[Show abstract][Hide abstract] ABSTRACT: An important challenge in nasopharyngeal carcinoma (NPC) research is to develop effective predictors of tumor recurrence following treatment to determine whether immediate adjuvant therapy is necessary. We retrospectively analyzed archived specimens collected from 45 patients with paired samples of primary NPC (pNPC) and recurrent NPC (rNPC). Clinical samples were collected from the Cancer Center Databases of the First People's Hospital of Foshan and Shantou Central Hospital (affiliates of Sun Yat-Sen University) between 2001 and 2012. Expression levels of phosphor-Stat3 (p-Stat3), signalosome complex subunit 5 (Jab1/Csn5), Akt1, C/EBP homologous protein (CHOP), Ki-67, and apoptosis were determined by immunohistochemistry in pNPC and rNPC samples from the same patients. Differences in these markers between the short-term interval to recurrence (ITR) group (ITR <18 months) and long-term ITR group (ITR ≥18 months) were further analyzed. In Cox's regression analysis, the ITR was significantly associated as an independent‑negative prognostic factor for overall survival (hazard ratio, 0.211; 95% confidence interval, 0.053-0.841; P=0.027). p-Stat3 was increased in the short-term ITR group (ITR <18 months) and tended to be lower in the long-term ITR group (ITR ≥18 months). In the short-term ITR group, nuclear Akt expression was significantly increased in paired rNPC (P=0.028). In the long-term ITR group, the expression of nuclear Jab1/Csn5 (P=0.047) and assessment of apoptosis measured with TdT-mediated dUTP nick end‑labeling (TUNEL) (P=0.003) was significantly increased in paired rNPC. The results suggest that differences between short- and long-term ITR may predict outcome in rNPC. Furthermore, the overexpression of Jab1/Csn5 and Akt may contribute to the carcinogenesis of rNPC, and Akt seems to promote the progression of short-term ITR. Intra-individual changes of Jab1/Csn5, Akt, and TUNEL may help to identify short-term ITR.
[Show abstract][Hide abstract] ABSTRACT: Little is known about the clinical implication of hepatitis B virus (HBV) infection in Epstein–Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC).Objective
This study aimed to investigate the clinical characteristics and prognostic factors in patients with newly-diagnosed NPC with HBV infection.Study designA total of 722 patients with pathologically-diagnosed NPC who received comprehensive treatment at First People's Hospital of Foshan between June 2006 and December 2011 were enrolled in this retrospective study; 79 and 643 patients were HBsAg(+) and HBsAg(−), respectively. The correlations between HBV (HBsAg status and HBV DNA load) and EBV DNA were analyzed, further long-term survival and prognostic factors also were explored.ResultsWe reported NPC patients with HBsAg(+) represented worse outcome, and distant-failure especially liver metastasis was more common in these patients. HBV infection was more frequent in younger patients and male patients. No correlation was observed between the pre-treatment plasma EBV DNA load (cutoff, 1500 copies/ml) and HBsAg status (positive or negative; r = −0.036, P = 0.392), or the pre-treatment plasma EBV DNA load and HBV DNA load (r = 0.042, P = 0823).Conclusions
Both HBV and EBV infection is an independent negative prognostic factor for long-term survival, distant metastasis, especially liver metastasis, was more common in NPC patients with HBsAg(+), and it seemed no link between EBV DNA load and HBsAg status in NPC.
No preview · Article · Nov 2014 · Journal of Clinical Virology
[Show abstract][Hide abstract] ABSTRACT: Nasopharyngeal carcinoma (NPC) is a malignant tumor originating in the epithelium. Radiotherapy is the standard therapy, but tumor resistance to this treatment reduces the 5-year patient survival rate dramatically. Studies are urgently needed to elucidate the mechanism of NPC radioresistance. Epigenetics - particularly microRNAs (miRNAs) and DNA methylation - plays an important role in carcinogenesis and oncotherapy. We used qRT-PCR analysis and identified a miRNA signature from differentially expressed miRNAs. Our objectives were to identify the role of miR-24 in NPC tumorigenesis and radioresistance and to identify the mechanisms by which miR-24 is regulated. We found that miR-24 inhibited NPC cell growth, promoted cell apoptosis, and suppressed the growth of NPC xenografts. We showed that miR-24 was significantly downregulated in recurrent NPC tissues. When combined with irradiation, miR-24 acted as a radiosensitizer in NPC cells. One of the miR-24 precursors was embedded in a CpG island. Aberrant DNA methylation was involved in NPC response to radiotherapy, which linked inactivation of miR-24 through hypermethylation of its precursor promoter with NPC radioresistance. Treating NPC cells with the DNA-hypomethylating agent 5-aza-2'-deoxycytidine compensated for the reduced miR-24 expression. Together, our findings showed that miR-24 was negatively regulated by hypermethylation of its precursor promoter in NPC radioresistance. Our findings defined a central role for miR-24 as a tumor-suppressive miRNA in NPC and suggested its use in novel strategies for treatment of this cancer.
No preview · Article · Oct 2014 · Molecular Cancer Therapeutics
[Show abstract][Hide abstract] ABSTRACT: In this retrospective study, the correlation between pre- and post-treatment plasma Epstein-Barr virus (EBV) DNA and circulating immune subsets as well as the prognostic implications was investigated in nasopharyngeal carcinoma (NPC) patients. Patients (n = 356) were diagnosed and received comprehensive treatment at the First People's Hospital of Foshan from 2006 to 2010. Pre- and post-treatment plasma EBV DNA load and circulating immune subsets (percentage of CD3+ T cell, CD3+ CD4+ T cells, CD3+ CD8+ T cells, CD19+ B cells and CD56+ NK cells) were analyzed by real-time PCR and flow cytometry. Patient age correlated negatively with CD3+ T cells (r = -0.264, P = 0.001) and positively with CD56+ NK cells (r = 0.272, P = 0.001). Pre-treatment plasma EBV DNA correlated negatively with CD19+ B cells (r = -0.223, P = 0.009) and CD4/CD8 ratio (r = -0.177, P = 0.047). Patients with low CD19+ B cell had poorer 5-year progression-free survival (PFS) (66.6 vs. 81.8 %, P = 0.036) and 5-year overall survival (OS) (70.5 vs. 81.5 %, P = 0.097) than patients with high CD19+ B cells. Low CD19+ B cells was identified as a negative prognostic factor for 5-year PFS (hazard ratio [HR] 0.487; P = 0.040), but not for 5-year OS (HR 0.550; P = 0.102) in multivariate analysis. Post-treatment plasma EBV DNA was the most important prognostic factor for 5-year PFS (HR 2.983; P = 0.006) and 5-year OS (HR 3.927; P < 0.001). This study demonstrates the clinical value of circulating CD19+ B cell measurements in NPC patients.
No preview · Article · Oct 2014 · Medical Oncology
[Show abstract][Hide abstract] ABSTRACT: Jab1/CSN5 is a multifunctional protein that plays an important role in integrin signaling, cell proliferation, apoptosis, and the regulation of genomic instability and DNA repair. Dysregulation of Jab1/CSN5 activity has been shown to contribute to oncogenesis by functionally inactivating several key negative regulatory proteins and tumor suppressors. In this review, we discuss our current understanding of the relationship between Jab1/CSN5 and DNA damage and summarize recent findings regarding opportunities for and challenges to therapeutic intervention.
Preview · Article · Feb 2014 · Cancer biology & therapy
[Show abstract][Hide abstract] ABSTRACT: Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus--associated malignancy that is most common in East Asia, Africa, and Alaska. Radiotherapy is the main treatment option; unfortunately, disease response to concurrent radiotherapy and chemotherapy varies among patients with NPC, and in many cases, NPC becomes resistant to radiotherapy. Our previous studies indicated that Jab1/CSN5 was overexpressed and plays a role in the pathogenesis and radiotherapy resistance in NPC. Therefore, it is important to seek for innovative therapeutics targeting Jab1/CSN5 for NPC. In this study, we explored the antitumor effect of a curcumin analogue T83 in NPC, and found T83 exhibits antitumor activity and induces radiosensitivity through inactivation of Jab1 in NPC.
NPC cell viability and proliferation were detected by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation assays. Cell cycle distribution was detected with use of flow cytometry. Apoptosis was examined by using the Annexin V/propidium iodide staining assay and cleavage poly(ADP-ribose polymerase (PARP) and cleavage caspase-3 expression. Jab1 expression was examined by Western blotting.
A growth inhibitory effect was observed with T83 treatment in a dose- and time-dependent manner. T83 significantly induced G2/M arrest and apoptosis in NPC. In addition, T83 inhibited Jab1 expression and sensitized NPC cells to radiotherapy.
Our data indicate that T83 exhibits potent inhibitory activity in NPC cells and induces radiotherapy sensitivity. Thus, T83 has translational potential as a chemopreventive or therapeutic agent for NPC.
[Show abstract][Hide abstract] ABSTRACT: The aim of the present study was to evaluate the antitumor effects of the curcumin analogue GL63 on radioresistant nasopharyngeal carcinoma (NPC) CNE2R cells and parental CNE2 cells. The cell viability and proliferation of NPC cells were detected by MTT assay and colony formation assay. The suppressive effect on tumor growth was examined using in vivo subcutaneously inoculated NPC tumor models using nude mice. The cell cycle distribution was detected using flow cytometry. Apoptosis was examined by Hoechst 33342 and Annexin V/PI staining assay. The protein expression of endoplasmic reticulum (ER) stress pathway markers, XBP-1, ATF-4 and CHOP, were examined by western blotting. A growth inhibitory effect was observed following treatment with GL63 in a dose-dependent manner and was more potent when compared to curcumin. GL63 at 5 µM induced significant G2/M arrest and apoptosis in NPC. The tumor-suppressive activity of GL63 in NPC xenograft models was more potent when compared to curcumin. Furthermore, GL63 induced an ER stress response, upregulation of CHOP, XBP-1 and ATF-4 expression, while the same concentration of curcumin had no effect on ER stress. These results suggest that GL63 has more potent antitumor activity than curcumin, which is associated with activation of ER stress, induction of G2/M arrest and apoptosis in NPC cells.
[Show abstract][Hide abstract] ABSTRACT: Radioresistance is a major obstacle to the treatment of human nasopharyngeal carcinoma (NPC). Emerging evidence has demonstrated that miRNAs are involved in cancer therapy resistance. Our research group established the radioresistant NPC cell line CNE2R derived from the CNE2 cell line, and demonstrated that irradiation-induced miR-205 determined the resistance of NPC through directly targeting PTEN. However, specific inhibitors targeting miRNAs are largely undetermined. SZ-685C was expected to abrogate the radioresistance of CNE2 cells through the miR-205‑PTEN-Akt pathway. SZ-685C exhibited a similar cytotoxic effect on both cell lines, and we demonstrated that both intrinsic and extrinsic pathways were activated by SZ-685C in the cell lines. Importantly, the miR-205-PTEN-Akt pathway was the key cell signaling pathway activated in the CNE2R cells upon SZ-685C treatment; however, the Stat3-Jab1-p27 pathway might participate in the pro-apoptotic effect in CNE2 cells but not in CNE2R cells. SZ-685C is a promising anticancer agent for treatment of NPC, and it exhibited pro-apoptotic activity in both radiosensitive and radioresistant NPC cells. Although the mechanisms between the two cell lines were not identical, the pro-apoptotic effects were similar between the two cell lines.
[Show abstract][Hide abstract] ABSTRACT: Subunit 6 of the COP9 signalosome complex, CSN6, is known to be critical to the regulation of the MDM2-p53 axis for cell proliferation and anti-apoptosis, but its many targets remain unclear. Here we show that p57 (Kip2) is a target of CSN6, and that CSN6 is a negative regulator of p57 (Kip2) . CSN6 associates with p57 (Kip2) , and its overexpression can decrease the steady-state expression of p57 (Kip2) ; accordingly, CSN6 deficiency leads to p57 (Kip2) stabilization. Mechanistic studies show that CSN6 associates with p57 (Kip2) and Skp2, a component of the E3 ligase, which, in turn, facilitates Skp2-mediated protein ubiquitination of p57 (Kip2) . Loss of Skp2 compromised CSN6-mediated p57 (Kip2) destabilization, suggesting collaboration between Skp2 and CSN6 in degradation of p57 (Kip2) . CSN6's negative impact on p57 (Kip2) elevation translates into cell growth promotion, cell cycle deregulation and potentiated transformational activity. Significantly, univariate Kaplan-Meier analysis of tumor samples demonstrates that high CSN6 expression or low p57 expression is associated with poor overall survival. These data suggest that CSN6 is an important negative regulator of p57 (Kip2) , and that overexpression of CSN6 in many types of cancer could lead to decreased expression of p57 (Kip2) and result in promoted cancer cell growth.
[Show abstract][Hide abstract] ABSTRACT: HER2/neu oncogene is frequently overexpressed in various types of cancer, and the (PI3K)-Akt signaling pathway is often activated in HER2-overexpressing cancer cells. CSN6, subunit 6 of the COP9 signalosome complex, is pivotal in regulating MDM2 to destabilize p53, but its upstream regulators remain unclear. Here we show that the HER2-Akt axis is linked to CSN6 regulation, and that Akt is a positive regulator of CSN6. Ectopic expression of Akt can increase the expression of CSN6; accordingly, Akt inhibition leads to CSN6 destabilization. Mechanistic studies show that Akt causes CSN6 phosphorylation at Ser 60, which, in turn, reduces ubiquitin-mediated protein degradation of CSN6. Significantly, Akt's positive impact on CSN6 elevation translates into p53 degradation, potentiating transformational activity and increasing DNA damage. Akt inhibition can attenuate these defects caused by CSN6. These data suggest that Akt is an important positive regulator of CSN6, and that activation of Akt in many types of cancer could lead to abnormal elevation of CSN6 and result in downregulated p53 and increased DNA damage, which promotes cancer cell growth.
[Show abstract][Hide abstract] ABSTRACT: Endoplasmic reticulum (ER) stress-induced cancer cell apoptosis has become a novel signaling target for the development of therapeutic drugs for cancer treatment. Curcumin, a dietary phytochemical, exhibits growth-suppressive activity against cancer cells via multitarget mechanisms. However, the low stability and poor pharmacokinetics significantly limit its clinical applications. Thus, we designed and synthesized a novel monocarbonyl analog of curcumin, 1,5-bis(2-methoxyphenyl) penta-1,4-dien-3-one (B63). This compound exhibited a higher chemical stability in cultural medium and a better intracellular profile than curcumin. Treatment with B63 potently induced apoptosis of human non-small cell lung cancer (NSCLC) cells in a dose-responsive manner, while exhibiting no cytotoxicity in normal lung fibroblast cells. Its antitumor effect was associated with the ER stress-mediated apoptotic pathway and, ultimately, the activation of the caspase cascades. However, curcumin at the same concentrations did not cause ER stress in H460 cells. Further, C/EBP homologous protein knockdown by siRNA attenuated B63-induced cell apoptosis, indicating that the apoptotic pathway is ER stress-dependent. In vivo, the volume and weight of the tumor were reduced significantly by pretreating the H460 tumor cells with B63 before implantation. Taken together, these insights on the novel compound B63, from both chemical and biological perspectives, may provide a novel anticancer candidate for the treatment of NSCLC.
Full-text · Article · Sep 2012 · International Journal of Cancer
[Show abstract][Hide abstract] ABSTRACT: Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus-associated malignancy most common in East Asia and Africa. Aberrant expression of Jab1/CSN5, a negative regulator of the cell-cycle inhibitor p27, is correlated with reduced p27 expression and associated with advanced tumor stage and poor prognosis in several human cancers. In this study, we examined the functional relationship between Jab1 and p27 protein expression in NPC. Immunohistochemical analysis showed an inverse association between Jab1 and p27 in NPC tissue samples, and overexpression of Jab1 correlated with poor survival in patients with NPC. Mechanistically, Jab1 and p27 were found to interact directly in NPC cells, with Jab1 mediating p27 degradation in a proteasome-dependent manner. Knockdown of Jab1 resulted in a remarkable increase in p27 levels and inhibition of cell proliferation, indicating that Jab1 targets p27 for degradation, thereby controlling its stability. Jab1 depletion also enhanced the antitumor effects of cisplatin in NPC cells. Together, our findings suggest that Jab1 overexpression plays an important role in the pathogenesis of NPC through Jab1-mediated p27 degradation. Jab1 therefore represents a novel diagnostic marker and therapeutic target in patients with NPC.
[Show abstract][Hide abstract] ABSTRACT: Radiotherapy is the primary treatment for nasopharyngeal carcinoma (NPC), but radioresistance severely reduces NPC radiocurability. Here, we have established a radio-resistant NPC cell line, CNE-2R, and investigate the role of miRNAs in radioresistance. The miRNAs microarray assay reveals that miRNAs are differentially expressed between CNE-2R and its parental cell line CNE-2. We find that miR-205 is elevated in CNE-2R. A target prediction algorithm suggests that miR‑205 regulates expression of PTE N, a tumor-suppressor. Introducing miR-205 into CNE-2 cells suppresses PTE N protein expression, followed by activation of AKT, increased number of foci formation and reduction of cell apoptosis postirradiation. On the other hand, knocking down miR-205 in CNE-2R cells compromises the inhibition of PTE N and increases cell apoptosis. Significantly, immunohistochemistry studies demonstrate that PTE N is downregulated at late stages of NPC, and that miR-205 is significantly elevated followed the radiotherapy. Our data conclude that miR-205 contributes to radioresistance of NPC by directly targeting PTE N. Both miR-205 and PTE N are potential predictive biomarkers for radiosensitivity of NPC and may serve as targets for achieve successful radiotherapy in NPC.
[Show abstract][Hide abstract] ABSTRACT: Lung cancer is a leading cause of morbidity and mortality worldwide. Cyclooxygenase-2 (COX-2) expression is upregulated in lung carcinomas and is considered an attractive therapeutic target. In this study, the effect of curcumin and curcumin analogues on COX-2 expression induced by phorbol 12-myristate 13-acetate (PMA) were investigated. We found that a novel curcumin analogue (GL63) inhibited PMA-induced COX-2 mRNA and protein levels in H460 cells to a greater degree than curcumin. To understand the molecular mechanisms governing COX-2 regulation, the effect on COX-2 mRNA degradation was examined; we found that GL63 significantly decreased COX-2 mRNA stability by reducing cytoplasmic localization and protein abundance of human antigen R (HuR). The 3'-untranslated region (3'-UTR) report gene assay also showed GL63 substantially reduced the 3'-UTR green fluorescent protein values, indicating that the destabilizing effect on COX-2 mRNA may be couple with the posttranscriptional inhibition of COX-2. Taken together, our results provide evidence that the novel curcumin analogue can effectively inhibit PMA-induced COX-2 expression in H460 cells, a mechanism associated with COX-2 mRNA stability and post-transcriptional regulation.
[Show abstract][Hide abstract] ABSTRACT: Cell cycle dysregulation is a critical feature of tumor cells. Numerous cell cycle regulators act either as oncogenes or tumor suppressors and their aberrations result in proliferative advantage for cancer cells. Many molecular targets and their use in either abrogating the growth advantage of oncogenic mediators or enhancing the growth suppressive activity of tumor suppressors, have been filed for patents. The molecular targets associated with cell cycle inhibition are of particular interest because they are potential therapeutic agents of promise in the control of inappropriate cellular proliferation. This review focuses on the recent discovery of potential molecular targets involved in cell cycle inhibition and their evaluation as therapeutic agents for cancers. In this review, the strategies employed to control oncogenesis and their possible clinical applications are discussed.
[Show abstract][Hide abstract] ABSTRACT: Anaplastic thyroid cancer (ATC) is extremely aggressive, and no effective treatment is available. Combretastatin A4 phosphate (CA4P), a vascular disrupting agent, has limited activity against ATC in a clinical trial, and so does paclitaxel.
We hypothesized that a triple-drug combination including CA4P and paclitaxel would improve efficacy against ATC. Therefore, we evaluated two such combinations in vivo.
We used a nude mouse xenograft model with ARO and KAT-4 cells.
The first combination consisted of CA4P, paclitaxel, and manumycin A (a farnesyltransferase inhibitor), and the second, CA4P, paclitaxel, and carboplatin.
Main outcome measures included tumor growth curves and tumor weights.
Tumor growth curve analysis (linear mixed models, P < 0.05) and xenograft weight analysis (Kruskal-Wallis one-way ANOVA on ranks, post hoc pairwise comparison, Dunn's test, P < 0.05) demonstrated that both triple-drug combinations were significantly better than placebo for both cell lines. Anti-bromodeoxyuridine immunostaining of xenograft sections from animals injected with bromodeoxyuridine before being killed showed that CA4P alone did not inhibit DNA synthesis, but manumycin A and paclitaxel did. CA4P decreased the depth of the viable outer rim of tumor cells on xenograft sections. Using electron microscopy, we found blebbing/budding of endothelial cells into capillary lumens and autophagy of tumor cells in CA4P-treated xenografts.
Both triple-drug combinations demonstrated excellent antineoplastic activity against ATC. The correlative findings in xenografts were consistent with vascular disruption but not direct inhibition of cell proliferation as the primary antineoplastic mechanism contributed by CA4P. These regimens warrant further investigation in clinical trials for ATC.
Full-text · Article · Aug 2007 · Journal of Clinical Endocrinology & Metabolism
[Show abstract][Hide abstract] ABSTRACT: Cyclin-dependent kinase (CDK) inhibitor p27 Kip1, a haplo-insufficient tumor suppressor, is downregulated by oncogenic signal of HER2, a receptor tyrosine kinase oncogene. HER2 promotes mitogenic growth and transformation of cancer cells. HER2 signaling can enhance p27 Kip1 ubiquitination, thereby promoting p27 degradation and subsequent activation of CDK activity. p27 ubiquitination and degradation is enhanced by JAB1 binding as well as by phosphorylation on Thr187. In this study, we generated modified p27 proteins, which are mutated at Thr 187 or deleted at JAB1 binding domain. We applied these modified p27 genes as novel anticancer agents for HER2-overexpressing cells under the control of a tetracycline (tet)-regulated gene expression system. Induction of p27 T187A and p27 T187A DeltaJAB inhibits HER2-activated cell growth, CDK2 activity, cell proliferation, and transformation. Significantly, a modified protein (p27 T187ADeltaJAB) reduced the tumor volume in a HER2-overexpressing tumor model efficiently. These findings demonstrate the applicability of employing modified p27 proteins as a therapeutic intervention in HER2-overexpressing cancers.
No preview · Article · May 2006 · Journal of Cellular Biochemistry
[Show abstract][Hide abstract] ABSTRACT: 14-3-3 sigma is induced by tumor suppressor protein p53 in response to DNA damage. p53 can directly transactivate the expression of 14-3-3 sigma to cause a G(2) cell cycle arrest when cell DNA is damaged. The expression of 14-3-3 sigma protein is down-regulated in various tumors, but its function has not been fully established. Protein kinase B/Akt, a crucial regulator of oncogenic signal involved in cell survival and proliferation, is deregulated in many types of cancer. Akt activation can enhance p53 degradation, but its role in DNA damage response is not clear. Here, we show that Akt activation is diminished when p53 and 14-3-3 sigma is up-regulated in response to DNA damage. Evidence is provided that 14-3-3 sigma binds and inhibits Akt. In keeping with this concept, Akt-mediated cell survival is inhibited by 14-3-3 sigma. Significantly, we show that 14-3-3 sigma inhibits Akt-mediated cell growth, transformation, and tumorigenesis. Low expression of 14-3-3 sigma in human primary breast cancers correlates with Akt activation. These data provide an insight into Akt regulation and rational cancer gene therapy by identifying 14-3-3 sigma as a molecular regulator of Akt and as a potential anticancer agent for Akt-activated cancers.
[Show abstract][Hide abstract] ABSTRACT: Our previous studies demonstrated that manumycin A, a farnesyltransferase inhibitor, induced apoptosis of anaplastic thyroid cancer cells via the intrinsic apoptosis pathway and induced reactive oxygen species (ROS), which mediated DNA damage. In this study, we investigated the hypothesis that the mechanism of apoptosis induced by manumycin in anaplastic thyroid cancer cells fits the general pattern of the "xenobiotic apoptosis pathway," the hallmarks of which are induction of oxidative stress, mitogen-activated protein kinase (MAPK) signaling, and cytochrome c release, which activates the intrinsic apoptosis pathway. We found that manumycin reduced intracellular glutathione and generated ROS: nitric oxide and superoxide anions. Manumycin-induced apoptosis correlated with increase in ROS. Quenching of ROS with N-acetyl-L-cysteine prevented cytochrome c release by manumycin. Manumycin induced phosphorylation of p38 MAPK, which was blocked by N-acetyl-L-cysteine. p38 MAPK may be an important signaling mediator in the activation of the intrinsic apoptotic pathway by manumycin because the p38 MAPK inhibitor SB203580 inhibited cytochrome c release and activation of caspase-3 by manumycin. In conclusion, manumycin activated the intrinsic apoptosis pathway via activation of p38 MAPK by oxidative stress. The mechanism of apoptosis induced by manumycin fits the emerging general pattern for apoptosis induced by xenobiotics.