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Umetani, N; Sasaki, S; Masaki, T; Watanabe, T; Matsuda, K; Muto, T
doi: 10.1054/bjoc.1999.0869pmid: 10638959
The aim of this study was to clarify the role of APC and K- ras mutations in non-polypoid colorectal tumorigenesis. DNA from 63 adenomas (31 polypoid, 17 superficial elevated, 15 superficial depressed), 66 submucosally invasive carcinomas (47 polypoid, 19 non-polypoid) and 34 advanced carcinomas were examined for K- ras codon 12 point mutations and APC mutations in the mutation cluster region. K- ras mutation: the frequency in superficial depressed adenomas was lower than that in polypoid adenomas (0% vs 31%: P = 0.018). The frequency in non-polypoid carcinomas was lower than that in polypoid carcinomas (11% vs 56%: P = 0.0008), and was relatively low compared with that in polypoid adenomas (11% vs 31%). APC mutation: the frequency in superficial depressed adenomas was lower than that in polypoid adenomas (7% vs 43%: P = 0.016), and that in polypoid carcinomas was similar to that in non-polypoid carcinomas. Polypoid adenomas, polypoid carcinomas and advanced carcinomas had almost the same frequency. There may be some pathway other than the conventional adenoma-carcinoma sequence in development of non-polypoid carcinomas. The precursors of most non-polypoid carcinomas are considered to be de novo or superficial depressed adenomas. In this non-polypoid pathway, APC mutation seems to be requisite but K- ras mutation not. It is possible that new APC mutations are acquired after the development of superficial depressed adenomas. © Copyright 2000 Cancer Research Campaign
Dohjima, T; Ohno, T; Banno, Y; Nozawa, Y; Wen-yi, Y; Shimizu, K
doi: 10.1054/bjoc.1998.0870pmid: 10638960
EWS-Fli-1, a fusion gene found in Ewing’s sarcoma and primitive neuro-ectodermal tumour (PNET), encodes a transcriptional activator and promotes cellular transformation. We have made stable Ewing’s sarcoma cells expressing antisense EWS-Fli-1 transcripts by transfecting the antisense EWS-Fli-1 expression plasmid. These cells showed partial loss of endogenous EWS-Fli-1 proteins and suppression of the cell growth. To elucidate the molecular mechanisms underlying the growth inhibition, we examined the changes of signal transducing proteins by immunoblot analysis in Ewing’s sarcoma cells stably expressing antisense EWS-Fli-1 transcripts. Western blotting of the cell proteins revealed that expressions of phospholipase Cβ2 and β3 (PLCβ2, PLCβ3), and also protein kinase C α and β (PKCα, β) were significantly reduced by transfecting with antisense EWS-Fli-1. The inositol phosphates production by bradykinin (BK), but not platelet-derived growth factor (PDGF), was suppressed in these cells. These results suggest that the PLCβ2 and PLCβ3 may play a role in tumour proliferation in Ewing’s sarcoma cells. © 2000 Cancer Research Campaign
Durany, N; Joseph, J; Jimenez, O M; Climent, F; Fernández, P L; Rivera, F; Carreras, J
doi: 10.1054/bjoc.1999.0871pmid: 10638961
We have compared the levels of phosphoglycerate mutase (EC 5.4.2.1), 2,3-bisphosphoglycerate phosphatase (EC 3.1.3.13), creatine kinase (EC 2.7.3.2) and enolase (EC 4.2.1.11) activities and the distribution of their isoenzymes in normal breast tissue and in breast carcinoma. Tumour tissue had higher phosphoglycerate mutase and enolase activity than normal tissue. Creatine kinase activity was higher in seven out of 12 tumours. In contrast 2,3-bisphosphoglycerate phosphatase activity was lower. Phosphoglycerate mutase, enolase and 2,3-bisphosphoglycerate phosphatase presented greater changes in the oestrogen receptor-negative/progesterone receptor-negative breast carcinomas than in the steroid receptor-positive tumours. Determined by electrophoresis, type BB phosphoglycerate mutase, type BB creatine kinase and αα-enolase were the major isoenzymes detected in normal breast tissue. Types αγ and γγ enolase, types MB and MM phosphoglycerate mutase were detected in much lower proportions. In tumours a decrease of phosphoglycerate mutase isoenzymes possessing M-type subunit and some increase of enolase isoenzymes possessing γ-type subunit was observed. No detectable change was observed in the creatine kinase phenotype. © 2000 Cancer Research Campaign
doi: 10.1054/bjoc.1999.0872pmid: 10638962
Molecular mechanisms and/or intrinsic factors controlling cellular radiosensitivity are not fully understood in mammalian cells. The recent studies have suggested that nuclear factor κB (NF-κB) is one of such factors. The activation and regulation of NF-κB are tightly controlled by IκB-α, a cellular inhibitory protein of NF-κB. Most importantly, phosphorylation regulates activity of the inhibitor IκB-α, which sequesters NF-κB in the cytosol. Two different pathways for the phosphorylation of IκB-α are demonstrated, such as serine (at residues 32 and 36) and tyrosine (at residue 42) phosphorylations. To assess a role of the transcription factor, NF-κB, on cellular sensitivity to DNA damaging agents, we constructed three different types of expression plasmids, i.e. S-IκB (mutations at residues 32 and 36), Y-IκB (mutation at residue 42) and SY-IκB (mutations at residues 32, 36 and 42). The cell clones expressing S-IκB and Y-IκB proteins became sensitive to X-rays as compared with the parental and vector-transfected cells. The cell clones expressing SY-IκB were further radiosensitive. By the treatment with herbimycin A, an inhibitor of phosphorylation, the X-ray sensitivity of cells expressing SY-IκB did not change, while that of the cells expressing S-IκB and Y-IκB and the parental cells was enhanced. Change in the sensitivity to adriamycin and UV in those clones was very similar to that in the X-ray sensitivity. The inhibition of IκB-α phosphorylation at serine and tyrosine acts independently on the sensitization to X-rays, adriamycin and UV. These findings suggest that the transcriptional activation induced by NF-κB may play a role in the DNA damage repair. The present study proposes a possibility that the inactivation of NF-κB by inhibition of both serine and tyrosine phosphorylations may be useful for the treatment of cancer in radio- and chemotherapies. © 2000 Cancer Research Campaign
Isonishi, S; Ohkawa, K; Tanaka, T; Howell, S B
doi: 10.1054/bjoc.1999.0873pmid: 10638963
Down-regulation of protein kinase C (PKC) by 12- O -tetradecanoylphorbol-13-acetate (TPA) enhances the sensitivity of human ovarian carcinoma 2008 cells to various types of platinum compounds such as cisplatin (DDP), carboplatin and (–)-(R)-2-aminomethylpyrrolidine (1,1-cyclobutanedicarboxylato)-platinum(II) monohydrate (DWA) by a factor of two- to threefold. TPA enhanced the sensitivity of the DDP-resistant 2008/C13*5.25 subline to each of these three drugs to the same extent as for the 2008 cells. The extent of PKC down-regulation and drug sensitization depended on the duration of TPA exposure; maximum effect was achieved with a 48 h pretreatment. Sensitization was TPA concentration-dependent and was maximal at 0.05 μM TPA. 2008 cells expressed only the PKCα and PKCζ isoforms. Western blot analysis revealed that whereas the expression of PKCα was reduced by TPA the level of PKCζ was not affected. These results suggest that PKCα is the isotype responsive to TPA in these cells and that platinum drug sensitivity can be modulated by this isoform alone. In parallel to its effect on PKCα, TPA decreased cellular glutathione content by 30 ± 3 (standard deviation (s.d.) % in 2008 cells and by 41 ± 3 (s.d.) % in 2008/C13*5.25 cells. TPA also increased accumulation of DDP and DWA by 70%, although this effect was limited to the 2008/C13*5.25 cells. TPA rendered 2008 and 2008/C13*5.25 cells resistant to cadmium chloride by a factor of 3.7 and 3.6-fold respectively, suggesting a significant increase in cellular metallothionein content. Although the mechanism of TPA induced sensitization is not yet fully understood, this study points to a central role for PKCα in modulating platinum drug sensitivity. © 2000 Cancer Research Campaign
Cronauer, M V; Nessler-Menardi, C; Klocker, H; Maly, K; Hobisch, A; Bartsch, G; Culig, Z
doi: 10.1054/bjoc.1999.0874pmid: 10638964
Interactions between polypeptide growth factors and the androgen receptor (AR) are important for regulation of cellular events in carcinoma of the prostate. Basic fibroblast growth factor (bFGF), the prototype of heparin-binding growth factors, and the AR are commonly expressed in prostate cancer. bFGF diminished prostate-specific antigen protein in the supernatants of androgen-stimulated human prostate cancer cells LNCaP by 80%. In the present study, we asked whether the bFGF effect on prostate-specific antigen is preceded by action on AR expression. LNCaP cells were treated with bFGF and AR protein expression was determined by immunoblotting and ligand binding assay. bFGF down-regulated AR protein in a dose-dependent manner showing a maximal effect at 50 ng ml−1both in the presence or absence of dihydrotestosterone. Down-regulation of AR protein expression occurred already after 8 h of bFGF treatment and a maximal inhibition was observed 24 h after addition of bFGF to culture media. As AR expression can be reduced by an increase in intracellular calcium levels, we investigated whether the bFGF effect on AR protein is mediated by this mechanism. Calcium release from intracellular stores and store-operated calcium influx after treatment with either bFGF or calcium ionophore A 23187 were measured by single cell fluorescence technique. The ionophore A 23187 was able to induce calcium influx and an increase in cytoplasmic calcium concentration in LNCaP cells. In contrast, bFGF was incapable of eliciting a similar effect. In contrast to AR protein, AR mRNA levels were not affected by bFGF as shown by semiquantitative reverse transcription polymerase chain reaction. In summary, these studies show that bFGF is a potent negative regulator of AR protein expression in the human prostate cancer cell line LNCaP. © 2000 Cancer Research Campaign
Kunisue, H; Kurebayashi, J; Otsuki, T; Tang, C K; Kurosumi, M; Yamamoto, S; Tanaka, K; Doihara, H; Shimizu, N; Sonoo, H
doi: 10.1054/bjoc.1999.0875pmid: 10638965
Anti-oestrogen is effective for the treatment of oestrogen receptor (ER)-positive breast carcinomas, butmost of these tumours become resistant to anti-oestrogen. It has been suggested that anti-oestrogen therapy may induce a HER2signalling pathway in breast cancer cells and this may cause resistance to anti-oestrogen. Thus, it is conceivable that combinedtherapy with anti-oestrogen and anti-HER2 antibody might be more effective. In the present study, we investigated the effect ofcombined treatment with a humanized anti-HER2 monoclonal antibody, rhumAbHER2 (trastuzumab), and an anti-oestrogen, ICI 182,780, onthe cell growth of three human breast cancer cell lines which respectively express different levels of ER and HER2. The combinedtreatment enhanced the growth inhibitory effect on ML-20 cells, which express a high level of ER and a moderate level of HER2, butshowed no additive effect on either KPL-4 cells, which express no ER and a moderate level of HER2, or MDA-MB-231 cells, which express no ER and a low level of HER2. It is also suggested that both the antibody and anti-oestrogen induce a G1–S blockade and apoptosis. These findings indicate that combined treatment with anti-HER2 antibody and anti-oestrogen may be useful for thetreatment of patients with breast cancer expressing both ER and HER2. © 2000 Cancer Research Campaign
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