Daizo Oka

Sesquiterpene lactone parthenolide suppresses tumor growth in a xenograft model of renal cell carcinoma by inhibiting the activation of NF-κB

The transcription factor nuclear factor‐κB (NF‐κB) has been shown to be constitutively activated in various human malignancies, including leukemia, lymphoma and a number of solid tumors. NF‐κB regulates the transcriptional of genes important for tumor invasion, metastasis and chemoresistance. The sesquiterpene lactone parthenolide, an inhibition of NF‐κB, has been used conventionally to treat migraines and inflammation. In this study, renal cancer cell lines OUR‐10 and ACHN were used for in vitro experiments to evaluate growth‐inhibitory effects of parthenolide. An OUR‐10 xenograft model in nude mice was also used to investigate the in vivo growth‐inhibitory effects of parthenolide. Apoptosis in response to treatment of OUR‐10 cells with parthenolide was confirmed. Localization of NF‐κB in response to parthenolide treatment was examined of by immunofluorostaining of OUR‐10 cells with antibody against NF‐κB p65 and by Western blot analysis of OUR‐10 cell and tumor nuclear and cytosol fraction. Parthenolide effectively inhibited proliferation of cultured OUR‐10 cells and triggered apoptosis in vitro. Subcutaneous injection or oral administration of parthenolide showed significant tumor growth inhibition in the xenograft model via decreased production of interleukin‐8 (IL‐8) or vascular endothelial growth factor (VEGF). Immunohistochemistry and Western blot analysis showed decreased nuclear localization of NF‐κB and phosphorylated NF‐κB protein and subsequently expression of MMP‐9, Bcl‐xL and Cox‐2 in response to parthenolide treatment. These results indicate that parthenolide is a useful in the treatment of renal cell carcinoma and acts via inhibition of NF‐κB.

Excision Repair Cross-Complementing Group 1 May Predict the Efficacy of Chemoradiation Therapy for Muscle-Invasive Bladder Cancer

Purpose: Chemoradiation therapy (CRT) is now widely recognized as bladder-preserving therapy for muscle-invasive bladder cancer (MIBC). However, some patients who fail CRT may miss the chance to be cured by cystectomy. Therefore, it is important to select patients with MIBC who are expected to have a good response to CRT. Several reports indicate that the excision repair cross-complementing group 1 (ERCC1) gene is associated with resistance to cisplatin and radiation therapy. In this study, we examined the correlation between ERCC1 and CRT in vitro and in vivo in bladder cancer. Experimental Design: Bladder cancer cell lines T24, 5637, Cl8-2 (multidrug-resistant subline of T24), and CDDP10-3 (cisplatin-resistant subline of T24) were used for in vitro assays to measure ERCC1 expression level and growth inhibition with cisplatin or ionizing radiation (IR). We then examined by immunohistochemistry that whether ERCC1 nuclear staining correlates with the efficacy of CRT using cisplatin in 22 patients with MIBC. Results: Cl8-2 cells expressed ERCC1 mRNA 5.96-fold higher than did T24. Cl8-2 and CDDP10-3 were more resistant to cisplatin or IR than was T24. Resistance to IR, but not to cisplatin, was removed by suppressing ERCC1 using siRNA in both Cl8-2 and CDDP10-3 cells. In immunohistochemistry with ERCC1, 6 of 8 positive cases did not have complete response to CRT, whereas 12 of 14 negative cases had complete response. Sensitivity and specificity were 75% and 85.7%, respectively (P = 0.008). Conclusion: Although further study is needed, ERCC1 expression level may predict the efficacy of CRT for MIBC. Clin Cancer Res; 17(8); 2561–9. ©2010 AACR.

SERPINE2 is a possible candidate promotor for lymph node metastasis in testicular cancer

Testicular germ cell tumors (TGCTs) commonly metastasize to the lymph node or lung. However, it remains unclear which genes are associated with TGCT metastasis. The aim of this study was to identify gene(s) that promoted human TGCT metastasis. We intraperitoneally administered conditioned medium (CM) from JKT-1, a cell-line from a human testicular seminoma, or JKT-HM, a JKT-1 cell sub-line with high metastatic potential, into mice with JKT-1 xenografts. Administration of CM from JKT-HM significantly promoted lymph node metastasis. A cDNA microarray analysis showed that JKT-HM cells highly expressed the Serpine peptidase inhibitor, clade E, member 2 (SERPINE2), which encodes a secreted protein. Administration of CM from SERPINE2-silenced JKT-HM cells inhibited lymph node metastasis in the xenograft model, compared with administration of CM from JKT-HM cells. There was no significant difference in xenograft volume. Moreover, administration of CM from SERPINE2-over-expressing JKT-1 was likely to promote lymph node metastasis in the xenograft model. There was no difference in the in vitro proliferation or migration of JKT-1 cells cultured with CM from JKT-HM cells, compared to that with CM from JKT-1. There was no promotion of proliferation or lymphangiogenesis in the xenografts, as measured by Ki-67 and LYVE-1 immunohistochemistry, respectively. Although we could not clarify how SERPINE2 promoted lymph node metastasis, it may be a promoter in the development of lymph node metastasis in the human seminoma cells in a mouse xenograft model.

Brain metastases from testicular germ cell tumors: A retrospective analysis

Objectives:  To review our series of testicular germ cell tumors with brain metastases and to establish an optimal treatment strategy for them.

The Growth-Inhibitory Effects of Dexamethasone on Renal Cell Carcinoma In Vivo and In Vitro

Background: Recently, several kinase inhibitors have been reported to exert stronger growth inhibitory effects on metastatic renal cell carcinomas (RCCs) than cytokines such as interferons (IFNs) and interleukin-2 (IL-2). On the contrary, the adverse effects of these drugs are also severe. The aim of this study is to analyze the growth-inhibitory effects of DEXamethasone (DEX) on RCC in vivo and in vitro. Methods: The MTT assay was performed using three RCC cell lines, OUR-10, Caki-1, and NC65. OUR-10 cells were subcutaneously transplanted to the dorsal area of nude mice. The nuclear translocation of glucocorticoid receptor (GR) and NF-κ B was examined using appropriate antibodies. Concentrations of interleukin-6 (IL-6), IL-8, and vascular endothelial cell growth factor (VEGF) in the conditioned media and cytosol were measured by enzyme-linked immunosorbent assay (ELISA). Results: All three RCC cell lines responded to DEX treatment. The growth of OUR-10 xenografts was significantly inhibited by administration of DEX. GR was translocated into the nucleus on DEX treatment. Intracellular IL-6, as well as IL-6 in the conditioned medium, decreased in OUR-10 cells following treatment with increasing amounts of DEX. Concentrations of IL-8 and VEGF in the conditioned medium of OUR-10 and NC65 cells also decreased following DEX treatment, with the inhibition of nuclear translocation of NF-κ B. Conclusion: DEX treatment is a candidate for advanced RCC therapy by inhibiting the activation of NF-κ B and its downstream products such as IL-6, IL-8 and VEGF.

Paclitaxel, ifosfamide, and nedaplatin (TIN) salvage chemotherapy for patients with advanced germ cell tumors.

Background:  The paclitaxel, ifosfamide, and cisplatin regimen has been used to treat metastatic testicular cancer with successful results. We investigated the usefulness of a paclitaxel, ifosfamide, and nedaplatin (TIN) regimen as salvage therapy for patients with advanced testicular germ cell tumors (GCTs).

Decreased immunostaining for macrophage scavenger receptor is associated with poor prognosis of prostate cancer

The aim of this study is to evaluate the expression of the macrophage scavenger receptor (MSR) in prostate needle biopsy specimens as a possible prognostic factor for prostate cancer. As MSR reportedly has a role in recognizing foreign pathogenic substances, MSR‐positive inflammatory cells are often detected in solid tumours, and there is a correlation between the relative risk of prostate cancer and polymorphism of the MSR gene.

1998 IMPACT OF HYPONATREMIA ON SURVIVAL OF PATIENTS WITH METASTATIC RENAL CELL CARCINOMA TREATED WITH MOLECULAR TARGETED THERAPY

Atsunari Kawashima*, Hitoshi Takayama, Suita, Japan; Yasuyuki Arai, Osaka, Japan; Mikio Nin, Sakai, Japan; Go Tanigawa, Yutaka Yasunaga, Osaka, Japan; Masatoshi Mukai, Toyonaka, Japan; Hironori Nomura, Daizo Oka, Toshiaki Yoshioka, Osaka, Japan; Satoko Fukuda, Ikeda, Japan; Kenji Nishimura, Nishinomiya, Japan; Nobukazu Murosaki, Itami, Japan; Minoru Koga, Minoh, Japan; Yasuyuki Kojima, Suita, Japan; Miyaji Kyakuno, Osaka, Japan; Takahiro Yoshida, Koji Hatano, Mototaka Sato, Motohide Uemura, Yasutomo Nakai, Suita, Japan; Kazuo Nishimura, Osaka, Japan; Akira Tsujimura, Norio Nonomura, Suita, Japan

Abstract 2475: Excision repair cross complementing group 1 (ERCC1) predicts the efficacy of chemoradiotherapy for invasive bladder cancer

Purpose: Chemoradiation therapy (CRT) is now widely recognized as bladder-preserving therapy for muscle-invasive bladder cancer (MIBC). However, some patients who fail CRT may miss the chance to be cured by cystectomy. Therefore, it is important to select patients with MIBC who are expected to have a good response to CRT. Several reports indicate that the excision repair cross-complementing group 1 (ERCC1) gene is associated with resistance to cisplatin and radiation therapy. In this study, we examined the correlation between ERCC1 and CRT in vitro and in vivo in bladder cancer. Experimental Design: Bladder cancer cell lines T24, 5637, Cl8-2 (multi-drug-resistant subline of T24), and CDDP10-3 (cisplatin-resistant subline of T24) were used for in vitro assays to measure ERCC1 expression level and growth inhibition with cisplatin or irradiation (IR). To clarify the association between ERCC1 and cisplatin resistance in bladder cancer cells, we knocked down ERCC1 in Cl8-2 and CDDDP10-3 with siRNA (C18-2ΔERCC1 and CDDP10-3ΔERCC1). To further prove the cause of the radiation sensitivity of ERCC1 knockdown cells, we measured the phosphorylated histone variant H2A.X, a marker of DNA damage. In the clinical study, we then examined by immunohistochemistry whether ERCC1 nuclear staining correlates with the efficacy of CRT using cisplatin in 22 patients with MIBC. Results: Cl8-2 cells expressed ERCC1 mRNA 5.96-fold higher than did T24. Cl8-2 and CDDP10-3 were more resistant to cisplatin or IR than was T24. Our ERCC1 knockdown experiments showed that there was statistical difference in the resistance to IR exposure not cisplatin compared with C18-2CTL and CDDP10-3CTL. C18-2ΔERCC1 and CDDP10-3ΔERCC1 recovered more slowly in terms of the number of phospho-H2A.X foci than did C18-2cont and CDDP10-3cont, suggesting continued accumulation or persistence of DSBs. In immunohistochemistry with ERCC1, six of eight positive cases did not have complete response to CRT, whereas 12 of 14 negative cases had complete response. Sensitivity and specificity were 75% and 85.7%, respectively (p = 0.008). Conclusion: Our results suggest that in some bladder cancer cells, ERCC1 expression correlates with IR resistance but not with cisplatin resistance. Moreover, the lack of ERCC1 expression correlated well with the efficacy of CRT, and especially with that of IR, in our clinical study. Although further study is needed, ERCC1 expression level may predict the efficacy of CRT for MIBC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2475. doi:10.1158/1538-7445.AM2011-2475

Abstract 3293: Possible role of SERPINE2 in lymph node metastasis of testicular cancer

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Objective Testicular germ cell tumors (TGCTs) commonly metastasize to the lymph node or lung. However, it remains unclear which genes are associated with TGCT metastasis. The aim of this study was to identify gene(s) that promote TGCT metastasis. Methods JKT-1 cell-line derived from human testicular seminoma and JKT-HM cell-line, which is the sub-line of JKT-1 with highly metastatic potential, were used. We intraperitoneally administered PBS or conditioned medium (CM) from JKT-1 or JKT-HM into mice with JKT-1 xenografts for 16 days. Metastasis to lymph nodes were examined 5 weeks after JKT-1 inoculation. We performed MTS assay and transwell assay to clarify if CM from JKT-1 and JKT-HM cells promote a proliferation and a migration of JKT-1 in vitro. Then we analyzed RNA expression pattern of JKT-1 and JKT-HM cells using cDNA microarray. The gene of interest was silenced with siRNA or overexpressed using gene transfer technique and confirmed with mouse xenograft model administrated CM itnraperitoneally as we mentioned above. Results In the xenograft model, metastasis of lymph node metastases were identified in 2/5 (40%), 5/9 (56%) and 12/13 (92%) of mice treated with PBS, CM from JKT-1 and CM from JKT-HM, respectively. There was a significant difference in the frequency of lymph node metastasis between mice treated with CM from JKT-1 cells and those treated with CM from JKT-HM cells (p = 0.043). There was no difference in the in vitro proliferation or migration of JKT-1 cells cultured with CM from JKT-HM cells, compared to that with CM from JKT-1. A cDNA microarray analysis showed that JKT-HM cells highly expressed the Serpine peptidase inhibitor, clade E, member 2 (SERPINE2), which encodes a secreted protein. In the xenograft model, lymph nodes metastases were identified in 13/14 (93%), 14/15 (93%), and 8/14 (57%) of mice treated with CM from JKT-HM cells, CM from JKT-HM cells transfected with control siRNA, and CM from SERPINE2-silenced JKT-HM cells, respectively. There was a significant difference in lymph node metastasis between mice treated with CM from JKT-HM cells transfected with control siRNA and those treated with CM from SERPINE2-silenced JKT-HM cells (p = 0.023). Moreover, there was a significant difference in lymph node metastasis between mice treated with CM from JKT-1 cells(3/12, 23%) and those treated with CM from SERPINE2-over-expressing JKT-1 cells (10/12, 83%) (p = 0.0026). Lymph node metastasis occurred more frequently in CM from SERPINE2-over-expressing JKT-1 cells (83%) than in that from JKT-1 cells transfected control vector (8/12, 67%). However, the difference was not statistically significant (p = 0.3458). Conclusion We identified SERPINE2 as a possible promoter of TGCT metastasis. Although further study would be expected, SERPINE2, which is a secreted protein, may be a new target for the therapy against human TGCT. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3293.