UCN-01去除放射后肿瘤细胞G2期阻滞及其相关机制

背景与目的:辐射对肿瘤细胞的影响常表现为细胞周期的改变。本研究观察X线照射后肿瘤细胞G2期阻滞及药物UCN鄄01去除此阻滞的情况,并进一步研究其相关机制。方法:选用已知p53突变的人鼻咽癌CNE鄄1细胞系和人肺腺癌973细胞系进行研究,并以p53功能正常的人纤维母细胞瘤HT鄄1080细胞系作为对照。采用细胞培养和流式细胞仪技术分析X线照射对以上细胞系的细胞周期的影响,并观察UCN鄄01对放射后细胞G2期阻滞的影响;应用蛋白印迹法(Westernblot)测定在UCN鄄01去除CNE鄄1细胞G2期阻滞的过程中,细胞内磷酸化CDC2鄄Tyr15含量的相应变化。结果:X线照射明显导致CNE鄄1细胞和973细胞G2阻滞,照射2Gy后两组的G2期细胞比例分别由18.4%和14.8%上升至43.6%和42.8%,两组细胞的G1期阻滞不显著,衡量G1期阻滞的指标“S期消耗率”均较低,分别为14.8%和-1.2%,明显低于p53功能正常的HT鄄1080细胞(57.0%)。UCN鄄01能够去除放射后CNE鄄1细胞和973细胞G2阻滞,两组的G2期细胞比例分别从单纯照射组的63.5%和35.4%降至16.1%和16.3%。CNE鄄1细胞照射后随着细胞G2期阻滞增加,磷酸化CDC2鄄Tyr15的含量相应增加,而UCN鄄01能够降低照射后细胞内磷酸化CDC2鄄Tyr15的含量,并与该药去除细胞G2期阻滞的过程相一致。结论:对于p53突变的人鼻咽癌CNE     

UCN-01增加鼻咽癌细胞株放射敏感性的研究

背景与目的：应用药物提高肿瘤组织的放射敏感性,是改善放疗疗效的主要研究方向之一。本研究通过观察已知p53基因突变的鼻咽癌CNE-1细胞照射后G2期阻滞的情况,以及药物7-hydroxystaurosporine(UCN-01)是否能够去除此G2期阻滞并增加放射敏感性,探讨UCN-01增加放射敏感性的机制。方法：用细胞培养和流式细胞仪（FCM）技术分析X线照射对CNE-1细胞周期（特别是G2期阻滞）的影响,观察UCN-01对放射引起的G2期阻滞的影响。应用细胞克隆形成实验观察UCN-01去除G2期阻滞对放射敏感性的影响。结果：照射明显导致CNE-1细胞G2期阻滞,未照射组（对照组）及照射2Gy、4Gy和6Gy组的细胞G2期比例分别为18.4％、43.6％、77.4％和86.4％,G2期阻滞的程度与照射剂量正相关。UCN-01能够去除放射引起的CNE-1细胞G2期阻滞,50nmol/L、100nmol/L、200nmol/L和400nmol/L UCN-01组细胞的G2比例由对照组的63.5％分别降至28.5％、25.0％、16.1％和13.7％,UCN-01去除G2期阻滞的程度与药物浓度正相关。UCN-01能明显降低放射后CNE-1细胞的克隆形成率,100nmol/L和200nmol/LUCN-01组的放射增敏比分别为2.60和3.09。结论：UCN-01对CNE-1细胞有放射增敏作用,其机制可能与UCN-01去除放射引起的G2期阻滞有关。     

Phase I and pharmacokinetic study of UCN-01 in combination with irinotecan in patients with solid tumors

Purpose 7-Hydroxystaurosporine (UCN-01) is a protein kinase inhibitor that inhibits several serine–threonine kinases including PKC and PDK1. Due to the preclinical synergistic effects seen with topoisomerase I inhibitors and non-overlapping toxicity, UCN-01 and irinotecan were combined in a dose-finding study designed to determine the maximum tolerated dose (MTD), toxicity profile, and pharmacokinetics (PK) of UCN-01 and irinotecan. Methods Patients with incurable solid malignancies received UCN-01 intravenously (IV) as a 3-h infusion on day 1 and irinotecan IV over 90 min on days 1 and 8 of a 21-day cycle. Doses of UCN-01 for subsequent cycles were half the starting dose. Dose level 1 (DL1) consisted of UCN-01 and irinotecan doses of 50 and 60 mg/m², respectively. Blood samples were collected in cycle 1 for UCN-01, irinotecan, and irinotecan metabolites. Results A total of 16 patients were enrolled on the trial at UCN-01/Irinotecan doses of 50/60 mg/m² (DL1; n = 1), 70/60 mg/m² (DL2; n = 6), 90/60 mg/m² (DL3; n = 4), and 70/90 mg/m² (DL4; n = 5). Two dose-limiting toxicities were observed each in DL3 and DL4 (2 grade 3 hypophosphatemia, 1 grade 4 hyperglycemia and grade 3 hypophosphatemia, 1 grade 4 febrile neutropenia). Fatigue, diarrhea, nausea, and anorexia were the most prevalent toxicities. No objective responses were documented, and four patients had stable disease for at least ten cycles. The long half-life (292.0 ± 135.7 h), low clearance (0.045 ± 0.038 l/h), and volume of distribution (14.3 ± 5.9 l) observed for UCN-01 are consistent with prior UCN-01 data. There was a significant decrease in C _max of APC, AUC of APC and SN-38, and AUC ratio of SN-38:irinotecan when comparing days 1 and 8 PK. Conclusions APC and SN-38 exposure decreased when administered in combination with UCN-01. The MTD of the combination based on protocol criteria was defined as 70 mg/m² of UCN-01 on day 1 and 60 mg/m² of irinotecan on days 1 and 8 in a 21-day cycle. Presented in part at the 16th EORTC-NCI-AACR Symposium on Molecular Targets and Cancer Therapeutics, September 2004.     

G1–checkpoint Function Including a Cyclin-dependent Kinase 2 Regulatory Pathway as Potential Determinant of 7–Hydroxystaurosporine (UCN-01)-induced Apoptosis and G1–phase Accumulation

7–Hydroxystaurosporine (UCN-01), which was originally identified as a protein kinase C selective inhibitor, is currently in clinical trials as an anti-cancer drug. We previously showed that UCN-01 induced preferential G1–phase accumulation in tumor cells and this effect was associated with the retinoblastoma (Rb) protein and its regulatory factors, such as cyclin-dependent kinase 2 (CDK2) and CDK inhibitors p21^Cip1/WAF1 and p27^kipl. We demonstrate here that G1–phase accumulation was induced by UCN-01 in Rb-proficient cell lines (WiDr and HCT116 human colon carcinomas and WI-38 human lung fibroblast), and it was accompanied by dephosphorylation of Rb. In addition, UCN-01–induced G1–phase accumulation was also demonstrated in a Rb-defective cell line (Saos-2 human osteosarcoma), but not in a simian virus 40 (SV40)-transformed cell line (WI-38 VA13). Apoptosis was induced by UCN-01 in the two Rb-deficient cell lines, but not in the other Rb-proficient cell lines. These observations suggest that G1–checkpoint function might be important for cell survival during UCN-01 treatment. In addition, there may be a UCN-01–responsive factor in the G1–checkpoint machinery other than Rb which is targeted by SV40. Further studies revealed a correlation between UCN-01–induced G1–phase accumulation and reduction of cellular CDK2 kinase activity. This reduction was strictly dependent on down-regulation of the Thr160–phosphor-ylated form of CDK2 protein, and coincided in part with up-regulation of p27^Kip1, but it was independent of the level of the p21^Cip1/WAF1 protein. These results suggest that G1–checkpoint function, including a CDK2–regulatory pathway, may be a significant determinant of the sensitivity of tumor cells to UCN-01.     

Enhancement of camptothecin-induced cytotoxicity with UCN-01 in breast cancer cells: abrogation of S/G2 arrest

Purpose: To determine the ability of UCN-01 to abrogate the cell cycle arrest induced by camptothecin (CPT) in tumor cells that lack p53 function, and therefore enhance the cytotoxicity of CPT in these cells in relation to normal cells with wild-type p53. Methods: The responses of MDA-MB-231 and GI 101A breast cancer cells were compared to those of normal bovine endothelial cells. Cytotoxicity was assessed by the MTT assay, and the resulting data were modeled using median-effect analysis. Inhibition of DNA synthesis was determined by loss of [³H]thymidine incorporation, and cell cycle status was determined by flow cytometric analysis of propidium-iodide-stained nuclei. Results: UCN-01, a specific inhibitor of protein kinase C (PKC) presently in clinical trials, abrogated CPT-induced activation of S and G₂ checkpoints in human MDA-MB-231 and GI 101A breast carcinoma cells, both of which are mutants for the p53 gene. This abrogation occurred with the use of sublethal doses (100 nM) of UCN-01 and correlated with the enhancement of CPT-induced cytotoxicity. Median-effect analysis showed that synergistic cytotoxic interactions existed between CPT and UCN-01 against these tumor cells. In normal cells, however, abrogation of the S phase arrest caused accumulation in G₀/G₁ phase, perhaps by the presence of wild-type p53 activity, with no change in CPT-induced cytotoxicity. Conclusion: We have shown previously that the cytotoxicity of CPT is correlated with cell cycle response in normal and tumor cells. Low doses of CPT arrest cells in the G₂/M phase and inhibit DNA synthesis, but higher doses cause arrest of cells in S phase. Thus modulation of events at the S and G₂ checkpoints may provide an opportunity to enhance CPT-induced cytotoxicity in tumor cells. The results of this study indicate that UCN-01 enhances the progression of tumor cells through S phase thus greatly increasing CPT-induced cytotoxicity. Normal cells, however, are able to arrest in G₀/G₁ and thus avoid the increased toxicity induced by CPT. Our findings suggest potential usefulness of combining UCN-01 in topoisomerase I inhibitor-based drug therapy for the treatment of breast cancer with a dysfunctional p53 gene. Received: 25 February 1999 / Accepted: 4 October 1999     

G2 checkpoint abrogation and checkpoint kinase-1 targeting in the treatment of cancer

Rigorous quality control steps, termed checkpoints, tightly regulate progression through the cell cycle. DNA-damaging chemotherapy and radiation activate functional cellular checkpoints. These checkpoints can facilitate DNA repair and promote cell death in unrepaired cells. There are at least three DNA damage checkpoints - at G1/S, S, and G2/M - as well as a mitotic spindle checkpoint. Most cancer cells harbour mutations in tumour suppressors and/or oncogenes, which impair certain cell checkpoints. Inhibiting the remaining cell checkpoints - particularly after exposure of cancer cells to chemotherapy and/or radiation - allows cell death, a strategy now being employed in cancer therapeutics. With our increasing knowledge of cell cycle regulation, many compounds have been developed to inhibit specific checkpoint components, particularly at the G2/M transition. One such target is checkpoint kinase-1 (Chk1). We review here the molecular framework of the cell cycle, the rationale for targeting Chk1, the preclinical concepts related to the development of Chk1 inhibitors, and the efficacy and safety results from Chk1 inhibitors now in phase I/II trials.     

CHFR-associated early G2/M checkpoint defects in breast cancer cells

Cell division is a highly regulated process. Checkpoints can halt cell-cycle progression due to adverse conditions such as misalignment of chromosomes to prevent missegregation. The search for new regulators of the cell cycle revealed the mitotic checkpoint gene CHFR (checkpoint with forkhead-associated and ring finger). CHFR coordinates an early mitotic phase by delaying chromosome condensation in response to a mitotic stress. Because aneuploidy and chromosome instability are common in malignant breast tumors, we screened 24 breast cancer cell lines for CHFR expression and demonstrated that 50% (12 of 24) of breast cancer cell lines had low CHFR levels. Expression of CHFR was reactivated with the demethylating agent 5-aza-2'-deoxycytidine (5-aza-dC) in two low-CHFR-expressing cell lines. Eleven of these 12 (92%) low-CHFR-expressing cell lines had an unusually high number of condensed chromosomes and high mitotic indices in response to nocodazole treatment. Transfection of CHFR in one of these cancer cell lines lowered the mitotic index after nocodazole treatment. In conclusion, our data suggested that low CHFR expression associated with high mitotic indices in response to nocodazole treatment were common in the breast cancer cell lines studied. Additional flow cytometry studies and analysis of a protein that interacts with CHFR in vitro, polo-like kinase 1 (PLK1), suggests that this CHFR-associated early G(2)/M checkpoint is complex, involving additional, as yet unidentified, proteins. Further analysis of CHFR in breast cancer cells will be important for understanding the complex mechanisms leading to aneuploidy and chromosomal instability observed in breast cancer.     

柴胡皂苷D通过诱导C2-M期阻滞抑制结直肠癌SW480细胞的增殖

目的:观察柴胡皂苷D（SSD）对于结直肠癌肿瘤细胞增殖的影响并初步探索其潜在的分子机制。方法:运用MTT、细胞计数试验及克隆形成试验观察SSD对细胞增殖的影响。流式细胞仪检测SSD对细胞凋亡及周期分布情况的影响。RT-PCR和Western-blot技术检测G2-M周期阻滞关键调控因子在SW480细胞中的表达。结果:MTT试验发现SSD能够显著抑制结直肠癌肿瘤细胞SW480的增殖,而对正常结直肠细胞FHC的增殖无明显影响。细胞计数试验和克隆形成试验进一步验证了SSD对于SW480细胞增殖的影响(P<0.05)。流式细胞术检测发现SSD不影响细胞的凋亡(P>0.05),但却显著诱导G2-M周期阻滞(P<0.05)。SSD在mRNA水平下调了G2-M周期调控因子CCNA1、CCNA2、CCNB1和CCNB2的表达(P<0.05);在蛋白水平,CCNA2、CCNB1和p34/cdc2明显被下调,p-H3S10上调,p21WAF1/CIP1被明显上调。结论:SSD可以通过上调p21WAF1/CIP1的表达诱导G2-M周期阻滞来抑制结直肠肿瘤细胞SW480的增殖。     

High-mobility group box 2 (HMGB2) modulates radioresponse and is downregulated by p53 in colorectal cancer cell

Overexpression of high-mobility group box 2 (HMGB2) is recently reported in several malignant cancers and was correlated with poor response to preoperative chemoradiotherapy of colorectal cancer patients. To enhance the chemoradiotherapy efficacy, the biological function of HMGB2 was investigated with respect to radiation response. HMGB2 gene knockdown cells were constructed by infecting shRNA expressing lentivirus and clonogenic assay was performed to count the radiosensitivity. HMGB2 knockdown sensitized HCT-116 and HT-29 colorectal cancer cells to ionizing radiation. This could be due to an increased DNA damage and an inefficient DNA damage repair in HMGB2 knockdown cells. In addition, an exposure to radiation downregulated HMGB2 expression in colorectal cancer cells with an intact TP53 gene. HMGB2 gene expression of TP53-mutant cell was not affected by irradiation. p53-mediated downregulation of HMGB2 was confirmed by direct activation of p53 using Nutlin-3 or by inducing p53 expression using Tet-On system. Luciferase reporter assay showed that HMGB2 promoter activity was inversely correlated with the amount p53 cotransfected. Our study revealed that HMGB2 is necessary to protect colorectal cancer cells from DNA damage and efficient DNA repair and p53-mediated downregulation is a critical mechanism of modulating HMGB2 expression.     

Expression of the mitotic checkpoint gene MAD2L2 has prognostic significance in colon cancer

Aneuploidy and genetic instability are a hallmark of colorectal cancer and other solid tumors, and they are thought to enhance tumor progression. The gene MAD2L2 (mitotic arrest deficient 2-like 2) encodes the spindle checkpoint protein MAD2L2 (or MAD2B), a key component of a surveillance system that delays anaphase until all chromosomes are correctly oriented. Defects in this mitotic checkpoint are known to contribute to genetic instability, i.e., numerical and structural aberrations of chromosomes. We have previously identified MAD2L2 as significantly upregulated in locally restricted colorectal tumors by gene expression profiling. So far, MAD2L2 has not been reported to play a major role in human cancer in contrast to its homologue MAD2. To address this question, 118 histologically confirmed colorectal lesions were analyzed by quantitative real-time PCR for expression of MAD2L2, and compared to normal colon tissue from 11 patients. Twenty-five out of 118 tumor samples (21%) showed MAD2L2 overexpression of 3-fold or more compared to normal colon, and the fraction of overexpressing tumors increased with tumor stage. Correspondingly, protein levels of MAD2L2 were found to be significantly upregulated in tumors as compared to matched normal tissue. Tumors with upregulated MAD2L2 expression had significantly higher numbers of aberrant mitotic figures (anaphase bridges), an indication of chromosomal instability. Elevated expression of MAD2L2 was significantly correlated with reduced patient survival. By multivariate analysis, MAD2L2 expression was retained as an independent prognostic parameter for patient survival. Thus, our results demonstrate that overexpression of MAD2L2 correlates with bad prognosis in colorectal cancer.     

In vitro phosphorylation of BRCA2 by the checkpoint kinase CHEK2

Germline mutations in both BRCA2 and CHEK2 are associated with an increased risk for male breast cancer. To search for potential interactions between the products of these breast cancer susceptibility genes, we undertook systematic mapping of BRCA2 for potential phosphorylation sites by CHEK2. In vitro kinase assays and mass spectrometric analysis identified a 50 amino-acid fragment within the N-terminus of BRCA2 potentially targeted by CHEK2, containing two major phosphopeptides. Inducible overexpression of this peptide, but not a derivative with mutated phosphorylation sites, leads to increased chromosome fragmentation and suppression of cellular proliferation. These results suggest a link between CHEK2 and BRCA2 pathways, which may contribute to the spectrum of cancers associated with germline CHEK2 mutations.     

The amount of DNA damage needed to activate the radiation-induced G2 checkpoint varies between single cells

Background and purpose

The radiation-induced G2 checkpoint helps facilitate DNA repair before cell division. However, recent work has revealed that human cells often escape the G2 checkpoint with unrepaired DNA breaks. The purpose was to explore whether G2 checkpoint activation occurs according to a threshold level of DNA damage.

Materials and methods

G2 checkpoint activation was assayed at 75-90 min and 24-48 h after X-ray irradiation of BJ diploid fibroblasts and U2OS osteosarcoma cells. Multiparameter flow cytometry with pacific blue barcoding, and flow cytometry-based sorting of phospho-H3 positive cells to microscope slides, were used to examine the DNA damage marker γ-H2AX in individual mitotic cells that had escaped the G2 checkpoint.

Results

For all radiation doses and times tested, the number of γ-H2AX foci varied between individual mitotic cells. At 75 min the median levels of γ-H2AX in mitotic cells increased with higher radiation doses. At 24-48 h, following a prolonged G2 checkpoint, cells were more resistant to checkpoint re-activation by a second dose of radiation.

Conclusion

Our results suggest that different amounts of DNA damage are needed to activate the G2 checkpoint in individual cells. Such single cell variation in checkpoint activation may potentially contribute to radiation-induced genomic instability.     

5-Fluorouracil response in a large panel of colorectal cancer cell lines is associated with mismatch repair deficiency

Background:

Colorectal cancer is (CRC) one of the commonest cancers and its therapy is still based on few drugs. Currently, no biological criteria are used to choose the most effective of the established drugs for treatment.

Methods:

A panel of 77 CRC cell lines was tested for sensitivity to 5-fluorouracil (5FU) using the SRB assay. The responses were grouped into three categories and correlated with genetic changes in the cell lines.

Results:

The strongest and most clearcut correlation was between 5-fluorouracil response and replication error status (mismatch repair deficiency). All the other significant correlations (loss of heterozygosity for DCC and mutations in TGFbIIR) are secondary to the association with replication error status.

Interpretation and conclusion:

Our findings validate previous analyses based mainly on clinical data, and indicate that replication error status could be a useful guide to 5-fluorouracil-based CRC therapy. Essentially, all previously described correlations with 5FU response are secondary to the association with replication error status.     

c-Myc alters the DNA damage-induced G2/M arrest in human mammary epithelial cells

Effects of c-Myc overexpression on the DNA damage-induced G2/M checkpoint were studied in finite lifespan, normal human mammary epithelial cells (HMECs). Previously, we showed that c-Myc attenuates G1/S arrest and leads to an inappropriate entry of cells with damaged DNA into the S phase, following treatment with ionising radiation (IR). Here we show that, in striking contrast to control cells, c-Myc-overexpressing HMECs demonstrate a significant attenuation of the G2/M arrest, following IR, and enter into inappropriate mitoses. At the molecular level, ectopic overexpression of c-Myc leads to an unusually high level of expression of cyclin B1, and the elevated levels of cyclin B1 were maintained, after gamma-irradiation. Introduction of DNA damage in c-Myc-overexpressing, normal mammary epithelial cells eventually induces apoptosis, indicating a dramatic sensitisation by c-Myc of DNA damage-induced apoptosis. These two remarkable phenotypes, checkpoint attenuation and sensitisation to apoptosis, resulting from a deregulation of the protooncogene c-myc, may produce a unique pattern of alternating cycles, consisting first of amplification of DNA damage, followed by apoptosis-assisted selective pressure. The result of this alternating pattern of damage apoptosis could facilitate the selection of certain genomic alterations required for cellular survival and cellular transformation.     

免疫检查点抑制剂在转移性结直肠癌患者治疗中的研究进展

免疫检查点抑制剂的出现,增加了许多实体肿瘤的治疗选择。尽管在黑素瘤和肺癌的治疗中效果良好,但大多数转移性结直肠癌患者无法从免疫治疗中获益。免疫检查点抑制剂在错配修复功能缺失转移性结直肠癌患者中明确有显著和持久的临床反应,即使在既往多线治疗失败的群体中也是如此。然而,这种临床获益仅限于小部分肿瘤患者,约占转移性结直肠癌的4%。事实上抗程序性死亡受体1(programmed cell death protein 1,PD-1)单抗对错配修复功能缺失转移性结直肠癌患者是无效的。迫切需要新颖的治疗策略使这些肿瘤具有免疫应答。破坏肿瘤的疗法(化学疗法,放射疗法和靶向疗法),从而释放肿瘤抗原,是免疫检查点抑制和其他疗法相结合的最直接的策略。这些标准疗法远没有像曾经担心的那样削弱免疫反应,反而还可以增强免疫应答。     

Apoptosis induced by cryo-injury in human colorectal cancer cells is associated with mitochondrial dysfunction

Cryotherapy, a method of in situ ablation, is used in the treatment of colorectal liver metastases with variable results. During the treatment, the central area of treated tumor undergoes necrotic destruction by lethal cryo-injury; however, the cellular response of tumor exposed to sublethal cryo-injury at the peripheral zone is unclear. In our study, we have identified the induction of apoptosis by cryo-injury at -10 degrees C in 4 colorectal cancer cell lines (HT29, HCT116, KM12C and KM12SM). The apoptosis was characterized by chromatin condensation, transferase-mediated dUTP nick end-labeling (TUNEL) staining, proteolytic cleavage of poly(ADP-ribose) polymerase (PARP) and cytokeratin 18, and activation of caspase-3. The occurrence and intensity of cryo-induced apoptosis did not correlate with the functional status of p53 in the cell lines studied. The expression of anti-apoptotic proteins (Bcl-2, Bcl-X(L)) and pro-apoptotic proteins (Bax, Bcl-X(S), Bad, and Bak) in response to cryo-injury varied in this cell line panel. The basal level of Bcl-2/Bax protein ratio correlated inversely to the apoptotic rate. We further demonstrated that Bax level decreased in cytosol and increased in mitochondria, followed by a loss of mitochondrial membrane potential after cryo-injury in HT29 cells. These findings indicate that cryo-injury induces apoptosis in colorectal cancer cells via disruption of mitochondrial integrity. The cryo-induced apoptosis was also identified in a nude mouse tumor xenograft model. Our elucidation of the apoptosis pathway induced by cryo-injury implies that synergistic combination of cryosurgery with pharmacological agents that augment of apoptosis induction may have clinical relevance in treating colorectal liver metastasis.     

Epithelial expression of FHL2 is negatively associated with metastasis-free and overall survival in colorectal cancer

Background:

Four-and-a-half LIM domains protein 2 (FHL2) is a component of the focal adhesion structures and has been suggested to have a role in cancer progression. It has been shown to be overexpressed in the colorectal cancer (CRC).

Methods:

Here, we examined a possible prognostic value of FHL2 in CRC. Immunohistochemistry for FHL2 was performed on 296 CRCs without distant metastases at the time of surgery. Staining in the epithelial compartment was quantitatively evaluated using image analysis, and results were related to clinical variables. Antibody specificity was tested using small-interfering RNA transfection in hTERT-immortalised myofibroblasts.

Results:

Varying degrees of cytoplasmic FHL2 expression by neoplastic epithelial cells were detectable in all cases. Higher FHL2 expression in the epithelial compartment was an independent adverse prognostic factor. Multivariate Cox analysis shows that expression in the tumour invasion front (P<0.001) as well as in the centre of the tumour (P<0.001) was associated with metachronous metastases independently of the clinicopathological variables; expression in the tumour invasion front was also associated with overall survival independently of the clinicopathological variables (P<0.01).

Conclusion:

Higher FHL2 expression is involved in CRC progression and correlates with the development of metachronous metastases and overall survival, suggesting that FHL2 is an independent adverse prognostic indicator for CRC.     

UCN-01提高肺腺癌细胞放射敏感性的实验观察

目的：观察p53基因突变的人肺腺癌细胞系973照射后G2期阻滞以及药物7－Hydroxystaurosporine(UCN-01)是否能去除此阻滞并增加放射敏感性，探讨UCU－01增加放射敏感性的机制。方法：用细胞培养和流式细胞仪技术分析X射线照射对973细胞周期（特别是G2期阻滞）的影响，观察UCN－01对放射引起的G2期阻滞的影响。应用细胞克隆形成实验观察UCN－01对放射敏感性的影响。结果：照射明显导致973细胞G2期阻滞，未照射组（对照组）及2、4和6Gy组的G2期比例分别为14．8％、42．8％、55．9％和60．5％，G2期阻滞的程度与照射剂量呈正相关。UCN－01能够去除放射引起的973细胞G2期阻滞，50、100、200和400nmol/L UCN-01组G2期比例由对照组的35．4％分别降至23．5％、19．6％、16．3％和12．4％，UCN－01去除G2期阻滞的程度与药物浓度呈正相关。UCN－01能明显降低放射后973细胞的存活分数，200和400nmol/L UCN-01组的放射增敏比分别为1．13和1．31。结论：UCN－01对973细胞有放射增敏作用，机制可能与UCN－01去除放射引起的G2期阻滞有关。