Corrigendum: Enhanced Cytotoxic Effects of Combined Valproic Acid and the Aurora Kinase Inhibitor VE465 on Gynecologic Cancer Cells.

[This corrects the article on p. 58 in vol. 3, PMID: 23519775.].

Enhanced Cytotoxic Effects of Combined Valproic Acid and the Aurora Kinase Inhibitor VE465 on Gynecologic Cancer Cells.

Increasing evidence shows that targeting epigenetic changes including acetylation and deacetylation of core nucleosomal histones as well as Aurora kinases hold promise for improving the treatment of human cancers including ovarian cancer. We investigated whether the histone deacetylase (HDAC) inhibitor, valproic acid (VPA), and the Aurora kinase inhibitor VE465 can have additive or synergistic effects on gynecologic cancer cells. We tested the in vitro antitumor activity of VPA and VE465, alone and in combination, in gynecologic cancer cells and assessed potential mechanisms of action. 3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide (MTT) analysis revealed that 72 h of treatment with VPA or VE465 alone induced dose-dependent cytotoxic effects in nine gynecologic cancer cell lines (ovarian: 2008/C13, OVCAR3, SKOV3, and A2780; cervical: ME180 and CaSki; endometrial: HEC-1B; and uterine sarcoma: MES-SA and MES-SA/D×5). Co-treatment with VPA and VE465 enhanced cytotoxic effects on five of these cell lines: ovarian: 2008/C13, A2780, and OVCAR3; endometrial: HEC-1B; and cervical: ME180. In ovarian 2008/C13 cells, co-treatment with VPA (2 mM) and VE465 (1 µM) induced more apoptosis than either VPA or VE465 alone. Western blot analysis showed that VPA alone increased the expression of cleaved PARP and p21 in a dose-dependent manner in 2008/C13 cells, while co-treatment with VPA and VE465 induced more cleaved PARP than treatment with VPA or VE465 alone did. The combined use of VPA and VE465 enhanced cytotoxic effects in some ovarian cancer cells, via enhanced induction of apoptosis. Targeting epigenetics with the HDAC inhibitor, in combination with Aurora kinase inhibitors, holds promise for more effective therapy of ovarian cancer.

Biological roles of the Delta family Notch ligand Dll4 in tumor and endothelial cells in ovarian cancer.

Emerging evidence suggests that the Notch/Delta-like ligand 4 (Dll4) pathway may offer important new targets for antiangiogenesis approaches. In this study, we investigated the clinical and biological significance of Dll4 in ovarian cancer. Dll4 was overexpressed in 72% of tumors examined in which it was an independent predictor of poor survival. Patients with tumors responding to anti-VEGF therapy had lower levels of Dll4 than patients with stable or progressive disease. Under hypoxic conditions, VEGF increased Dll4 expression in the tumor vasculature. Immobilized Dll4 also downregulated VEGFR2 expression in endothelial cells directly through methylation of the VEGFR2 promoter. RNAi-mediated silencing of Dll4 in ovarian tumor cells and tumor-associated endothelial cells inhibited cell growth and angiogenesis, accompanied by induction of hypoxia in the tumor microenvironment. Combining Dll4-targeted siRNA with bevacizumab resulted in greater inhibition of tumor growth, compared with control or treatment with bevacizumab alone. Together, our findings establish that Dll4 plays a functionally important role in both the tumor and endothelial compartments of ovarian cancer and that targeting Dll4 in combination with anti-VEGF treatment might improve outcomes of ovarian cancer treatment.

Nuclear cyclin B1 is overexpressed in low-malignant-potential ovarian tumors but not in epithelial ovarian cancer.

OBJECTIVE: To investigate the role of cyclin G1 and cyclin B1 in ovarian tumorigenesis. STUDY DESIGN: We examined cyclin B1 and G1 expression in 58 epithelial ovarian cancer, 18 low-malignant-potential ovarian tumors, and 6 normal ovarian epithelium samples using immunohistochemistry. We also examined cyclin G1 and p53 expression in 7 epithelial ovarian cancer cell lines using Western blot analysis. RESULTS: Nuclear cyclin B1 expression was significantly higher in low-malignant-potential tumors than in normal ovarian epithelium. There was no difference in nuclear or cytoplasmic cyclin B1 or cyclin G1 expression between epithelial ovarian cancer and normal ovarian epithelium. Cyclin G1 and B1 expression was not associated with p53 expression or clinicopathologic features in patients with epithelial ovarian cancer or low-malignant-potential tumors. CONCLUSION: Our data demonstrated that nuclear cyclin B1 is overexpressed in low-malignant-potential tumors, which may contribute to the development of low-malignant-potential tumors. Cyclin B1 and G1 may not be suitable targets for epithelial ovarian cancer treatment.

Azacitidine enhances sensitivity of platinum-resistant ovarian cancer cells to carboplatin through induction of apoptosis.

OBJECTIVE: The objective of the study was to investigate whether azacitidine sensitizes platinum-resistant ovarian cancer cells to carboplatin and the possible mechanisms involved. STUDY DESIGN: We tested the in vitro antitumor activity of azacitidine both alone and combined with carboplatin in the ovarian cancer cell line 2008/C13 and Hey by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assays and investigated the potential mechanisms by flow cytometry, terminal transferase deoxyuridine 5-triphosphate nick-end labeling assay, Western blot, reverse transcriptase-polymerase chain reaction (PCR), and promoter methylation-specific PCR. RESULTS: Sequential treatment (ie, 24-hour azacitidine pretreatment followed by 48-hour cotreatment with azacitidine and carboplatin) significantly inhibited growth in 2008/C13 and Hey cells. More apoptotic cells were induced in 2008/C13 cells by sequential treatment than by a single drug. Increased cleaved caspase-3 and -8 were seen in 2008/C13 cells after sequential treatment with azacitidine and carboplatin. DR4 was demethylated, and DR4 messenger ribonucleic acid expression was increased in 2008/C13 cells after the 24-hour azacitidine treatment. CONCLUSION: Azacitidine enhanced the sensitivity of platinum-resistant ovarian cancer cells to carboplatin associated with caspase-3- and -8-dependent apoptosis pathway and reexpression of DR4.

Diammine dicarboxylic acid platinum enhances cytotoxicity in platinum-resistant ovarian cancer cells through induction of apoptosis and S-phase cell arrest.

PURPOSE: Polysaccharides such as chondroitin play a potent role in tumor growth, tissue repair and angiogenesis. These properties make chondroitin a good candidate for novel drug delivery systems. Diammine dicarboxylic acid platinum (DDAP), a novel polymeric platinum compound, was developed by conjugating the platinum analogue to aspartate-chondroitin for drug delivery to tumor cells. DDAP improves platinum solubility which may reduce systemic toxicity and be more efficacious than cisplatin in killing tumor cells. METHODS: We tested and compared the cytotoxic effects of DDAP and CDDP on the platinum-sensitive 2008 and A2780 ovarian cancer cell lines and their platinum-resistant sublines 2008.C13 and A2780cis; we also investigated DDAP's mechanism of action. RESULTS: In the platinum-sensitive cell lines, the cytotoxic effects of DDAP and CDDP were comparable. However, in the platinum-resistant sublines, significantly greater cell-growth inhibition was induced by DDAP than by CDDP, especially at lower doses. DDAP also induced more apoptosis than CDDP did in the 2008.C13 subline, which was partially mediated by the caspase 3-dependent pathway. In addition, lower (but not higher) doses of DDAP arrested 90% of S-phase 2008.C13 cells, which might be associated with up-regulation of p21 and maintenance of low cyclin A expression. Furthermore, greater cellular uptake of DDAP was seen in platinum-resistant than in platinum-sensitive ovarian cancer cells. CONCLUSIONS: Low-dose DDAP enhances drug delivery to platinum-resistant ovarian cancer cells and substantially inhibits their growth by inducting apoptosis and arresting cells in the S-phase, suggesting that DDAP may overcome platinum resistance in ovarian cancer.

Deficiencies in myeloid antigen-presenting cells in women with cervical squamous intraepithelial lesions.

BACKGROUND: There is little information on the function of dendritic cells in women with human papillomavirus (HPV)-related cervical squamous intraepithelial lesions (SILs). In the current study the functions of dendritic cells in the development of T-cell immunity in women with cervical SILs were assessed. METHODS: The percentage of myeloid dendritic cells (MDCs) and plasmacytoid dendritic cells (PDCs) in peripheral blood were enumerated of 44 patients with SIL (low-grade, 19; high-grade, 25), 19 patients with atypical squamous cells of undetermined significance (ASCUS), and 18 controls. The expression of costimulatory receptors was assessed and the ability of monocyte-derived dendritic cells (MDDC) to present HPV16-E6 and HPV16-E7 antigens to autologous T cells. RESULTS: Patients with either low (L)-grade or high (H)-grade SIL had significantly lower median plasma levels of interferon-gamma than did the controls (P = .038 and .031, respectively). Compared with the controls, patients with ASCUS or LSILs had significantly lower median percentages of MDCs (P = .002 and P < .001, respectively), and significantly lower median percentages of MDDCs that expressed CD86 (P < .001 and P = .003, respectively) and major histocompatability complex class-II antigen human leukocyte antigen DR (HLA-DR) (P = .012 and P < .001, respectively). T cells of patients with ASCUS or LSILs proliferated less than those of the controls in response to HPV16-E7 (P = .002 and .046, respectively). CONCLUSIONS: Low levels of peripheral blood MDCs and of MDDCs expressing CD86 and HLA-DR suggest that deficiencies in the ability of MDDC to present antigen to autologous T cells may lead to persistent infection with HPV and the development of cervical SILs in HPV-infected women.

Depressed type 1 cytokine synthesis by superantigen-activated CD4+ T cells of women with human papillomavirus-related high-grade squamous intraepithelial lesions.

Carcinoma of the cervix is causally related to infection with the human papillomavirus (HPV), and T cells play a pivotal role in the immune response of the host to rid itself of HPV infection. Therefore, we assessed the T-cell function of women with HPV-related cervical neoplasia against a superantigen, Staphylococcus enterotoxin B (SEB). Each woman provided a cervical brush specimen for HPV DNA testing and Papanicolaou (Pap) smears for the staging of cervical lesions. They also provided a blood specimen for determination of the ability of CD4(+) T and CD8(+) T cells to synthesize Th1 (interleukin-2 [IL-2], gamma interferon [IFN-gamma], and tumor necrosis factor alpha [TNF-alpha]) and Th2 (IL-10) cytokines in response to activation with SEB. Compared with control subjects with self-attested negative Pap smears, women with high-grade squamous intraepithelial lesions (HSIL) had significantly lower percentages of activated CD4(+) T cells that produced IL-2 (P = 0.045), IFN-gamma (P = 0.040), and TNF-alpha (P = 0.015) and a significantly lower percentage of activated CD8(+) T cells that produced IL-2 (P < 0.01). These data indicate that women with HPV-related cervical HSIL show a decrease in Th1 cytokine production by activated CD4(+) T cells and suggested that compromised T-helper functions may negatively impact the function of cytotoxic CD8(+) T cells.