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Budiman Kharma,
Tsukasa Baba,
Masaki Mandai,
Noriomi Matsumura,
Susan K Murphy,
Hyun Sook Kang,
Koji Yamanoi,
Junzo Hamanishi, Ken Yamaguchi,
Yumiko Yoshioka,
Ikuo Konishi
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ABSTRACT: Objectives: Endometrial cancer, one of the most common gynecologic malignancies, is increasing in Japan, nearly doubling over the last decade. High grade disease patients are often resistant to conventional chemotherapy with platinum agents; therefore discovery of efficacious new drugs in this setting is required to benefit chemo-refractory cases. Methods: The 50% Growth-Inhibitory (GI50) concentration of 27 clinically relevant drugs was measured in the NCI60 panel of cell lines. Gene expression data were analyzed using Bayesian binary regression, to first generate a response signature for each drug, then to calculate individual susceptibility scores using in vivo endometrial cancer data (GSE2109; http://www.ncbi.nlm.nih.gov/geo) and in vitro data (GSE25458), as well as to identify candidate drugs for chemo-refractory cases. Using these candidates, cell proliferation, apoptosis, and caspase assays were performed in vitro. The tumor growth-inhibitory effect of the candidate was also assessed in vivo using nude mice. Results: Through microarray analysis, Fludarabine and Temsirolimus showed higher susceptibility scores in high grade cases compared with cisplatin, doxorubicin, and paclitaxel. Fludarabine significantly inhibited cell proliferation and increased apoptosis in the cisplatin-resistant endometrial cancer cell line, HEC1A, relative to HEC50B (p<0.001). Fludarabine treatment also enhanced Caspase 3/7 activity in HEC1A relative to HEC50B cells (p<0.001), and inhibited the growth of HEC1A xenograft tumors relative to cisplatin (p<0.05). Conclusions: These results support that identification and use of genomic signatures can lead to identification of new therapeutic candidates that may prove beneficial to chemo-resistant cases. Fludarabine may be useful in targeting high grade, chemo-refractory endometrial cancer. © 2013 Wiley Periodicals, Inc.
International Journal of Cancer 04/2013; · 5.44 Impact Factor
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Kaoru Abiko,
Masaki Mandai,
Junzo Hamanishi,
Yumiko Yoshioka,
Noriomi Matsumura,
Tsukasa Baba, Ken Yamaguchi,
Ryusuke Murakami,
Ayaka Yamamoto,
Budiman Kharma,
Kenzo Kosaka,
Ikuo Konishi
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ABSTRACT: PURPOSE: Ovarian cancer often progresses by disseminating to the peritoneal cavity, but how the tumor cells evade host immunity during this process is poorly understood. Programmed cell death 1 ligand 1 (PD-L1) is known to suppress immune system and to be expressed on cancer cells. The purpose of this study is to elucidate the function of PD-L1 in peritoneal dissemination. EXPERIMENTAL DESIGN: Ovarian cancer cases were studied by microarray and immunohistochemistry. PD-L1 expression in mouse ovarian cancer cell line in various conditions was assessed by flow cytometry. PD-L1-overexpression cell line and PD-L1-depleted cell line were generated, and cytolysis by CTLs was analyzed, and alterations in CTLs were studied by means of timelapse and microarray. These cell lines were injected intraperitoneally to syngeneic immunocompetent mice. RESULTS: Microarray and immunohistochemistry in human ovarian cancer revealed significant correlation between PD-L1 expression and peritoneal positive cytology. PD-L1 expression on mouse ovarian cancer cells was induced upon encountering lymphocytes in the course of peritoneal spread in vivo and co-culture with lymphocytes in vitro. Tumor cell lysis by CTLs was attenuated when PD-L1 was overexpressed and promoted when it was silenced. PD-L1 overexpression inhibited gathering and degranulation of CTLs. Gene expression profile of CTLs caused by PD-L1-overexpressing ovarian cancer was associated with CTLs exhaustion. In mouse models, PD-L1 depletion resulted in inhibited tumor growth in the peritoneal cavity and prolonged survival. CONCLUSIONS: PD-L1 expression on tumor cell promotes peritoneal dissemination by repressing CTL function. PD-L1-targeted therapy is a promising strategy for preventing and treating peritoneal dissemination.
Clinical Cancer Research 01/2013; · 7.74 Impact Factor
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ABSTRACT: Although it is well known that ovarian endometriosis occasionally gives rise to ovarian cancers with specific histology such as endometrioid and clear cell carcinomas, its etiology is not fully understood. We have shown that a stressful microenvironment within the endometriotic cyst may lead to cancer development by inducing unique gene expressions, which potentially serves as a molecular marker for treatment modality. In this review, by referring to other articles in this field, we explore how the carcinogenic microenvironment affects the phenotype and gene expression of a cancer, and how we can develop new treatment based on this concept.
Cancer letters 11/2011; 310(2):129-33. · 4.86 Impact Factor
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Shogo Yamamura,
Noriomi Matsumura,
Masaki Mandai,
Zhiqing Huang,
Tomonori Oura,
Tsukasa Baba,
Junzo Hamanishi, Ken Yamaguchi,
Hyun Sook Kang,
Takako Okamoto,
Kaoru Abiko,
Seiichi Mori,
Susan K Murphy,
Ikuo Konishi
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ABSTRACT: Peritoneal dissemination including omental metastasis is the most frequent route of metastasis and an important prognostic factor in advanced ovarian cancer. We analyzed the publicly available microarray dataset (GSE2109) using binary regression and found that the transforming growth factor (TGF)-beta signaling pathway was activated in omental metastases as compared to primary sites of disease. Immunohistochemical analysis of TGF-beta receptor type 2 and phosphorylated SMAD2 indicated that both were upregulated in omental metastases as compared to primary disease sites. Treatment of the mouse ovarian cancer cell line HM-1 with recombinant TGF-β1 promoted invasiveness, cell motility and cell attachment while these were suppressed by treatment with A-83-01, an inhibitor of the TGF-β signaling pathway. Microarray analysis of HM-1 cells treated with TGF-β1 and/or A-83-01 revealed that A-83-01 efficiently inhibited transcriptional changes that are induced by TGF-β1. Using gene set enrichment analysis, we found that genes upregulated by TGF-β1 in HM-1 cells were also significantly upregulated in omental metastases compared to primary sites in the human ovarian cancer dataset, GSE2109 (false discovery rate (FDR) q = 0.086). Therapeutic effects of A-83-01 in a mouse model of peritoneal dissemination were examined. Intraperitoneal injection of A-83-01 (150 μg given three times weekly) significantly improved survival (p = 0.015). In summary, these results show that the activated TGF-β signaling pathway in peritoneal metastases is a potential therapeutic target in ovarian cancer.
International Journal of Cancer 02/2011; 130(1):20-8. · 5.44 Impact Factor
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Noriomi Matsumura,
Masaki Mandai,
Takako Okamoto, Ken Yamaguchi,
Shogo Yamamura,
Tomonori Oura,
Tsukasa Baba,
Junzo Hamanishi,
Hyun S Kang,
Shigeyuki Matsui,
Seiichi Mori,
Susan K Murphy,
Ikuo Konishi
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ABSTRACT: The purpose of this study was to investigate a new modality of therapy against ovarian clear cell carcinoma (OCCC), a chemoresistant subtype of ovarian cancer. Microarray datasets of ovarian cancer cell lines and cancer tissues were analyzed using bioinformatic tools. The gene expression profile of OCCC was similar to that of renal cell carcinoma (RCC). This similarity was at least partially due to hepatocyte nuclear factor 1 pathway activation common to both malignancies. In addition, oncogenic pathway alterations were characteristic of OCCC including hypoxia inducible factor 1 alpha subunit and relatively high Ras activities. Therefore, we predicted that the multi-kinase inhibitor sorafenib, which is approved for RCC and suppresses Ras activity, would also be effective against OCCC. Orally administered sorafenib (40 mg/kg per day) significantly inhibited tumor growth in nude mice when it was given after inoculation with the OCCC cell line RMG-2 (P = 0.002). Furthermore, sorafenib significantly reduced tumor size when it was administered to established RMG-2 tumors (P = 0.0002), while intraperitoneal injection of cisplatin (5 mg/kg per week) did not. In conclusion, the prominent anti-tumor effect of sorafenib against OCCC indicates that sorafenib is a promising candidate drug and supports the need for clinical trials using sorafenib against OCCC. This report demonstrates a method to utilize genome-wide information to facilitate translational research for treatments against less common subtypes of cancers.
Cancer Science 09/2010; 101(12):2658-63. · 3.33 Impact Factor
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ABSTRACT: Although tumor microenvironments play a key role in successful tumor immunotherapy, effective manipulation of local immunity is difficult because of the lack of an appropriate target system. It is well known that bone marrow-derived endothelial progenitor cells (EPCs) are actively recruited during tumor angiogenesis. Using this feature, we attempted to establish a novel therapeutic modality that targets tumor vessels of multiple metastases using embryonic endothelial progenitor cells (eEPCs) transduced with an immune-activating gene. The eEPCs were retrovirally transduced with the mouse CC chemokine ligand 19 (CCL19) gene, a lymphocyte-migrating chemokine. The mouse ovarian cancer cell line OV2944-HM-1 (HM-1) was inoculated subcutaneously into B6C3F1 mice, along with CCL19-tranduced eEPCs (eEPC-CCL19), resulting in immunologic activity and tumor-inhibitory effects. In this model, eEPC-CCL19 showed tumor repression accompanied by increased tumor-infiltrating CD8+ lymphocytes compared with the control group. In contrast, no tumor repression was observed when the same experiment was done in immunodeficient (SCID) mice, suggesting a crucial role of T-cell function in this system. Next, we established a lung metastasis model by injecting HM-1 cells or B16 melanoma cells via the tail vein. Subsequent intravenous injection of eEPC-CCL19 leads to a decrease in the number of lung metastasis and prolonged survival. Antitumor effects were also observed in a peritoneal dissemination model using HM-1. These results suggest that systemic delivery of an immune-activating signal using EPCs can alter the tumor immune microenvironment and lead to a therapeutic effect, which may provide a novel strategy for targeting multiple metastases of various malignancies.
Stem Cells 11/2009; 28(1):164-73. · 7.78 Impact Factor
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ABSTRACT: Recent molecular and pathological evidence suggests that endometriosis is a monoclonal, neoplastic disease. Moreover, endometriosis serves as a precursor of ovarian cancer (endometriosis-associated ovarian cancer; EAOC), especially of the endometrioid and clear cell subtypes. Although a variety of molecular events, such as p53 alteration, PTEN silencing, K-ras mutations, and HNF-1 activation, have been identified in EAOC, its precise carcinogenic mechanism remains poorly understood. Our recent data indicate that microenvironmental factors, including oxidative stress and inflammation, play an important role in the carcinogenesis and phenotype of EAOC. The management of endometriosis from the standpoint of EAOC is not standardized yet. To this end, clarification of the precise natural course and the risk factors that contribute to malignant transformation remain important goals. Among the phenotypes of EAOC, clear cell carcinoma, seems to require a specific treatment strategy, including molecular targeting.
International Journal of Clinical Oncology 10/2009; 14(5):383-91. · 1.41 Impact Factor
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Eiji Kondoh,
Seiichi Mori, Ken Yamaguchi,
Tsukasa Baba,
Noriomi Matsumura,
J Cory Barnett,
Regina S Whitaker,
Ikuo Konishi,
Shingo Fujii,
Andrew Berchuck,
Susan K Murphy
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ABSTRACT: Advanced ovarian cancer has a high rate of recurrence and mortality despite relative chemosensitivity at the time of initial treatment. Conventional chemotherapeutic agents typically target rapidly dividing cells. Disease relapse may therefore result from the survival and later emergence of latent slow-proliferating and/or quiescent cancer cells. We sought to identify drugs that target this cell population and to investigate the influence of these cells on outcome of patients in remission from advanced ovarian cancer. Drugs with increased efficacy against slower proliferating cells were identified using correlation-based screening of 44,657 compounds tested on the NCI-60 panel of cancer cell lines. Validation of candidates was performed in comparison with Cisplatin or Paclitaxel and by manipulation of proliferation rates by serum deprivation. Cytostatic and cytocidal effects were evaluated using MTT assays and active caspase-3 immunocytochemistry. Ki-67 proliferation indices were determined for tumors from 104 patients in remission. UCN-01 efficacy was correlated with longer doubling time among the NCI-60 cell lines (R = 0.54, p < 0.0001) and in a panel of 24 ovarian cancer cell lines (R = 0.42, p = 0.04), whereas this was not the case for Cisplatin (R = -0.10, p = 0.65) and Paclitaxel efficacy correlated with shorter doubling time (R = -0.52, p = 0.009). Cytostatic and cytocidal effects of UCN-01 were increased in serum-deprived cells. A low proliferation index was associated with presence of persistent disease at second-look surgery (p = 0.01) and poor survival (disease-free survival, p = 0.002; overall survival, p = 0.04). These results suggest that targeting quiescent ovarian cancer cells may be a worthwhile therapeutic approach to improving survival of women with ovarian cancer.
International Journal of Cancer 09/2009; 126(10):2448-56. · 5.44 Impact Factor
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ABSTRACT: To investigate the clinical significance of the expression of the NKG2D ligands MICA/B and ULBP2 in ovarian cancer.
Eighty-two ovarian cancer patients and six patients without ovarian cancer from Department of Obstetrics and Gynecology of Kyoto University Hospital were enrolled in this study between 1993 and 2003. Expression of MICA/B, ULBP2, and CD57 in ovarian cancer tissue and normal ovary tissue was evaluated by immunohistochemical staining, and the relationship of these results to relevant clinical patient data was analyzed. Expression of MICs, ULBP2, and HLA-class I molecules in 33 ovarian cancer cell lines and two normal ovarian epithelial cell lines, as well as levels of soluble MICs and ULBP2 in the culture supernatants, were measured.
Expression of MICA/B and ULBP2 was detected in 97.6 and 82.9% of ovarian cancer cells, respectively, whereas neither was expressed on normal ovarian epithelium. The expression of MICA/B in ovarian cancer was highly correlated with that of ULBP2. Strong expression of ULBP2 in ovarian cancer cells was correlated with less intraepithelial infiltration of T cells and bad prognoses for patients, suggesting that ULBP2 expression is a prognostic indicator in ovarian cancer. The expression of NKG2D ligands did not correlate with the levels of the soluble forms of the ligands.
High expression of ULBP2 is an indicator of poor prognosis in ovarian cancer and may relate to T cell dysfunction in the tumor microenvironment.
Cancer Immunology and Immunotherapy 10/2008; 58(5):641-52. · 3.70 Impact Factor
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ABSTRACT: The aim of this study was to seek for factors which lead to the early diagnosis of malignant transformation from mature cystic teratoma. Fourteen patients with malignant transformation from mature cystic teratoma of the ovary were analyzed retrospectively for precise clinicopathology and prognosis. The results demonstrated that although all the patients with stage Ia disease were disease-free, only 2 out of 7 patients were stage Ic to IV and disease-free in the follow-up period. Pre-operative imaging correctly diagnosed tumors as malignant in all stage Ic to IV cases, but only in 2 out of 4 stage Ia cases with magnetic resource and none of the 2 cases with computed tomography, respectively. In malignant cases, elevation of the serum SCC and CEA was observed in 90.9 and 88.9%, respectively. On the other hand, in benign cases, a false positive elevation of the serum SCC and CEA was observed in 23.5 and 14.3%, which turned out to be normal in 40 and 52.9% cases in the repeated study, respectively. In conclusion neither imaging analysis nor tumor markers including SCC and CEA accurately diagnose malignant transformation of mature cystic teratoma in its early stage, suggesting that a combination of diagnostic means is important. In the follow-up cases, repeated measurement of serum markers proved useful in ruling out false positive cases.
Oncology Reports 04/2008; 19(3):705-11. · 1.84 Impact Factor
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ABSTRACT: Systemic administration of Fms-like tyrosine kinase 3 ligand (FLT3L) has been considered to be a major route of delivery for tumor immunotherapy because expression of its receptor, FLT3, was detected predominantly in hematopoetic progenitor cells. However, several studies indicate that FLT3L locally overexpressed in tumor or dendritic cells (DCs) also shows an anti-tumor effect. In the current study, we found that FLT3 expression is not present in monocytes but is instead induced in DCs through the differentiation process resulting from stimulation by GM-CSF and IL-4. Addition of FLT3L further augmented FLT3 induction and also increased CD40 expression in DCs, leading to enhanced induction of lymphoblastoid cell line-targeted cytotoxic T-lymphocyte response and CD107a mobilization in CD8+ T cells. Furthermore, FLT3L also induced FLT3 expression in peripheral blood NK cells that showed an enhanced response detected by CD107a mobilization. In a murine ovarian cancer model, locally expressed FLT3L showed anti-tumor effects. Collectively, the current study indicates that FLT3L has an immunostimulatory effect on peripheral blood cells and FLT3L targeted to mature peripheral blood cells may serve as a useful tool for cancer immunotherapy.
Oncology Reports 03/2008; 19(2):505-15. · 1.84 Impact Factor
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ABSTRACT: Endometriotic cysts are known to transform into ovarian cancers, such as clear cell and endometrioid carcinomas. We hypothesized that an iron-rich environment produced by the repetition of hemorrhage in the endometriotic cysts during the reproductive period may play a crucial role in carcinogenesis in the cysts through the iron-induced persistent oxidative stress.
Contents of human ovarian cysts, including 21 endometriotic cysts, 4 clear cell carcinomas, and 11 nonendometriotic cysts, were analyzed for the concentrations of free "catalytic" iron, lactose dehydrogenase, potential antioxidant, lipid peroxide, and 8-hydroxy-2'-deoxyguanosine (8-OHdG). Iron deposition and 8-OHdG levels were also analyzed histologically. Reactive oxygen species and the mutagenicity of the contents in endometriotic cyst were determined in vitro.
The concentration of free iron in endometriotic cysts (100.9 mmol/L) was significantly higher than that in nonendometriotic cysts (0.075 mmol/L; P < 0.01). The average concentrations of lactose dehydrogenase, potential antioxidant, lipid peroxide, and 8-OHdG were also significantly higher in endometriotic cysts (P < 0.01). There was a correlation between the concentration of free iron and that of 8-OHdG (P < 0.01). Histologically, we could observe iron deposits more abundantly in endometriotic cysts than in nonendometriotic cysts (P < 0.01). The level of 8-OHdG in carcinoma associated with endometriosis was higher than that of carcinoma without endometriosis (P < 0.05). In vitro analyses showed that the contents of endometriotic cyst could produce more reactive oxygen species and could induce gene mutations more frequently than the contents in the other cysts.
Abundant free iron in the contents of endometriotic cysts was strongly associated with greater oxidative stress and frequent DNA mutations. A long-standing history of the RBCs accumulated in the ovarian endometriotic cysts during the reproductive period produces oxidative stress that is a possible cause for the malignant change of the endometriotic cyst.
Clinical Cancer Research 01/2008; 14(1):32-40. · 7.74 Impact Factor
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ABSTRACT: Malignant transformation is caused by multi-step genetic mutations, and growth factors are believed to play important roles in developing and maintaining malignant phenotype. However, there is no direct evidence that a specific growth factor contributes to malignant transformation of phenotypically normal cells. In order to assess the function of Acrogranin (also known as granulin epithelial precursor; GEP) in ovarian carcinogenesis, ovarian surface epithelial (OSE) cells, which are supposed to be the origin of primary ovarian epithelial cancer, were transfected with combined genes of hTERT, SV40 LT, and Acrogranin. Introduction of hTERT and SV40 LT was sufficient for immortalizing OSE cells but not enough for tumor formation in nude mice. In contrast, transfection and overexpression of Acrogranin in immortalized OSE cells showed augmented clonogenicity in soft agar and obvious tumorigenicity in nude mice. This is the first study showing evidence that a specific growth factor plays a direct role in malignant transformation in ovarian cancer development.
Oncology Reports 03/2007; 17(2):329-33. · 1.84 Impact Factor
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Junzo Hamanishi,
Masaki Mandai,
Masashi Iwasaki,
Taku Okazaki,
Yoshimasa Tanaka, Ken Yamaguchi,
Toshihiro Higuchi,
Haruhiko Yagi,
Kenji Takakura,
Nagahiro Minato,
Tasuku Honjo,
Shingo Fujii
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ABSTRACT: The ligands for programmed cell death 1 (PD-1), an immunoinhibitory receptor belonging to CD28/cytotoxic T lymphocyte antigen 4 family, are PD-1 ligand 1 and 2 (PD-Ls). Recent reports suggest that the aberrant expression of PD-Ls on tumor cells impairs antitumor immunity, resulting in the immune evasion of the tumor cells. Although an inverse correlation between the expression level of PD-Ls and patients' prognosis has been reported for several malignant tumors, the follow-up period was limited because of the lack of the antibody (Ab) applicable to paraffin-embedded specimens. Here we generated a new Ab against PD-1 ligand 1 (PD-L1) and analyzed the expression level of PD-Ls in human ovarian cancer using paraffin-embedded specimens. Patients with higher expression of PD-L1 had a significantly poorer prognosis than patients with lower expression. Although patients with higher expression of PD-1 ligand 2 also had a poorer prognosis, the difference was not statistically significant. A significant inverse correlation was observed between PD-L1 expression and the intraepithelial CD8(+) T lymphocyte count, suggesting that PD-L1 on tumor cells directly suppresses antitumor CD8(+) T cells. Multivariate analysis showed the expression of PD-L1 on tumor cells and intraepithelial CD8(+) T lymphocyte count are independent prognostic factors. The PD-1/PD-L pathway can be a good target for restoring antitumor immunity in ovarian cancer.
Proceedings of the National Academy of Sciences 03/2007; 104(9):3360-5. · 9.68 Impact Factor
