SEARCH · PubMed Health

Results for “HCT116 Cells”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources

PubMed2006

Effect of indomethacin on Bfl-1, WISP-1 and proliferating cell nuclear antigen in colon cancer cell line HCT116 cells.

BACKGROUND: Non-steroidal anti-inflammatory drugs such as indomethacin can inhibit the growth of tumors through both the cyclooxygenase-2 (COX-2) dependant and COX-2 independent pathways, but the exact mechanism has not yet been shown. In our previous study, COX-2 independent proteins (Bfl-1, WISP-1 and proliferating cell nuclear antigens [PCNA]) in indomethacin-treated colorectal cancer cells with the use of proteomics technology had been identified. OBJECTIVES: To study and confirm the effect of indomethacin on the expression of Bfl-1, WISP-1 and PCNA in human colon cancer line HCT116 cells and the COX-2 independent tumor inhibiting pathway. METHODS: Human colon cancer cell line HCT116 cells were divided into a treatment with indomethacin (IC 50) group, and a treatment with dimethyl sulfoxide (DMSO) as a control group for 48 h. The expression of Bfl-1, WISP-1 and PCNA, mRNA and protein were determined by a real-time quantitative PCR and Western blot, respectively. RESULTS: Indomethacin down-regulated the expression of Bfl-1, WISP-1 and PCNA mRNA in vitro (9.53 +/- 0.15 vs 27.87 +/- 0.12, 7.37 +/- 0.58 vs 20.17 +/- 0.58, 5.17 +/- 0.06 vs 0.87 +/- 0.06). Indomethacin also down-regulated the expression of Bfl-1, WISP-1 and PCNA protein (40.01 +/- 1.61 vs 43.76 +/- 1.63, 22.50 +/- 1.17 vs 30.30 +/- 1.55, 17.69 +/- 1.18 vs 20.80 +/- 1.08). CONCLUSIONS: Inducing apoptosis and inhibiting proliferation contribute to the anticancer activity of indomethacin via COX-2 independent pathway of Bfl-1, WISP-1 and PCNA. This further confirms the results of our previous study.

Apoptosis↗

PubMed2004

Influence of p53 and p21Waf1 expression on G2/M phase arrest of colorectal carcinoma HCT116 cells to proteasome inhibitors.

Ubiquitin-mediated protein degradation in vertebrates has been implicated in cell cycle control. In this report we explored the effects of proteasome inhibitors (MG132, lactacystin and ALLN) on cell cycle distribution. Colorectal carcinoma HCT116 cells were treated with proteasome inhibitor MG132. The results showed that MG132 inhibited cell proliferation in a dose-dependent manner. MG132 arrested HCT116 cells at G2/M phase, which was associated with drug-induced blockade of p53 degradation and/or induction of p53-related gene expression along with the accumulation of cyclin B, cyclin A and p21. MG132 treated HCT116 (wild-type) had a similar cell cycle distribution as the MG132 treated HCT116 (p53-/-) and HCT116 (p21-/-) cells, suggesting that p53 and p21 may not be essential for MG132-induced G2/M phase arrest. The release experiments from nocodazole-induced mitotic phase cells indicated that MG132 inhibits the proliferation of HCT116 cells via arrest in the G2 phase. In addition, when HCT116 cells were exposed to combination of sodium butyrate and MG132 enhanced cell growth inhibition and induction of apoptosis were observed.

Acetylcysteine↗

PubMed2005

Microarray analysis identifies apoptosis regulatory gene expression in HCT116 cells infected with thermostable direct hemolysin-deletion mutant of Vibrio parahaemolyticus.

The thermostable direct hemolysin (TDH) is considered as a major virulence factor of Vibrio parahaemolyticus. We observed this potential in several human cancer cell lines by using the TDH-producing wild-type (RIMD2210633) as well as tdh-deletion mutant of V. parahaemolyticus and found that the deletion of tdh did not affect cytotoxicity to any of the cell lines tested. DNA fragmentation and annexin V staining showed that both wild-type and tdh-mutant trigger apoptosis in these cells. To understand the molecular basis of cell death in the absence of TDH, gene expression profile of human colon cancer cell line HCT116 infected with tdh-deletion mutant was carried out using human cDNA microarrays consisting of 33,000 known genes. In infected cells, differentially expressed genes including genes for early growth response, growth arrest and DNA damage, and activating transcription factor that affect programmed cell death pathways were detected. Interestingly, mutant strains having a deletion in type III secretion system 1 (TTSS1) failed to elicit DNA fragmentation in HCT116 cells. Our results strongly suggest that apoptosis requires functional TTSS1 and TTSS1-dependent translocation factor(s) to be associated with the host cell death, and thus pathogenesis of V. parahaemolyticus.

Annexin A5↗

PubMed2002

Nocodazole-induced p53-dependent c-Jun N-terminal kinase activation reduces apoptosis in human colon carcinoma HCT116 cells.

Microtubule-interfering agents are widely used in cancer chemotherapy, and prognostic results vary significantly from tumor to tumor, depending on the p53 status. In preliminary experiments, we compared the expression and phosphorylation profiles of more than 100 protein kinases and protein phosphatases in human colorectal carcinoma cell line HCT116 between p53+/+ and p53-/- cells in response to short term nocodazole treatment through application of Kinetworks immunoblotting screens. Among the proteins tracked, the regulation of the phosphorylation of c-Jun N-terminal kinase (JNK)1/2 at Thr-183/Tyr-185 was the major difference between p53+/+ and p53-/- cells. With the loss of the p53 gene, the levels of phosphorylation of Ser-63 of c-Jun and Thr-183/Tyr-185 of JNK1/2 in p53-/- cells did not increase as markedly as in p53+/+ cells in response to a 1-h treatment with nocodazole or other microtubule-disrupting drugs such as vinblastine and colchicine. Similar observations were also made in MCF-7 and A549 tumor cells, which were rendered p53-deficient by E6 oncoprotein expression. However, arsenate-induced JNK activation in p53-/- cells was preserved. Inhibition of p53 expression by its antisense oligonucleotide also attenuated nocodazole-induced JNK activation in p53+/+ cells. Surprisingly, cotransfection of p53+/+ cells with dominant negative mutants of JNK isoforms and treatment of p53+/+ cells with the JNK inhibitor SP600125 actually further enhanced apoptosis in p53+/+ cells by up to 2-fold in response to nocodazole. These findings indicate that inhibition of p53-mediated JNK1/2 activity in certain tumor cells could serve to enhance the apoptosis-inducing actions of cancer chemotherapeutic agents that disrupt mitotic spindle function.

Antineoplastic Agents↗

PubMed2003

Bax plays a pivotal role in thapsigargin-induced apoptosis of human colon cancer HCT116 cells by controlling Smac/Diablo and Omi/HtrA2 release from mitochondria.

Bax is a crucial mediator of the mitochondrial pathway for apoptosis, and loss of this proapoptotic Bcl-2 family protein contributes to drug resistance in human cancers. We report here that the endoplasmic reticulum Ca(2+)-ATPase inhibitor thapsigargin (THG) induces apoptosis of human colon cancer HCT116 cells through a Bax-dependent signaling pathway controlling the cytosolic release of mitochondrial apoptogenic molecules. Treating HCT116 cells with THG results in caspase-8 activation; Bid cleavage; Bax conformational change and mitochondrial translocation; the release of cytochrome c, Smac/Diablo, and Omi/HtrA2 into the cytosol; caspase-3 activation; and apoptosis. In contrast, knockout of Bax completely abrogates the full processing/activation of caspase-3 but has no effect on the processing of caspase-8 and the initial cleavage of caspase-3 to p24 fragment after THG treatment. The caspase-8-specific inhibitor z-IETD-fmk, as well as pan-caspase inhibitor z-VAD-fmk, but not the calpain inhibitor E-64d, prevents Bid cleavage, Bax conformational change, and subsequent caspase-3 processing and apoptosis. Caspase-8 processing is dependent on de novo protein synthesis; DR5 expression is strongly up-regulated by THG treatment. Moreover, the absence of Bax blocks THG-induced Omi and Smac release from mitochondria, and expression of cytosolic Omi (GFP-IETD-Omi) or Smac (GFP-IETD-Smac) restores the sensitivity of Bax-knockout HCT116 cells to apoptosis in response to THG treatment. Taken together, our results indicate that Bax-dependent Smac and Omi release plays an essential role in caspase-3 activation and apoptosis induced by THG in human colon cancer HCT116 cells.

Apoptosis↗

PubMed2001

Bile acid hydrophobicity is correlated with induction of apoptosis and/or growth arrest in HCT116 cells.

Faecal bile acids have long been associated with colon cancer; highly hydrophobic bile acids, which induce apoptosis, have been implicated in the promotion of colon tumours. The moderately hydrophobic chemopreventive agent ursodeoxycholic acid (UDCA) does not induce apoptosis; rather, it causes colon-derived tumour cells to arrest their growth. To investigate the relationship between bile acid hydrophobicity and biological activity we examined 26 bile acids for their capacity to induce apoptosis or alter cell growth. We found that the rapidity with which, and the degree to which, bile acids could induce apoptosis or growth arrest was correlated with their relative hydrophobicities. Of the bile acids tested, only deoxycholic acid (DCA) and chenodeoxycholic acid, the most hydrophobic bile acids tested, could induce apoptosis in less than 12 h in the human colon cancer cell line HCT116. The moderately hydrophobic bile acids hyoDCA, lagoDCA, norDCA, homoUDCA and isoUDCA induced growth arrest at 12 h but longer incubations resulted in apoptosis. Conjugation of glycine or taurine to the bile acids decreased relative hydrophobicity and eliminated biological activity in our assays. In addition, we tested a subset of these bile acids for their ability to translocate across cell membranes. When (14)C-labelled and (3)H-labelled DCA, UDCA and lagoDCA were added to cell cultures, we found only minimal uptake by colon cells, whereas hepatocytes had considerably higher absorption. These experiments suggest that hydrophobicity is an important determinant of the biological activity exhibited by bile acids but that under our conditions these activities are not correlated with cellular uptake.

Apoptosis↗

PubMed2003

Enhanced radiosensitization with gemcitabine in mismatch repair-deficient HCT116 cells.

Gemcitabine [2',2'-difluoro-2'-deoxycytidine (dFdCyd)] is a potent ionizing radiation sensitizer in solid tumor cells in vitro and in vivo. Previously, we have demonstrated (Shewach et al., Cancer Res., 54: 3218-3223, 1994) a strong correlation between depletion of dATP (caused by dFdCyd diphosphate-mediated inhibition of ribonucleotide reductase) and radiosensitization. In addition, we and others (Latz et al., Int. J. Radiat. Oncol. Biol. Phys., 41: 875-882, 1998; Ostruszka and Shewach, Cancer Res., 60: 6080-6088, 2000) have shown that the accumulation of cells in S phase prior to irradiation is also important for radiosensitization with dFdCyd. This led us to hypothesize that the incorporation of incorrect nucleotides because of the dATP pool imbalance was important for radiosensitization with dFdCyd, and, therefore, cells deficient in mismatch repair (MMR) would exhibit greater radiosensitization. We tested this hypothesis by evaluating the ability of HCT116 colon carcinoma cell lines, which differ in MMR proficiency, to be radiosensitized by dFdCyd. The MMR-proficient cell line (HCT116 + ch3) was more sensitive to dFdCyd alone than were the MMR-deficient cell lines (HCT116, HCT116 + ch2, and HCT116 p53(-/-)). Interestingly, the MMR-proficient cells could not be radiosensitized at concentrations of dFdCyd IC(96)) enhanced cell killing with radiation. In contrast, the MMR-deficient cells were radiosensitized at concentrations of dFdCyd or=80% decrease in dATP within 4 h after drug addition, and this low dATP level was maintained for another 12-20 h. Although the IC(50) of dFdCyd was unable to sustain a >80% decrease in the dATP level in the MMR-proficient cells, the IC(90) did achieve this level of dATP depletion; however, it was unable to radiosensitize the MMR-proficient cells. Similar results were obtained with HCT116 cells, in which the MMR deficiency was corrected by transfection with a vector containing the hMLH1 cDNA. In addition, the deletion of p53 did not increase radiation enhancement ratios. These results demonstrate that MMR deficiency promotes radiosensitization with dFdCyd. We suggest that dATP depletion produces errors of replication in MMR-deficient cells, which, if left unrepaired, enhances cell death by ionizing radiation.

Base Pair Mismatch↗

PubMed2006

Inhibition of Fas expression by RNAi modulates 5-fluorouracil-induced apoptosis in HCT116 cells expressing wild-type p53.

Drug resistance to 5-fluorouracil (5-FU) is still a major limitation to its clinical use. In addition, the clinical value of p53 as a predictive marker for 5-FU-based chemotherapy remains a matter of debate. Here, we used HCT116 human colorectal cancer cells expressing wild-type p53 and investigated whether inhibition of Fas expression by interference RNA modulates 5-FU-induced apoptosis. Cells were treated with 5-FU (1, 4 or 8 microM) for 8-48 h. Cell viability was evaluated by trypan blue dye exclusion. Apoptosis was assessed by changes in nuclear morphology and caspase activity. The interference RNA technology was used to silence Fas expression. Caspase activation, p53, Fas, cytochrome c, and Bcl-2 family protein expression was evaluated by immunoblotting. 5-FU was cytotoxic in HCT116 cells (p<0.001). Nuclear fragmentation and caspase-3, -8 and -9 activities were also markedly increased in HCT116 cells after 5-FU (p<0.001). In addition, wild-type p53 and Fas expression were 25- and 4-fold increased (p<0.05). Notably, when interference RNA was used to inhibit Fas, 5-FU-mediated nuclear fragmentation and caspase activity were markedly reduced in HCT116 cells. Finally, western blot analysis of mitochondrial extracts from HCT116 cells exposed to 5-FU showed a 6-fold increase in Bax, together with a 3-fold decrease in cytochrome c (p<0.001). In conclusion, 5-FU exerts its cytotoxic effects, in part, through a p53/Fas-dependent apoptotic pathway that involves Bax translocation and mitochondrial permeabilization.

Antimetabolites, Antineoplastic↗

PubMed2001

Ecteinascidin 743 induces protein-linked DNA breaks in human colon carcinoma HCT116 cells and is cytotoxic independently of topoisomerase I expression.

Ecteinascidin 743 (Et743; NSC 648766) is a potent antitumor agent presently in clinical trials. Et743 selectively alkylates guanine N2 from the minor groove of duplex DNA and bends the DNA toward the major groove. This differentiates Et743 from other DNA-alkylating agents presently in the clinic. To date, the cellular effects of Et743 have not been elucidated. Recently, Et743 DNA adducts have been found to suppress gene expression selectively and to induce topoisomerase I (top1) cleavage complexes in vitro and top1-DNA complexes in cell culture. In the present study, we characterized the DNA damage and the cell cycle response induced by Et743 in human colon carcinoma HCT116 cells. Alkaline elution experiments demonstrated that micromolar concentrations of Et743 produced comparable frequencies of DNA-protein cross-links and DNA single-strand breaks. The single-strand breaks were protein-cross-linked and were not associated with detectable DNA double-strand breaks. By contrast with camptothecin, these lesions persisted for several hours after drug removal and were not formed at 4 degrees C. Et743 treatment induced transient p53 elevation, dose-dependent cell cycle accumulation in G2-M and in G1- and S-phase, and inhibition of DNA synthesis. The sensitivity of camptothecin-resistant mouse leukemia P388/ CPT45 cells, which fail to express detectable top1, was similar to the sensitivity of wild-type P388 cells, suggesting that top1 is not a critical target for the antiproliferative activity of Et743.

Antineoplastic Agents, Alkylating↗

PubMed2004

[Effect of indomethacin on human colorectal cancer cells HCT116 and SW480].

OBJECTIVE: To explore the effect of indomethacin on COX non-expressing colon cancer cells (HCT116 and SW480). METHODS: We used MTT assay to assess the effect of indomethacin on the growth of the cultured cells. Transplanted tumor models were established with BALB/c nude mice and were treated with indomethacin at 3 mg/(kg x d) for 4 weeks. Tumor volume and mouse weight were recorded every other day. Histologic examination was performed. RESULTS: After 48-hour incubation with various concentrations of indomethacin, the growth of the two cell lines was significantly suppressed in a dose-dependent manner, with IC50 being (318.2 +/- 12.7) micromol/L and (701.4 +/- 29.5) micromol/L, respectively. Indomethacin also effectively retarded the tumor growth within xenografts by 44.6% according to the tumor volume. No obvious toxicity was observed in these nude mice. CONCLUSION: Indomethacin can inhibit the growth of COX-negative human colon cancer cells both in vitro and in vivo, which indirectly suggests that indomethacin exerts its anti-tumor effect at least partly independent of the cyclooxygenase inhibitory profile.

Animals↗

PubMed1996

Induction of p21 mediated by reactive oxygen species formed during the metabolism of aziridinylbenzoquinones by HCT116 cells.

Aziridinylbenzoquinones are a group of antitumor agents that elicit cytotoxicity by generating either alkylating intermediates or reactive oxygen species. The mechanism of toxicity may not always, however, involve profound damage of cellular constituents, but may involve a cytostatic effect through interference with the cell cycle. In this context, we have examined the induction of the cell cycle inhibitor p21 (WAF1, CIP1, or sdi1), whose overexpression suppresses the growth of various tumor cells, in human tumor cells metabolizing 3,6-diaziridinyl-1,4-benzoquinone (DZQ) and its C2,C5-substituted derivatives: 2,5-bis-(carboethoxyamino) (AZQ) and 2, 5-bis-2(-hydroxyethylamino) (BZQ). Both DZQ and AZQ were effectively activated by HCT116 human colonic carcinoma cells; the activation of the former involved largely a dicoumarol-sensitive activity, whereas that of the latter appeared to be accomplished primarily by one-electron transfer reductases. BZQ was not a substrate for the dicoumarol-sensitive enzyme in HCT116 cells. Cellular activation of the first two quinones was associated with formation of oxygen-centered radicals as detected by EPR in conjunction with the spin trap 5,5'-dimethyl-1-pyrroline-N-oxide. The redox transitions of DZQ involved hydroxyl radical formation and were strongly inhibited by catalase, whereas those of AZQ showed a strong superoxide anion component sensitive to superoxide dismutase. These signals were suppressed by N-acetylcysteine with concomitant production of a thiyl radical adduct. This suggests an effective electron transfer between the thiol and free radicals formed during the activation of these quinones. DZQ and AZQ induced significantly the expression of p21 in HCT116 cells, but a 10-fold higher concentration of AZQ was required to achieve the level of induction elicited by DZQ. BZQ had little effect on p21 expression. p21 induction at both mRNA and protein levels correlated with the inhibition of either cyclin-dependent kinase activity or cell proliferation. p21 induction elicited by the above quinones was inhibited by N-acetylcysteine, whereas the non-sulfur analog, N-acetylalanine, was without effect. Catalase and superoxide dismutase did not effect p21 induction by aziridinylbenzoquinones in HCT116 cells, thus suggesting that extracellular sources of oxygen radicals generated by plasma membrane reductases have no influence in the expression of this gene. Hydrogen peroxide, a product of quinone redox cycling, elicited an increase of p21 mRNA levels in HCT116 and K562 human chronic myelogenous leukemia cells. The latter lacks p53, one of the activators of p21 transcription, thus suggesting that p21 expression can be accomplished in a p53-independent manner in these cells. This study suggests that p21 induction is mediated by an increase in the cellular steady-state concentration of oxygen radicals and that the greater effectiveness in p21 induction by DZQ may be related to its efficient metabolism by NAD(P)H:quinone oxidoreductase activity in HCT116 cells.

Acetylcysteine↗

PubMed2000

Apoptosis induction in HCT116 cells by cytochalasins isolated from the fungus Daldinia vernicosa.

We examined the induction of apoptosis by cytochalasin (cc) derivatives (1-14) isolated from the Japanese fungus Daldinia vernicosa to HCT116 human colon cancer cell line based on their cytotoxicity, DNA ladder and DNA fragmentation ratio in agarose gel electrophoresis, and morphological changes. Most cc derivatives tested here induced apoptosis. Particularly cytochalasin 1 (cc1), monoacetate of 1 (cc1Ac), and cc14 were the most potent apoptosis inducers. These apoptotic activities were stronger than that of cytochalasin D as a known apoptosis inducer in HCT116 cell. However, cc4 and cc12 induced necrosis. The structure-activity relationship including their cytotoxicity will be discussed.

Apoptosis↗

PubMedDate not supplied

Induction of apoptosis in human colon cancer HCT116 cells treated with an extract of the plant product, Chios mastic gum.

A hexane extract of the plant product Chios mastic gum (He-CMG) is demonstrated to kill human colon cancer cells in vitro via the process of anoikis. Specifically, the sequence of events includes He-CMG-induced GI-arrest of the cells, detachment of the cells from the substrate and subsequent apoptosis. Anoikis is dependent on the concentration and duration of treatment with He-CMG. Presence of the pan-caspase inhibitor, Z-VAD-fmk, did not prevent cell detachment, but it did prevent apoptosis of the detached cells indicating that the process of cell detachment, but not apoptosis, is independent of caspase activation. He-CMG-induced apoptosis is associated with activation of the initiator caspases-8, and -9 and the effector caspase-3. Caspases are activated in cells at a relatively long time after detachment, and caspase-3 activation may require caspase-8 or caspase-9 activation, as determined by using HCT116 isogenic clones impaired in apoptosis mechanisms that involve these two caspases. Finally, electron microscopy observations indicated a time-dependent appearance of morphological features both typical and non-typical of apoptosis in cells treated with He-CMG for various periods of time. Taken together, the results demonstrated that He-CMG induces apoptosis in HCT116 cells and, therefore, further in vivo and in vitro studies of the anticancer activities of this plant product are warranted.

Anoikis↗

PubMed2006

hRFI overexpressed in HCT116 cells modulates Bcl-2 family proteins when treated with 5-fluorouracil.

Exogenous overexpression of hRFI, originally isolated in our laboratory, inhibits not only death receptor-mediated apoptosis but also the mitochondrial apoptosis induced by several chemotherapeutic agents including 5-fluorouracil (5-FU). Recently, it has become clear that hRFI targets and degradates caspase-8 and -10 in death receptor-mediated apoptosis by E3 ubiquitin activity in a ring finger domain homologous to that of X-chromosome-linked inhibitor of apoptosis protein (XIAP). However, the cellular mechanism of the inhibition of mitochondrial apoptosis by hRFI has not been fully elucidated. We prepared HCT116 overexpressing hRFI (HCT116/hRFI) cells and comprehensively analyzed the expression changes of 51 apoptosis-related genes with or without 5-FU treatment between HCT116/hRFI and mock cells using microfluidic low-density arrays. As a result, we identified four genes (Bcl-2, Bcl-XL, cIAP2, and CFLAR) whose expression was four or more times higher in HCT116/ hRFI cells than in HCT116/LacZ cells, and found that Bcl-2 and the ratio of Bcl-2/Bax or Bcl-2/Bak were upregulated when HCT116/hRFI cells were treated with 5-FU. Furthermore, we also validated the up-regulation of Bcl-2 and Bcl-XL in HCT116/hRFI cells treated with 5-FU by Western blot analysis. Such evidence suggests that the modulation of Bcl-2 family proteins seen in 5-FU treatment plays an important role in the anti-apoptotic function of HCT116/hRFI cells.

Antimetabolites, Antineoplastic↗

PubMed2003

The DNA minor groove-alkylating cyclopropylpyrroloindole drugs adozelesin and bizelesin induce different DNA damage response pathways in human colon carcinoma HCT116 cells.

As members of the cyclopropylpyrroloindole family, adozelesin and bizelesin cause genomic DNA lesions by alkylating DNA. Adozelesin induces single-strand DNA lesions, whereas bizelesin induces both single-strand lesions and double-strand DNA cross-links. At equivalent cytotoxic concentrations, these agents caused different biological responses. Low adozelesin concentrations (e.g., 0.5 nM) induced a transient S-phase block and cell cycle arrest in G(2)-M, as well as increased induction of p53 and p21, whereas a high drug concentration (e.g., 2.5 nM) caused apoptosis but no p21 induction. In contrast, both low and high bizelesin concentrations enhanced p53 and p21 induction and triggered G(2)-M cell cycle arrest and eventual senescence without significant apoptotic cell death. However, in cells lacking p21, bizelesin, as well as adozelesin, triggered apoptosis, indicating that p21 was crucial to sustained bizelesin-induced G(2)-M arrest. Thus, despite similar abilities to alkylate DNA, the chemotherapeutic agents adozelesin and bizelesin caused a decrease in HCT116 tumor cell proliferation by different pathways (i.e., adozelesin induced apoptosis, and bizelesin induced senescence).

Antineoplastic Agents, Alkylating↗

PubMed1996

DNA-dependent protein kinase inhibitor (OK-1035) suppresses p21 expression in HCT116 cells containing wild-type p53 induced by adriamycin.

The tumor growth suppressor p21 has been shown to be induced by wild-type p53 (wt-p53) and to be a potent inhibitor of cyclin-dependent kinases and PCNA/DNA polymerase delta. Although wt-p53 is reported to be phosphorylated by several protein kinases, the function and significance of the phosphorylation of wt-p53 are not yet fully understood. Using OK-1035, a selective inhibitor of DNA-dependent protein kinase (DNA-PK), we demonstrated the importance of the phosphorylation of wt-p53 by DNA-PK in the DNA damage-mediated expression of the p21 gene. Treatment of HCT116, a human colon carcinoma cell line, with adriamycin induced the expression of wt-p53 and p21. By addition of OK-1035 to this culture, the induction of p21 protein was significantly decreased in a dose-dependent manner, whereas wt-p53 induction was not affected. Northern blot analysis revealed that suppression of p21 protein expression by OK-1035 resulted from reduction in the level of p21 mRNA. OK-1035 did not directly affect the binding ability of wt-p53 to its consensus DNA sequence. Our observations support the idea that wt-p53 induces the transcriptional activation of the p21 gene only after it is phosphorylated by DNA-PK.

Amino Acid Sequence↗

PubMed2003

Dephosphorylation of p53 during cell death by N-alpha-tosyl-L-phenylalanyl chloromethyl ketone.

The apoptotic function of N-alpha-tosyl-L-phenylalanyl chloromethyl ketone (TPCK) was investigated in cultured human colorectal carcinoma cells (HCT116). TPCK-induced apoptosis was shown to be p53-dependent in HCT116 cells during the early stage of incubation. The function of p53 was required for TPCK-induced activation of caspase-3 and caspase-7. TPCK promoted dephosphorylation of p53 on serine residues at 6, 9, 46, 376, and 378 in parallel with the activation of p53 transcriptional activity. HCT116 p53-/- cells expressing p53 mutant, in which serine residues at 6, 9, 46, 376, and 378 were replaced by aspartic acids, were resistant to TPCK-induced apoptosis suggesting the requirement of dephosphorylation of p53 on serine residues during TPCK-induced apoptosis.

Apoptosis↗

PubMed2006

Chronic but not acute conjugated linoleic acid treatment inhibits deoxycholic acid-induced protein kinase C and nuclear factor-kappaB activation in human colorectal cancer cells.

Conjugated linoleic acid (CLA) has anti-carcinogenic effects in a variety of cancers including colon cancer. Secondary bile acids on the other hand are known as tumour promoters in colon cancer with effects on protein kinase C (PKC) and nuclear factor kappa B (NF-kappaB) signalling pathways. The aim of this study was to examine acute and chronic, isomer-specific effects of CLA on bile salt-induced PKC and NF-kappaB signal transduction in human colon cancer cells. HCT116 cells were treated with 100 mumol/l and 50 mumol/l cis-9,trans-11-CLA and trans-10,cis-12-CLA for 24 h and 14 days, respectively. The cells were then transfected with DNA coding for PKC beta1-EGFP (enhanced green fluorescent protein), PKC delta-EGFP or PKC zeta-EGFP fusion protein and activated with deoxycholic acid (DCA), phorbol myristate acetate (PMA) or C2-ceramide. PKC translocation was observed using real-time photomicroscopy and fluorescent microscopy and NF-kappaB analyses by gel shift assays. Chronic c-9,t-11-CLA and t-10,c-12-CLA treatment inhibited DCA-induced PKC beta1 and PKC delta translocation and also inhibited NF-kappaB activation. Acute CLA treatment had no effect on PKC or NF-kappaB activation. In conclusion this study indicates that chronic CLA treatment inhibits DCA-induced PKC and NF-kappaB activation in colon cancer cells. These data suggest mechanisms by which CLA may influence the course of colonic cancer.

Colorectal Neoplasms↗