM for HU-S cells (P = 0.005). The cellular uptake of [3H]gemcitabine and its incorporation into DNA were increased in HU-R cells. Over an 18-h incubation with radiolabeled gemcitabine (0.25 microM), gemcitabine uptake was 286 +/- 37.3 fmol/10(6) cells for HU-R cells and 128 +/- 8.8 fmol/10(6) cells for HU-S cells (P = 0.03). The incorporation of gemcitabine into DNA was 75 +/- 6.7 fmol/10(6) cells for HU-R cells versus 22 +/- 0.6 fmol/10(6) cells for HU-S cells (P < 0.02). Our studies suggest that the increased sensitivity of HU-R cells to gemcitabine results from increased drug uptake by these cells. This, in turn, favors the incorporation of gemcitabine into DNA, resulting in enhanced cytotoxicity. The increased sensitivity of malignant cells to gemcitabine after the development of hydroxyurea resistance may be relevant to the design of chemotherapeutic trials with these drugs. (+info)
Inhibition of p53 transcriptional activity by Bcl-2 requires its membrane-anchoring domain.
(23/35913)
We show here that the anti-apoptosis protein Bcl-2 potently inhibits p53-dependent transcriptional activation of various p53-responsive promoters in reporter gene co-transfection assays in human embryonic kidney 293 and MCF7 cells, without affecting nuclear accumulation of p53 protein. In contrast, Bcl-2(Deltatransmembrane (TM)), which lacks a hydrophobic membrane-anchoring domain, had no effect on p53 activity. Similarly, in MCF7 cells stably expressing either Bcl-2 or Bcl-2(DeltaTM), nuclear levels of p53 protein were up-regulated upon treatment with the DNA-damaging agents doxorubicin and UV radiation, whereas p53-responsive promoter activity and expression of p21(CIP1/WAF1) were strongly reduced in MCF7-Bcl-2 cells but not in MCF7-Bcl-2(DeltaTM) or control MCF7 cells. The issue of membrane anchoring was further explored by testing the effects of Bcl-2 chimeric proteins that contained heterologous transmembrane domains from the mitochondrial protein ActA or the endoplasmic reticulum protein cytochrome b5. Both Bcl-2(ActA) and Bcl-2(Cytob5) suppressed p53-mediated transactivation of reporter gene plasmids with efficiencies comparable to wild-type Bcl-2. These results suggest that (a) Bcl-2 not only suppresses p53-mediated apoptosis but also interferes with the transcriptional activation of p53 target genes at least in some cell lines, and (b) membrane anchoring is required for this function of Bcl-2. We speculate that membrane-anchored Bcl-2 may sequester an unknown factor necessary for p53 transcriptional activity. (+info)
Induction of low density lipoprotein receptor (LDLR) transcription by oncostatin M is mediated by the extracellular signal-regulated kinase signaling pathway and the repeat 3 element of the LDLR promoter.
(24/35913)
Oncostatin M (OM) activates the transcription of the human low density lipoprotein receptor (LDLR) in HepG2 cells through a sterol-independent mechanism. Our previous studies showed that mutations within the repeat 3 sequence of the LDLR promoter significantly decreased OM activity on LDLR promoter luciferase reporter constructs that contain the sterol responsive element-1 (repeat 2) and Sp1 binding sites (repeats 1 and 3). In this study, we investigated the signal transduction pathways that are involved in OM-induced LDLR transcription. In HepG2 cells, OM induced a rapid increase in LDLR mRNA expression, with increases detected at 30 min and maximal induction at 1 h. This OM effect was not blocked by protein synthesis inhibitors, inhibitors of p38 kinase, phosphatidylinositol 3-kinase, or c-Jun N-terminal kinase, but OM activity was completely abolished by pretreating cells with inhibitors of the extracellular signal-regulated kinase (ERK) kinase (mitogen/ERK kinase (MEK)). To investigate whether the repeat 3 sequence of the LDLR promoter is the OM-responsive element that converts ERK activation at the promoter level, three luciferase reporters, pLDLR-TATA containing only the TATA-like elements of the promoter, pLDLR-R3 containing repeat 3 and the TATA-like elements, and pLDLR-234 containing repeats 1, 2, 3 and the TATA-like elements were constructed and transiently transfected into HepG2 cells. OM had no effect on the basal promoter construct pLDLR-TATA; however, including a single copy of repeat 3 sequence in the TATA vector (pLDLR-R3) resulted in a full OM response. The activity of OM on pLDLR-R3 was identical to that of pLDLR-234. Importantly, the ability of OM to increase luciferase activities in both pLDLR-R3- and pLDLR-234-transfected cells was blocked in a dose-dependent manner by inhibition of MEK. These results demonstrate that the mitogen-activated protein kinase MEK/ERK cascade is the essential signaling pathway by which OM activates LDLR gene transcription and provide the first evidence that the repeat 3 element is a new downstream target of ERK activation. (+info)