Mitsunobu Sato, Yoshio Hayashi and Hideo Yoshida : Immunopharmacological aspects of OK-432 in humans, --- Effect of immuno-therapy with streptococcal preparation,OK-432, on the peripheral killer lymphocyte population in patients with head and neck cancer ---, 1986.
5.
Mitsunobu Sato, Yoshio Hayashi and Hideo Yoshida : Recent Advances in Chemotherapy, --- Effect of postoperative immunotherapy with streptpcoccal preparation OK-432 on different immunologic parameters in patients with head and neck cancer ---, 1985.
学術論文(審査論文):
1.
Daisuke Uchida, Tomitaro Onoue, Yoshifumi Tomizuka, Nasima Mila Begum, Yoshihiro Miwa, Hideo Yoshida and Mitsunobu Sato : Involvement of an autocrine stromal cell derived factor-1/CXCR4 system on the distant metastasis of human oral squamous cell carcinoma., Molecular Cancer Research : MCR, Vol.5, No.7, 685-694, 2007.
(要約)
We have previously shown that a stromal cell-derived factor-1 (SDF-1; CXCL12)/CXCR4 system is involved in the establishment of lymph node metastasis, but not in that of distant metastasis, in oral squamous cell carcinoma (SCC). In this study, we investigated the role of the autocrine SDF-1/CXCR4 system, with a focus on distant metastasis in oral SCC cells. The immunohistochemical staining of SDF-1 and CXCR4 using primary oral SCCs and metastatic lymph nodes showed a significantly higher number of SDF-1-positive cases among the metastatic lymph nodes than among the primary oral SCCs, which was associated with a poor survival rate among those of the former group. The forced expression of SDF-1 in B88 cells, which exhibit functional CXCR4 and lymph node metastatic potential (i.e., the autocrine SDF-1/CXCR4 system), conferred enhanced cell motility and anchorage-independent growth potential onto the cells. Orthotopic inoculation of the transfectant into nude mice was associated with an increase in the number of metastatic lymph nodes and more aggressive metastatic foci in the lymph nodes. Furthermore, the SDF-1 transfectant (i.e., the autocrine SDF-1/CXCR4 system) exhibited dramatic metastasis to the lung after i.v. inoculation, whereas the mock transfectant (i.e., the paracrine SDF-1/CXCR4 system) did not. Under the present conditions, AMD3100, a CXCR4 antagonist, significantly inhibited the lung metastasis of the SDF-1 transfectant, ameliorated body weight loss, and improved the survival rate of tumor-bearing nude mice. These results suggested that, in cases of oral SCC, the paracrine SDF-1/CXCR4 system potentiates lymph node metastasis, but distant metastasis might require the autocrine SDF-1/CXCR4 system.
Masayuki Azuma, Yuki Ashida, Tetsuya Tamatani, Katsumi Motegi, Natsumi Takamaru, Naozumi Ishimaru, Yoshio Hayashi and Mitsunobu Sato : Cepharanthin,a biscoclaurine alkaloid,prevents destruction of acinar tissues in murine Sjogren's syndrome., The Journal of Rheumatology, Vol.33, No.5, 912-920, 2006.
(要約)
Our previous study suggested that suppression by cepharanthin of tumor necrosis factor-a (TNF-a)-induced matrix metalloproteinase-9 (MMP-9) could prevent destruction of the acinar structure in the salivary glands of patients with Sjögren's syndrome (SS). In this study, we observed that in vivo administration of cepharanthin prevented severe damage to acinar tissues in the murine model of human SS. Cepharanthin was intraperitoneally administered to thymectomized female NFS/sld mice. Inflammatory lesions in the salivary and lacrimal glands were then examined histologically. Expression of phosphorylated IkB-a, MMP-9, and type IV collagen was analyzed immunohistochemically. The apoptotic cell death of acinar cells was determined. Although extensive mononuclear cell infiltration and destruction of acinar tissue in salivary and lacrimal glands were observed in control mice, significant improvement of these lesions was evident in mice treated with cepharanthin. Immunohistochemical analysis revealed that p65, phosphorylated IkB-a, and MMP-9 were more strongly stained in the acinar cells of control mice than in cepharanthin-treated mice. Although no staining for type IV collagen was observed in the acinar tissues of control mice, continuity of staining for type IV collagen was observed in acinar tissues of cepharanthin-treated mice. Destruction of acinar tissues was attributed to the induction of apoptosis, suggesting that cepharanthin inhibits apoptosis by suppressing phosphorylation of IkB-a, followed by prevention of MMP-9 activation. Our findings suggest that cepharanthin may be a promising agent for use in preventing destruction of acinar tissues in murine SS.
Naonobu Tanaka, Mamoru Okasaka, Youko Ishimaru, Yoshihisa Takaishi, Mitsunobu Sato, Masato Okamoto, Tetsuya Oshikawa, Sharif Uddin Ahmed, L. Mark Consentino and Kuo-Hsiung Lee : Biyouyanagin A, an Anti-HIV Agent from Hypericum chinense L. var. salicifolium, Organic Letters, Vol.7, No.14, 2997-2999, 2005.
(要約)
[structure: see text] A structurally unique hydrophobic compound, biyouyanagin A, was isolated from the MeOH extract of the leaves of Hypericum chinense L. var. salicifolium. The structure of biyouyanagin A was elucidated on the basis of spectroscopic evidence. Biyouyanagin A showed a significant activity against HIV and inhibited cytokine production.
Ai Suga, Yoshihisa Takaishi, Hiroyuki Nakagawa, Tadashi Iwasa, Mitsunobu Sato and Masato Okamoto : Chemical Constituents from Fruits and Seeds of Myrica rubra (Myricaceae), Natural Medicines, Vol.59, No.2, 70-75, 2005.
(要約)
Two dammarane-type triterpenes and one steroid were isolated from the fruits of Myrica rubra and one furofran lignan was isolated from the seeds of M.rubra. Their structures were determined on the basis of spectral evidence. In addition, 16 known compounds were assayed for LPS-induced cytokine production.
Takaaki Sagawa, Yoshihisa Takaishi, Yoshinori Fujimoto, Carmenza Duque, Coralia Osorio, Freddy Ramos, Garzon Cristina, Mitsunobu Sato, Masato Okamoto, Tetsuya Oshikawa and Sharif Uddin Ahmed : Cyclobutane Dimers from the Colombian Medicinal Plant Achyrocline bogotensis, Journal of Natural Products, Vol.68, No.4, 502-505, 2005.
(要約)
Five cyclobutane dimers, achyrodimers A-E (1-5), along with 11 known compounds were isolated from the methanol extract of the Colombian medicinal plant Achyrocline bogotensis. Their structures were elucidated by spectroscopy. Several of these compounds were screened for cytokine-inducing activity in human peripheral blood mononuclear cells.
Katsumi Motegi, Masayuki Azuma, Tetsuya Tamatani, Yuki Ashida and Mitsunobu Sato : Expression of aquaporin-5 in and fluid secretion from immortalized human salivary gland ductal cells by treatment with 5-aza-2'-deoxycytidine: a possibility for improvement of xerostomia in patients with Sjögren's syndrome., Laboratory Investigation; a Journal of Technical Methods and Pathology, Vol.85, No.3, 342-353, 2005.
(要約)
The aim of the present study was to investigate the possibility that ductal cells, which preferentially survive and/or proliferate in Sjögren's syndrome (SS) salivary glands of patients with SS, could acquire the functional expression of membrane water channel aquaporin-5 (AQP5). Thus, in this study, we demonstrate that an immortalized normal human salivary gland ductal cell (NS-SV-DC) line, lacking the expression of AQP5, acquires AQP5 gene expression in response to treatment with 5-aza-2'-deoxycytidine (5-Aza-CdR), a DNA demethylating agent. Confocal microscopic analysis revealed the localization of AQP5 expression mainly at the apical and lateral sides of the plasma membrane. The expressed AQP5 protein was functionally active because AQP5 expression resulted in a significant increase in the osmotically directed net fluid rate across monolayers of NS-SV-DC cells. By the analysis of bisulfite sequencing of CpG islands in the AQP5 promoter, hypermethylation within the consensus Sp1-binding sites was commonly observed in parental cell clones, whereas demethylation at the CGs, one in the second consensus Sp1 element and the other outside of the third consensus Sp1 element in the AQP5 promoter, was detected in NS-SV-DC cells after treatment with 5-Aza-CdR. By analyzing the luciferase activity of transfected AQP5 promoter vectors, it became evident that demethylation at the CGs cooperatively functions between these two sites to induce AQP5 expression. Our data, therefore, suggest that treatment of ductal cells with 5-Aza-CdR could result in the expression of the AQP5 gene, thereby leading to increased fluid secretion from ductal cells in SS salivary glands.
Hiroyuki Nakagawa, Yoshihisa Takaishi, Yoshinori Fujimoto, Carmenza Duque, Cristina Garzon, Mitsunobu Sato, Masato Okamoto, Tetsuya Oshikawa and Sharif Uddin Ahmed : Chemical Constituents from the Colombian Medicinal Plant Maytenus laevis, Journal of Natural Products, Vol.67, No.11, 1919-1924, 2004.
(要約)
The methanol extract of the bark of the Colombian medicinal plant Maytenus laevis gave six new compounds and 28 known compounds. The structures of the new and known compounds were elucidated on the basis of spectroscopic evidence. Several of these compounds were screened for cytokine-inducing activity on human PBMCs to investigate antitumor effects, and canophyllol (12) demonstrated the most effective induction of the cytokines.
Daisuke Uchida, Nasima Mila Begum, Ammar Almofti, Hitoshi Kawamata, Hideo Yoshida and Mitsunobu Sato : Frequent Down-regulation of 14-3-3 sigma protein and Hypermethylation of 14-3-3 sigma gene in Salivary Gland Adenoid Cystic Carcinoma, British Journal of Cancer, Vol.91, No.6, 1131-1138, 2004.
(要約)
14-3-3 sigma:, a target gene of the p53 tumour suppressor protein, has been shown to regulate the cell cycle at the G2/M checkpoint. Recent studies have demonstrated that 14-3-3 sigma is downregulated by hypermethylation of the CpG island in several types of cancer. In this study, we investigated the expression and methylation status of 14-3-3 sigma in human salivary gland adenoid cystic carcinoma (ACC) and mucoepidermoid carcinoma (MEC). Immunohistochemical analysis revealed that the positive expression rate of 14-3-3 sigma in ACC (one out of 14) was markedly lower than that in MEC (ten out of 10). Since most of the ACCs carried the wild-type p53 protein, downregulation of 14-3-3 sigma in ACC may not be due to the dysfunction of p53 pathway. Microdissection-methylation-specific PCR revealed that frequent hypermethylation of the 14-3-3 sigma gene was observed in ACC when compared to that in MEC. In cultured-ACC cells, we confirmed the downregulation of 14-3-3 sigma via hemimethylation of the gene by sequencing analysis after sodium bisulphite treatment. Furthermore, re-expression of 14-3-3 sigma in the ACC cells was induced by the treatment with DNA demethylating agent, 5-aza-2'-deoxycytidine. Irradiation apparently induced the enhanced expression of 14-3-3 sigma and G2/M arrest in normal salivary gland cells; however, in the ACC cells, neither induction of 14-3-3 sigma nor G2/M arrest was induced by irradiation. These results suggest that downregulation of 14-3-3 sigma might play critical roles in the neoplastic development and radiosensitivity of ACC.
Masato Okamoto, Sachiko Furuichi, Yasuhiko Nishioka, Tetsuya Oshikawa, Tomoyuki Tano, Sharif Uddin Ahmed, Kiyoshi Takeda, Shizuo Akira, Yoshiki Ryoma, Yoichiro Moriya, Motoo Saito, Saburo Sone and Mitsunobu Sato : Expression of Toll-Like Receptor 4 on Dendritic Cells Is Significant for Anticancer Effect of Dendritic Cell-Based Immunotherapy in Combination with an Active Component of OK-432, a Streptococcal Preparation, Cancer Research, Vol.64, No.15, 5461-5470, 2004.
(要約)
A lipoteichoic acid-related molecule OK-PSA is an active component of OK-432, a Streptococcus-derived anticancer immunotherapeutic agent. In the present study, we first examined the effect of OK-PSA on the maturation of dendritic cells (DCs) in vitro by using the DCs derived from 5 healthy donors and 10 patients with head and neck cancer with or without expression of toll-like receptor 4 (TLR4) or MD-2 mRNA. OK-PSA treatment effectively increased the surface expression of MHC class II, CD80, CD83, and CD86. OK-PSA-stimulated DCs secreted the cytokines that can induce helper T-cell 1 (Th1)-type T-cell response, and stimulated allogeneic T cells to produce IFN-gamma and to elicit an allogeneic antigen-specific cytotoxicity. These activities almost depended on expression of TLR4 and MD-2 genes. We next investigated the in vivo anticancer effect of intratumoral administration of syngeneic DCs followed by OK-PSA against established tumors in mice. C57BL/6 mice, which express wild-type TLR4, and C57BL/6-derived TLR4-knockout (TLR4(-/-)) mice were used. Although OK-PSA accelerated the antitumor effect of intratumoral DC administration in wild-type mice bearing syngeneic tumors, the antitumor effect of OK-PSA as well as of the combination therapy with DCs and OK-PSA was not significant in TLR4(-/-) mice. Interestingly, an administration of wild-type-mouse-derived DCs followed by OK-PSA exhibited a marked antitumor effect even in the TLR4(-/-) mice. These findings suggest that OK-PSA may be a potent adjuvant for local DC therapy, and that DC therapy followed by OK-PSA is able to elicit anticancer activity even in a TLR4-deficient host when TLR4 is expressed only in DCs injected intratumorally.
Pycnodysostosis is an uncommon, autosomal recessive skeletal dysplasia characterized by osteosclerosis, bone fragility, osteomyelitis of the mandible, short stature, and actroosteolysis. Recently, we encountered a patient with pycnodysostosis associated with persistent osteomyelitis of the mandible. We report the treatment and mutation analysis of this patient. A 57-year-old man visited our clinic because of pus discharge from the mandibular skin. He had a history of three tibial fractures and had been given a diagnosis of pycnodysostosis at the department of orthopedic surgery. Resolution, sequesterectomy, and excision of the external fistula were performed for a diagnosis of osteomyelitis of the mandible with an external fistula. Subsequently, recrudescence occurred three times, but the patient responded to treatment with azithromycin (AZM). Mutations in the gene encoding cathepsin K, a cysteine protease localized exclusively in osteoclasts, have been recently demonstrated to be responsible for pycnodysostosis. In this study, we performed rapid mutation analysis by amplifying cathepsin K open reading frame (ORF) by reverse transcriptase-polymerase chain reaction (RT PCR), using highly sensitive DNA polymerase and total RNA derived from hematopoietic cells. We found that mutation of cathepsin K gene products converted leucine from proline at the position of the 9th amino acid (codon 9<SUP>leu-pro</SUP>; Pro; L9P) These results indicated that persistent osteomyelitis of the mandible in this patient resulted in a major symptom characterized by pycnodysostosis. Moreover, our experimental approach is rapid and sensitive and may be useful for the mutation analysis of diverse diseases.
Ammar Almofti, Daisuke Uchida, Nasima Mila Begum, Yoshifumi Tomizuka, Hiroki Iga, Hideo Yoshida and Mitsunobu Sato : The clinicopathological significance of the expression of CXCR4 protein in oral squamous cell carcinoma, International Journal of Oncology, Vol.25, No.1, 65-71, 2004.
(要約)
We have demonstrated the possibility that the stromal cell-derived factor-1 (SDF-1; CXCL12)/CXCR4 system might be involved in the establishment of lymph node metastasis in oral squamous cell carcinoma (SCC). In order to further clarify the role of the SDF-1/CXCR4 system in oral SCC, we examined the expression of CXCR4 and SDF-1 in biopsy specimens from 61 patients with oral SCC by means of immunohistochemistry, and investigated several clinicopathological factors, including age, sex, lymph node metastasis, invasion, recurrence and prognosis. The expression of CXCR4 and SDF-1 was observed in 57.3 and 11.4% of our subjects, respectively. Although we were unable to find a statistically significant association between the expression of SDF-1 and any clinicopathological factors, we did find a significant association between the expression of CXCR4 and lymph node metastasis (P=0.0417). Moreover, the mode of invasion (P=0.0002) and recurrence of the tumors (P=0.0185) were strongly associated with CXCR4 expression, and the CXCR4-positive group showed a significantly poorer prognosis than the CXCR4-negative group (P=0.0401). Recombinant SDF-1alpha stimulated in vitro invasiveness and scattering in CXCR4-expressing oral SCC cells, but the treatment did not affect the expression of matrix metalloproteinases or urokinase-type plasminogen activator. These results indicate that SDF-1/CXCR4 signaling in oral SCC cells might be involved in the diverse action of oral SCC, including invasion or micrometastasis at the primary site and lymph node metastasis.
Tetsuya Tamatani, Masayuki Azuma, Y Ashida, Katsumi Motegi, R Takashima, Koji Harada, S Kawaguchi and Mitsunobu Sato : Enhanced radiosensitization and chemosensitization in NF-κB-suppressed human oral cancer cells via the inhibition of g-irradiation- and 5-FU-induced production of IL-6 and IL-8, International Journal of Cancer, Vol.108, No.6, 912-921, 2004.
(要約)
We examined the mechanisms underlying the enhancement of radiosensitivity and chemosensitivity to gamma-irradiation (IR) and 5-Fluorouracil (5-FU) in human oral carcinoma cells (B88) in which NF-kappaB activity was constitutively suppressed. Three super-repressor form of IkappaBalpha cDNA-transfected cell (B88mI) clones and 1 empty vector-transfected cell clone (B88neo) have been established. We found that the tumor-forming ability in nude mice of B88mI clones was significantly lower than that of B88 or B88neo. This suppressed ability in tumorigenicity was attributed to the down-regulation of the expression of interleukin (IL)-1alpha, IL-6, IL-8, vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP)-9 in B88mI cell clones as compared to that in B88 or B88neo. IR and 5-FU induced a much greater degree of apoptosis, as evidenced by flow cytometry analysis and annexin V staining, in B88mI cell clones than in B88 or B88neo. When tumor-bearing nude mice were treated with IR or 5-FU, the suppression of tumor growth was significantly augmented in B88mI cell clones as compared to that in B88 or B88neo. ELISA analysis indicated that although a remarkable increase in production of IL-6 and IL-8 was observed in B88 and B88neo after in vitro exposure to IR or treatment with 5-FU, radiotherapy and chemotherapy-induced production of these cytokines was significantly suppressed in B88mI cell clones. These findings suggest that production of angiogenic factors and growth factors in response to radiotherapy and chemotherapy is a principal mechanism of inducible radioresistance and chemoresistance in human oral cancers, and establish the inhibition of NF-kappaB as a rational approach to improve conventional radiotherapy and chemotherapy outcomes.
Daisuke Uchida, Nasima Mila Begum, Yoshifumi Tomizuka, Takashi Bando, Ammar Almofti, Hideo Yoshida and Mitsunobu Sato : Acquisition of lymph node, but not distant metastatic potentials, by the over-expression of CXCR4 in human oral squamous cell carcinoma cells, Laboratory Investigation; a Journal of Technical Methods and Pathology, Vol.84, No.12, 1538-1546, 2004.
Ko-ichi Nakashiro, Mila Nasima Begum, Daisuke Uchida, Hitoshi Kawamata, Satoru Shintani, Mitsunobu Sato and Hiroyuki Hamakawa : Thiazolidinediones inhibit cell growth of human oral squamous cell carcinoma in vitro independent of peroxisome proliferator-activated receptor gamma, Oral Oncology, Vol.39, No.8, 855-861, 2003.
(要約)
Peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear receptor superfamily of ligand-activated transcription factors. Recently, we have demonstrated that PPARgamma is expressed in human salivary gland tumors and its ligands inhibit the growth of cultured salivary gland cancer cells. However, expression and function of PPARgamma in normal and neoplastic human oral squamous epithelium remains unclear. In the present study, we examined PPARgamma expression in human oral squamous cell carcinoma (OSCC) and tested its ligands for any antitumor effect. PPARgamma mRNA was detected by RT-PCR in some OSCC tissues and cultured cells, but the PPARgamma protein showed neither expression nor ligand-induced transcriptional activity. Despite loss of PPARgamma function, synthetic PPARgamma ligands caused significant dose-dependent inhibition of cancer cell growth. These results suggest that PPARgamma function is inactivated in OSCC cells and the anti-proliferative effect of its synthetic ligands is independent of PPARgamma.
Daisuke Uchida, Nasima Mila Begum, Ammar Almofti, Koichi Nakashiro, Hitoshi Kawamata, Yoshihisa Tateishi, Hiroyuki Hamakawa, Hideo Yoshida and Mitsunobu Sato : Possible role of stromal-cell-derived factor-1/CXCR4 signaling on lymph node metastasis of oral squamous cell carcinoma, Experimental Cell Research, Vol.290, No.2, 289-302, 2003.
(要約)
We examined the role of chemokine signaling on the lymph node metastasis of oral squamous cell carcinoma (SCC) using lymph node metastatic (HNt and B88) and nonmetastatic oral SCC cells. Of 13 kinds of chemokine receptors examined, only CXCR4 expression was up-regulated in HNt and B88 cells. CXCR4 ligand, stromal-cell-derived factor-1alpha (SDF-1alpha; CXCL12), induced characteristic calcium fluxes and chemotaxis only in CXCR4-expressing cells. CXCR4 expression in metastatic cancer tissue was significantly higher than that in nonmetastatic cancer tissue or normal gingiva. Although SDF-1alpha was undetectable in either oral SCC or normal epithelial cells, submandibular lymph nodes expressed the SDF-1alpha protein, mainly in the stromal cells, but occasionally in metastatic cancer cells. The conditioned medium from lymphatic stromal cells promoted the chemotaxis of B88 cells, which was blocked by the CXCR4 neutralization. SDF-1alpha rapidly activated extracellular signal-regulated kinase (ERK)1/2 and Akt/protein kinase B (PKB), and their synthetic inhibitors attenuated the chemotaxis by SDF-1alpha. SDF-1alpha also activated Src family kinases (SFKs), and its inhibitor PP1 diminished the SDF-1alpha-induced chemotaxis and activation of both ERK1/2 and Akt/PKB. These results indicate that SDF-1/CXCR4 signaling may be involved in the establishment of lymph node metastasis in oral SCC via activation of both ERK1/2 and Akt/PKB induced by SFKs.
Mitsunobu Sato, Masayuki Azuma, Tetsuya Tamatani, Yuki Ashida, Rina Takashima and Koji Harada : Cisplatin induces apoptosis in oral squamous carcinoma cells by the mitochondria-mediated but not the NF-kB-suppressed pathway, Oral Oncology, Vol.39, No.3, 282-289, 2003.
(要約)
Cisplatin (CDDP) is a potent DNA-damaging anticancer agent, and its cytotoxic action is exerted by the induction of apoptosis. However, activation of the transcription factor NF-kappaB results in protection against apoptosis. We examined the molecular mechanisms involved in the induction of apoptosis by CDDP as regards both suppression of NF-kappaB and activation of caspases. Human oral squamous carcinoma cells (B88) were employed in this study. We found that CDDP treatment affected neither NF-kappaB activity nor the expression levels of antiapoptotic proteins, including TRAF-1, TRAF-2, and cFLIP, in B88 cells. However, two apoptosome molecules, cytochrome c and Apaf-1, were significantly augmented in the cytoplasm by CDDP treatment. Further, the activation of caspase-9 and caspase-3, downstream molecules leading to mitochondria-mediated apoptosis, were detected after treatment with CDDP. Finally, apoptosis was also clearly observed, as evidenced by cleavage of PARP through the activation of caspase-3. These findings suggest that CDDP exerts its apoptotic action by the mitochondria-mediated activation of caspases but not by the activation of caspases due to the inhibition of NF-kappaB activity that follows the suppression of antiapoptotic proteins.
Tetsuya Oshikawa, Masato Okamoto, Go Ohe, Sachiko Furuichi, Hidetomo Nishikawa, Sharif Ahmed Uddin, Hideo Yoshida, Yoichiro Moriya, Shuzo Matsubara, Yoshiki Ryoma, Motoo Saito and Mitsunobu Sato : Anti-tumor immune response induced by the fractions derived from OK-432, a streptococcal preparation, by using a monoclonal antibody TS-2 that neutralizes the interferon-gamma-inducing activity of OK-432: comparison between the TS-2-binding and TS-2-unbinding fraction., International Immunopharmacology, Vol.3, No.5, 643-655, 2003.
(要約)
We have previously isolated a lipoteichoic acid (LTA)-related molecule (OK-PSA) from OK-432, a streptococcal agent, by affinity chromatography on a CNBr-activated Sepharose 4B bound TS-2 monoclonal antibody (mAb) that neutralizes the interferon (IFN)-gamma-inducing activity of OK-432. In the current study, we compared the cytokine-inducing and anti-tumor activities of OK-PSA, a TS-2-binding fraction, with those of OK-PTF, a TS-2-unbinding fraction, in order to determine the efficacy of OK-PSA for clinical use in affinity chromatography using TS-2. In the in vitro experiments using human peripheral blood mononuclear cells (PBMCs), OK-PSA markedly induced Th1-type cytokines, while interleukin (IL)-6 and IL-10, Th2-type cytokines, were induced by OK-PTF. Th1-cytokine induction by OK-PTF was not dose-dependent and was suppressed when PBMCs were treated with a high concentration of OK-PTF. In a mouse model, Th1 cytokines were also induced by OK-PSA and Th2 cytokines were induced by OK-PTF. Th2 cytokine-inducing activity of OK-PTF was accelerated in tumor-bearing mice relative to that in healthy mice. Although the anti-tumor effect of OK-PTF was statistically significant, it was much weaker than that of OK-PSA. A significant difference between the anti-tumor effect of OK-PSA and that of OK-PTF was observed (P<0.05). Finally, OK-PSA elicited its cytokine-inducing effect via Toll-like receptor (TLR) 4, whereas OK-PTF-induced signaling was mediated by both TLR2 and TLR4. These findings strongly suggested that the affinity chromatography using TS-2 is a useful strategy to separate the effective component for cancer therapy (OK-PSA) from other components.
Mitsunobu Sato, Masato Okamoto, Tetsuya Oshikawa, Tomoyuki Tano, Go Ohe, Sachiko Furuichi and Hidetomo Nishikawa : Involvement of Toll-like receptor 4 signaling in interferon-γ production and anti-tumor effect by a streptococcal agent OK-432., Journal of the National Cancer Institute, Vol.95, No.4, 316-326, 2003.
(要約)
The streptococcal agent OK-432 has been used for immunotherapy of head and neck cancer, among other malignancies, but its mechanism of action is unknown. Because the Toll-like receptor 4 (TLR4)/MD-2 complex is important in enabling the mammalian immune system to recognize bacterial components, we investigated whether expression of the TLR4 and MD-2 genes is associated with OK-432-induced anticancer immunity. Peripheral blood mononuclear cells (PBMCs) from 28 patients with head and neck cancer were analyzed for TLR4 and MD-2 mRNA expression by reverse transcription-polymerase chain reaction (RT-PCR) analysis. PBMCs were treated in vitro with OK-432 or with OK-PSA (a lipoteichoic-acid-related molecule that is an active component of OK-432), and interferon-gamma (IFN-gamma) mRNA expression, an immune response measure, was analyzed by RT-PCR. Patient sera collected 24 hours after OK-432 administration were examined for IFN-gamma protein using an enzyme-linked immunosorbent assay. Lewis lung carcinoma-bearing wild-type C57BL/6 and TLR4-deficient mice (four mice per group) received intraperitoneal injections of OK-432, and tumor volumes and sera IFN-gamma levels were measured over time. All statistical tests were two-sided. Twenty patients expressed both TLR4 and MD-2. Expression of TLR4 and MD-2 genes was associated with the in vivo IFN-gamma induction in 19 patients administered OK-432 (Fisher's exact test P<.001). Although both OK-432 and OK-PSA induced IFN-gamma expression from PBMCs in vitro, expression of TLR4 and MD-2 was associated only with IFN-gamma expression induced by OK-PSA (P<.001). In vivo intraperitoneal administration of OK-432 resulted in an increase of IFN-gamma in sera from wild-type mice but not in sera from TLR4-deficient mice. Tumors in wild-type mice treated with OK-432 were statistically significantly smaller than those in mice treated with saline (P =.007). By contrast, in TLR4-deficient mice, there was no difference in tumor volume between the two treatment groups. TLR4 and MD-2 may mediate OK-432-induced anticancer immunity.
Mitsunobu Sato, Masato Okamoto, Go Ohe, Sachiko Furuichi, Hidetomo Nishikawa, Tetsuya Oshikawa, Tomoyuki Tano, SU Ahmed and Hideo Yoshida : Enhancement of anti-tumor immunity by lipoteichoic acid-related molecule isolated from OK-432, a streptococcal agent, in athymic nude mice bearing human salivary adenocarcinoma: Role of natural killer cells, Anticancer Research, Vol.22, No.6A, 3229-3240, 2002.
(要約)
OK-PSA, a lipoteichoic acid (LTA)-related molecule isolated from a streptococcal agent OK-432, enhances anti-tumor immunity as a potent inducer of Th1-type cytokines. Recently, we obtained the data suggesting that natural killer (NK) cells may play a significant role for OK-PSA-induced cytokine production in vitro. We conducted the animal experiments using athymic nude mice bearing human salivary adenocarcinoma to examine the role of NK cells in OK-PSA-induced anti-tumor immunity. OK-PSA was peritumorally injected into the mice. Cytokines in the sera were analyzed by ELISA. mRNAs for cytokines were detected by RT-PCR. 51Cr release test was performed to measure killer cell activities. OK-PSA markedly increased the amounts of IFN-gamma, tumor necrosis factor (TNF)-alpha, interleukin (IL)-2, IL-12 and IL-18 that are generally called "Th1-type cytokines" in the sera derived from tumor-bearing nude mice, and also accelerated the killing activities of tumor-infiltrating lymphocytes as well as of draining lymph node cells. Furthermore, OK-PSA administration resulted in significant inhibition of tumor growth, but the effect of OK-PSA was almost completely inhibited by the deletion of NK cells using anti-asialo GM1 antibody. These findings strongly suggested that NK cells are closely involved in OK-PSA-mediated anti-tumor immunity.
Mitsunobu Sato, Koji Harada, Hideo Yoshida and Supriatno : Low p27Kip1 expression is associated with poor prognosis in oral squamous cell carcinoma., Anticancer Research, Vol.22, No.5, 2985-2989, 2002.
(要約)
p27Kip1 is a cyclin-dependent kinase inhibitor which regulates the progression of cells from the G1-into the S-phase in a cell cycle. Loss of p27Kip1 is associated with disease progression and an unfavorable outcome in several malignancies. The purpose of this study was to determine whether p27Kip1 expression can be a useful prognostic factor in oral squamous cell carcinoma (SCC) patients. p27Kip1 expression was investigated by immunohistochemistry in tissue samples from 81 patients with oral SCC. The associations between p27Kip1 expression and clinicopathological characteristics and patient survival were also analyzed. Significant associations were found between p27Kip1 expression and histological grade (p = 0.010), therapeutic effect (p = 0.004) and patient outcome (p = 0.005). The 5-year survival rates of p27Kip1 high- and low-expression tumours were 80.4% and 56.7%, respectively, this difference being significant (p = 0.009) by log-rank test. Multivariate analysis revealed that reduced term survival was related to low levels of p27Kip1 expression (p = 0.008) and advanced stage (stages III and IV) (p = 0.003). These results suggest that reduction of p27Kip1 plays an important role in the progression of oral SCC and is considered to be a useful prognostic factor in oral SCC patients.
MO Hoque, Hitoshi Kawamaha, Koh-ichi Nakashiro, Fumie Omotehara, Satoshi Hino, Daisuke Uchida, Koji Harada, NM Begum, Hideo Yoshida, Mitsunobu Sato and Takahiro Fujimori : Dysfunction of the p53 tumor suppressor pathway in head and neck cancer, International Journal of Oncology, Vol.21, No.1, 119-126, 2002.
(要約)
It is important to examine the abnormality of the entire p53 tumor suppressor pathway in head and neck cancer. We examined the mRNA expressions of p53 regulatory factors, p33ING1 and p14ARF, and a p53-target gene, p21WAF1 in head and neck cancer. Nine of 14 benign pleomorphic adenomas (PAs) and 7 of 8 malignant salivary gland tumors (MSGTs) expressed p33ING1 mRNA. Thirteen of 14 PAs expressed p14ARF mRNA, however, only 1 of 8 MSGTs expressed p14ARF mRNA. Eight of 14 PAs and 7 of 8 MSGTs expressed p21WAF1 mRNA. In salivary gland tumors, there was clear correlation between the expression of p33ING1 and p21WAF1 (p<0.0001, r2=0.53). However, there was no correlation between the expression of p14ARF and p21WAF1 (p=0.6543, r2=0.009). Twenty-six of 28 oral squamous cell carcinomas (SCCs) expressed p33ING1 mRNA. Nineteen of 28 oral SCCs expressed p14ARF mRNA. All of the oral SCCs expressed p21WAF1 mRNA. In oral SCCs, the expressions of both p33ING1 (p=0.009, r2=0.181) and p14ARF (p=0.0009, r2=0.271) correlated with the expression of p21WAF1. Interestingly, 24 of 26 oral SCCs (92%) showed either abnormality of p53 itself or loss of expression of p53 regulatory factors, p33ING or p14ARF. These results suggest that head and neck cancer often involve the dysfunction of p53 tumor suppressor pathway.
Masayuki Azuma, Keiko Aota, Tetsuya Tamatani, Katsumi Motegi, Yuki Ashida, Yoshio Hayashi, Mitsunobu Sato and Mitsunobu Sato : Suppression of TNF-α-induced MMP 9 production in human salivary gland acinar cells by cepharanthine occurs via down-regulation of NFκB, --- A possible therapeutic agent for preventing the destruction of the acinar structure in the salivary glands of Sjögren's syndrome patients ---, Arthritis and Rheumatism, Vol.46, No.6, 1585-1594, 2002.
(要約)
Our previous results suggested that suppression of tumor necrosis factor alpha (TNFalpha)-induced matrix metalloproteinase 9 (MMP-9) could prevent the destruction of acinar tissue in the salivary glands of patients with Sjögren's syndrome (SS). The present study was undertaken to investigate the effect of cepharanthine on the suppression of TNFalpha-induced MMP-9 production in NS-SV-AC, an SV40-immortalized normal human acinar cell clone. After pretreatment with or without cepharanthine, NS-SV-AC cells were treated with TNFalpha alone or with a combination of TNFalpha and cepharanthine. The expression of MMP-9 was then examined at the protein and messenger RNA levels. In addition, the effect of cepharanthine on the morphogenetic behavior of NS-SV-AC cells cultured on type IV collagen-coated dishes in the presence of TNFalpha was examined. Although TNFalpha induced the production of MMP-9 in NS-SV-AC cells, this production was greatly suppressed when cells were pretreated with cepharanthine, followed by treatment with both TNFalpha and cepharanthine. In addition, cepharanthine suppressed the TNFalpha-stimulated NF-kappaB activity by partly preventing the degradation of IkappaBalpha protein in NS-SV-AC cells. When NS-SV-AC cells were seeded on type IV collagen-coated dishes in the presence of both TNFalpha and plasmin, type IV collagen interaction with the cells was lost and the cells entered apoptosis. However, pretreatment with cepharanthine restored the aberrant in vitro morphogenesis of the NS-SV-AC cells. These results may indicate a molecular mechanism by which cepharanthine is able to protect against the destruction of the acinar structure in salivary glands from patients with SS.
Satoshi Hino, Hitoshi Kawamaha, Fumie Omotehara, Daisuke Uchida, Yoshihiro Miwa, NM Begum, Hideo Yoshida, Mitsunobu Sato and Takahiro Fujimori : Cytoplasmic TSC-22 (transforming growth factor-beta-stimulated clone-22) markedly enhances the radiation sensitivity of salivary gland cancer cells, Biochemical and Biophysical Research Communications, Vol.292, No.4, 957-963, 2002.
(要約)
We transfected a salivary gland cancer cell line, TYS, with three different forms of TSC-22 (transforming growth factor-beta-stimulated clone-22) gene: full-length TSC-22 (TSC-22FL) containing nuclear export signal, TSC-box and leucine zipper, truncated TSC-22 (TSC-22LZ) containing only TSC-box and leucine zipper, and truncated TSC-22 with nuclear localization signal (NLS-TSC-22LZ). High expression of TSC-22FL in the cytoplasm markedly enhanced the radiation-sensitivity of TYS cells, while, moderate expression of TSC-22FL marginally affected the radiation-sensitivity. TSC-22LZ, which was expressed in the cytoplasm and the nucleus, enhanced the radiation-sensitivity of TYS cells irrespective to its expression level. NLS-TSC-22LZ, which was expressed only in the nucleus, marginally affected the radiation-sensitivity of the cells even at high expression level. Interestingly, cytoplasmic TSC-22 translocates to nucleus concomitant with radiation-induced apoptosis. These results suggest that cytoplasmic localization of TSC-22 and translocation of TSC-22 from cytoplasm to nucleus is important for regulating the cell death signal after irradiation-induced DNA damage.
Satoshi Hino, Hitoshi Kawamata, Fumie Omotehara, Daisuke Uchida, NM Begum, Hideo Yoshida and Mitsunobu Sato : Leucine zipper structure of TSC-22 (TGF-b stimulated clone-22) markedly inhibits the anchorage-independent growth of salivary gland cancer cells, Oncology Reports, Vol.9, No.2, 371-374, 2002.
(要約)
Several investigators have demonstrated that TGF-beta stimulated clone-22 (TSC-22) regulates cell growth and differentiation, and cell death. TSC-22 is a putative transcriptional regulator containing a leucine zipper-like structure and a nuclear export signal. We previously showed the cytoplasmic localization of TSC-22 and the nuclear translocation of TSC-22 concomitant with induction of apoptosis in salivary gland cancer cells. In the present study, we attempted to identify the active domain of TSC-22 protein that regulated the biological functions of TSC-22. We constructed three mammalian expression vectors, which could produce full length TSC-22 only in cytoplasm, the leucine zipper structure of TSC-22 in both cytoplasm and nucleus, and the leucine zipper structure only in nucleus. Then, we transfected a salivary gland cancer cell line, HSG with these expression vectors, and investigated the growth profile of the transfectants. None of the TSC-22 constructs inhibited the monolayer growth and the anchorage-dependent colony formation of HSG cells. However, the leucine zipper structure of TSC-22 markedly inhibited the anchorage-independent colony formation of HSG cells (p<0.001; one way ANOVA). Full length TSC-22 also suppressed the anchorage-independent colony formation of HSG cells, although the effect of full length TSC-22 was much lower than those of the leucine zipper constructs. These observations suggest that the leucine zipper structure in TSC-22 protein is an active domain that negatively regulates the growth of salivary gland cancer cells.
Mila Nasima Begum, Ko-ichi Nakashiro, Hitoshi Kawamata, Daisuke Uchida, Satoru Shintani, Yoji Ikawa, Mitsunobu Sato and Hiroyuki Hamakawa : Expression of peroxisome proliferator-activated receptor _ and the growth inhibitory effect of its synthetic ligands in human salivary gland cancer cell lines, International Journal of Oncology, Vol.20, No.3, 599-605, 2002.
(要約)
Peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear receptor superfamily of ligand-activated transcription factors. It is expressed in several tissue types, including adipose tissue in which it stimulates adipogenesis. Recent studies have demonstrated that PPARgamma ligands induce cellular differentiation and inhibit cell growth in carcinomas of various organs including the breast, prostate, lung, colon, stomach, bladder, and pancreas. However, whether PPARgamma is expressed in human salivary gland tumors and its function in this tissue is unknown. In the present study, we examined PPARgamma gene expression in human salivary gland cancer cells and tested its ligands for any antitumor effect. PPARgamma mRNA was detected by RT-PCR in both benign and malignant salivary gland tumor tissues. The effect of PPARgamma on cell growth was investigated using four human salivary gland cancer cell lines; HSG, AZA3, HSY and TYS, which were confirmed to express PPARgamma1 mRNA and protein. Retinoid X receptor alpha protein, which forms heterodimers with PPARgamma, was also detected in all the cells tested. Data obtained by luciferase assay indicated that the intrinsic PPARgamma protein was activated by the synthetic ligands, troglitazone and pioglitazone, but not by the natural ligand, 15-deoxy-delta12, 14-prostaglandin J2. The synthetic PPARgamma ligands, particularly troglitazone, caused significant dose-dependent inhibition of cancer cell growth. Furthermore, the overexpression of PPARgamma1 or PPARgamma2 in cancer cells also reduced significantly their growth rate. These results suggest that PPARgamma and its synthetic ligands suppress the growth of human salivary gland cancer cells and it may be a useful molecular target for cancer treatment.
Mitsunobu Sato, Keiko Aota, Masayuki Azuma, Tetsuya Tamatani, Tuyoshi Yamashita and Yuki Ashida : Stable inhibition of NF-kB in salivary gland cells does not enhance sensitivity to TNF-a-induced apoptosis due to upregulation of TRAF-1 expression, Experimental Cell Research, Vol.276, No.1, 111-119, 2002.
(要約)
The transcription factor NF-kappa B inhibits the apoptotic response induced by TNF-alpha. However, in salivary gland cell clones (ACMT-6 and ACMT-7) in which NF-kappa B activation was suppressed by introduction of a super-repressor form of I kappa B-alpha cDNA, TNF-alpha did not cause apoptosis. Thus, to investigate the molecular mechanism involved in the unresponsiveness of these cell clones to TNF-alpha-induced apoptosis, we examined the effect of TNF-alpha on the expression of antiapoptotic proteins, including TNF receptor-associated factor (TRAF)-1, TRAF-2, cellular inhibitor of apoptosis protein (cIAP)-1, and cIAP-2. Here we show that expression of TRAF-1 was commonly detected by treatment with TNF-alpha in ACMT-6, ACMT-7, and an empty vector-transfected cell clone (ACpRc-1) and that downregulation of TRAF-1 protein by either treatment with an antisense oligonucleotide or introduction of an antisense plasmid resulted in the induction of apoptosis in these cell clones. Our results, therefore, suggest that one of the mechanisms by which cells acquire resistance to TNF-alpha-induced apoptosis is a TNF-alpha induction of TRAF-1.
Tetsuya Tamatani, Masayuki Azuma, Aota K, Yamashita T, Takashi Bando and Mitsunobu Sato : Enhanced IkB kinase activity is responsible for the augmented activity of NF-_B in human head and neck carcinoma cells, Cancer Letters, Vol.171, No.2, 165-172, 2001.
(要約)
The nuclear transcription factor kappaB (NF-kappaB) plays an important role in the development and progression of cancers. However, the mechanism by which cancer cells in the head and neck region acquire high NF-kappaB activity has not yet been clarified. In this study, we examined the NF-kappaB binding activity and the expression of the signal-transduction-related proteins of NF-kappaB in head and neck carcinoma cell lines. These cancer cells showed significantly higher NF-kappaB binding activity than normal oral epithelial and salivary gland cells. We also demonstrated the increased phosphorylation and degradation of IkappaB-alpha protein in cancer cells. Thus, enhanced NF-kappaB activity in cancer cells is attributable to the rapid phosphorylation and degradation of IkappaB-alpha protein. To further elucidate the mechanism involved in this phenomenon, we analyzed both the expression levels of upstream kinases (IkappaB kinase- (IKK-) alpha, IKK-beta, IKK-gamma, and NF-kappaB-inducing kinase (NIK)) and the IKK activity in cells. Although there was no significant difference in the expression levels of NIK, IKK-beta, or IKK-gamma in cancer cell lines compared to those in normal cells, increased expression of IKK-alpha protein was observed in cancer cells. In addition, IKK activity was significantly augmented in cancer cells as compared to normal cells. Thus, our results suggest that enhanced NF-kappaB activity in head and neck cancer cells may be due to the augmentation of IKK activity.
(キーワード)
Carcinoma, Squamous Cell / Head and Neck Neoplasms / Humans / I-kappa B Kinase / Isoenzymes / NF-kappa B / NF-kappa B p50 Subunit / Phosphorylation / Protein Binding / Protein-Serine-Threonine Kinases / RNA, Messenger / Reverse Transcriptase Polymerase Chain Reaction / Transcription Factor RelA / Tumor Cells, Cultured
MO Hoque, Hitoshi Kawamaha, Koh-ichi Nakashiro, Fumie Omotehara, Yasuhiro Shinagawa, Satoshi Hino, NM Begum, Daisuke Uchida, Hideo Yoshida, Mitsunobu Sato and Takahiro Fujimori : Dihydropyrimidine dehydrogenase mRNA level correlates with the response to 5-fluorouracil-based chemo-immuno-radiation therapy in human oral squamous cell cancer, International Journal of Oncology, Vol.19, No.5, 953-958, 2001.
(要約)
The measurement of the intra-tumoral level of thymidylate synthetase (TS), and dihydropyrimidine dehydrogenase (DPD), may be useful in predicting tumor sensitivity to 5-fluorouracil (5-FU). In this study, we examined the mRNA levels of DPD and TS in 28 oral squamous cell carcinomas (SCC), and 22 salivary gland tumors by semi-quantitative reverse transcription polymerase chain reaction. Then we examined the correlation of the responsiveness of the patients with oral SCC to 5-FU with the intra-tumoral levels of DPD and TS mRNA. All specimens were obtained at the biopsy before treatment, and then the patients were treated by oral administration of a 5-FU compound (UFT), the irradiation of cobalt-60 (upto 60 Gy) and injection of an immuno-potentiator (OK-432). Intra-tumoral levels of DPD mRNA in the patients who showed CR (complete response) and PR (partial response) were significantly lower than those in the patients who showed NC (no change). However, intra-tumoral levels of DPD mRNA did not correlate with the local recurrence of the tumor during the observation period after initial treatment with or without surgical resection of the residual tumors. On the other hand, TS mRNA levels in the tumors did not correlate with any clinico-pathological parameters. These observations suggest that intra-tumoral levels of DPD mRNA may predict the tumor response to 5-FU-based chemo-immuno-radiation therapy in the patients with oral SCC.
Daisuke Uchida, Hitoshi Kawamata, Fumie Omotehara, Koichi Nakashiro, Tetsuo Kimura-Yanagawa, Satoshi Hino, Nasima Mila Begum, MO Hoque, Hideo Yoshida, Mitsunobu Sato and Takahiro Fujimori : Role of HGF/C-MET system in invasion and metastasis of oral squamous cell carcinoma cells in vitro and its clinical significance, International Journal of Cancer, Vol.93, No.4, 489-496, 2001.
(要約)
We examined the role of the hepatocyte growth factor (HGF)/c-met system on invasion and metastasis of oral squamous cell carcinoma (SCC) cells. In monolayer culture, exogenous HGF marginally affected the growth of oral SCC cells (BHY, HN, IH) and human gingival epithelial cells (GE). In type I collagen matrix, however, HGF significantly enhanced the invasive growth of the cancer cells (p < 0.05). We detected the expression of c-met (HGF receptor) mRNA in all of the cancer cells, but not in human gingival fibroblasts (GF). Oral SCC cells did not secret HGF protein into the medium, but GF secreted a large amount of HGF protein (15 ng/ml). Furthermore, HGF markedly enhanced the migration of cancer cells in a Transwell invasion chamber. Then, we examined the serum levels of HGF in oral SCC patients, or HGF concentrations in oral cancer tissues. Serum levels of HGF in the patients were significantly higher than those in healthy volunteers (p < 0.05). After initial treatment, all of the tumor-free survivors showed a marked decline in the serum HGF levels. Furthermore, HGF concentrations in metastatic cancer tissues were significantly higher than those of non-metastatic cancer tissues and normal gingiva (p < 0.01). These results suggest that HGF plays an important role in invasion and metastasis of oral SCC cells as a paracrine factor, and an elevated HGF level in the cancer tissue can be a predictive marker for metastasis formation in patients with oral SCC.
Daisuke Uchida, Hitoshi Kawamata, Koichi Nakashiro, Fumie Omotehara, Satoshi Hino, MO Hoque, Nasima Mila Begum, Hideo Yoshida, Mitsunobu Sato and Takahiro Fujimori : Low-dose retinoic acid enhances in vitro invasiveness of human oral squamous-cell-carcinoma cell lines by up-regulation of tissue-type plasminogen activator and activation of pro-matrix metalloproteinases, British Journal of Cancer, Vol.85, No.1, 122-128, 2001.
(要約)
Retinoids inhibit the proliferation of several types of tumour cells, and are used for patients with several malignant tumours. In this study, we examined the effect of retinoic acids (RAs) on the invasive potentials of the oral squamous cell carcinoma (SCC) cells, BHY and HNt. BHY cells expressed all of retinoid nuclear receptors (RARalpha, beta, gamma, and RXRalpha) and cytoplasmic retinoic acid binding proteins (CRABP1 and CRABP2). HNt cells lacked the expression of RARbeta, but expressed other nuclear receptors and CRABPs. All-trans retinoic acid (ATRA) and 13-cis retinoic acid (13-cisRA) (10(-6)and 10(-7)M) inhibited the growth of the cells, but low-dose ATRA and 13-cisRA (10(-8)M) marginally affected the growth of the cells. Surprisingly, low-dose RAs enhanced the activity of tissue-type plasminogen activator (tPA), and activated pro-matrix metalloproteinases (proMMP2 and proMMP9). Activation of proMMP2 and proMMP9 was inhibited by aprotinin, a serine-proteinase, tPA inhibitor. Furthermore, low-dose RAs enhanced the in vitro invasiveness of BHY cells. These results indicate that low-dose RAs enhances the in vitro invasiveness of oral SCC cells via an activation of proMMP2 and proMMP9 probably mediated by the induction of tPA.
Satoshi Hino, Hitoshi Kawamata, Daisuke Uchida, Fumie Omotehara, Yoshihiro Miwa, Mila Nasima Begum, Hideo Yoshida and Mitsunobu Sato : Nuclear translocation of TSC-22 (TGF-beta-stimulated clone-22) concomitant with apoptosis: TSC-22 as a putative transcriptional regulator, Biochemical and Biophysical Research Communications, Vol.278, No.3, 659-664, 2000.
(要約)
We examined the alteration of the subcellular localization of TSC-22 (TGF-beta-stimulated clone-22) after induction of apoptosis and the transcription-regulatory activity of TSC-22. In the living cells, TSC-22-green fluorescent protein (GFP) fusion protein was clearly localized to the cytoplasm, however, in the apoptotic cells, the TSC-22-GFP fusion protein was translocated from the cytoplasm to the nucleus. TSC-22 fused to GAL4-DNA binding domain (GAL4BD) did not show the transcriptional activity on the reporter genes in yeast and in HSG (salivary gland cancer cells) and Hela. However, in CHO cells, TSC-22-GAL4BD fusion protein strongly activated the reporter gene. The transcriptional activity of the leucine zipper structure of TSC-22 is greater than that of the full-length TSC-22. These findings suggest that after receiving the apoptotic stimuli, TSC-22 translocates from the cytoplasm to the nucleus and shows the transcription-regulatory activity.
Masato Okamoto, Kenji Hiura, Go Ohe, Yasuo Oba, Kunihiro Terai, Tetsuya Oshikawa, Sachiko Furuichi, Hidetomo Nishikawa, Keiji Moriyama, Hideo Yoshida and Mitsunobu Sato : Mechanism for bone invasion of oral cancer cells mediated by interleukin-6 in vitro and in vivo, Cancer, Vol.89, No.9, 1966-1975, 2000.
(要約)
Osteoclastic bone resorption is an important step in bone invasion in several malignancies. Although interleukin (IL)-6 accelerates osteoclastic bone resorption, it remains unclear whether IL-6 may be involved in bone invasion of oral cancer. The pit formation assay with calf femur-derived bone slices was performed to examine the bone-resorbing activity of osteoclasts and cancer cells. The chemotaxis activity of the culture media was analyzed by the use of Boyden chamber technique. Nude mice, which were inoculated with IL-6-producing oral cancer cells into masseter, were treated with anti-IL-6 neutralizing antibody, and mandibular-bone invasion of the cells was assessed. BHY, a bone-invasive oral cancer cell line, but not HNT, a noninvasive cell line, produced large amounts of IL-6. In a pit formation assay, addition of conditioned medium (CM) derived from BHY but not HNT increased osteoclastic bone resorption, and the effects were inhibited by anti-IL-6 antibody. BHY-secreted IL-6 showed significant chemotaxis activity for osteoclasts. Of note, CM from the cocultivation of osteoclasts and BHY markedly enhanced the cancer cell migration, and the chemotaxis activity was significantly reduced when anti-IL-6 antibody was added into the coculture and then CM were collected, but not when the antibody was added into the CM after they were collected. Furthermore, treatment with anti-IL-6 antibody almost completely inhibited mandibular bone invasion of BHY in nude mice. These results strongly suggest that IL-6 secreted by oral cancer cells plays a significant role in bone invasion.
Masayuki Azuma, Tetsuya Tamatani, Katsumi Motegi, Keiko Aota, Tsuyoshi Yamashita, Kouji Harada, Yoshio Hayashi and Mitsunobu Sato : Suppression of tumor necrosis factor-a-induced matrix metalloproteinase-9-production by introduction of a super-repressor form of IkB-cDNA into immotalized human salivary gland acinar cells, Arthritis and Rheumatism, Vol.43, No.8, 1756-1767, 2000.
(要約)
We have previously shown that specific enhancement in acinar cells of proteolytic activity induced by tumor necrosis factor alpha (TNFalpha) may be responsible for the destruction of the acinar structure in the salivary glands of patients with Sjögren's syndrome. Because matrix metalloproteinase 9 (MMP-9) is regulated by nuclear factor kappaB (NF-kappaB), we investigated the effect of a super-repressor form of inhibitor of nuclear factor kappaBalpha (srIkappaBalpha) on the suppression of TNFalpha-induced MMP-9 production in acinar cells. Two srIkappaBalpha complementary DNA (cDNA)-transfected acinar cell clones (ACMT-6 and ACMT-7) and 1 empty vector-transfected cell clone (ACpRc-1) were established. After treatment of cell clones with TNFalpha, the expression of MMP-9 was examined. In addition, the effect of TNFalpha on cell growth and the morphogenetic behavior of cell clones cultured on type IV collagen-coated dishes were examined. TNFalpha induced the production of MMP-9 in the ACpRc-1 cell clone, but greatly suppressed MMP-9 production in ACMT-6 and ACMT-7 clones. No apparent cytotoxic effect of TNFalpha treatment was observed in these cell clones. When ACpRc-1 cells were seeded on type IV collagen-coated dishes in the presence of both TNFalpha and plasmin, type IV collagen interaction with the cells was lost and the cells entered apoptosis. However, even when ACMT-6 and ACMT-7 cells were cultured under the same culture conditions as those for ACpRc-1, these cell clones attached to the substrate and grew consistently without showing apoptosis. Conclusion. These observations indicate that suppression of TNFalpha-induced MMP-9 production by the introduction of srIkappaBalpha cDNA corrected the aberrant in vitro morphogenesis of acinar cells grown on type IV collagen.
Keiko Aota, Masayuki Azuma, Tsuyoshi Yamashita, Testuya Tamatani, Katsumi Motegi, Naozumi Ishimaru, Yoshio Hayashi and Mitsunobu Sato : 5-Fluorouracil Induces Apoptosis through the Suppression of NF-κB Activity in Human Salivary Gland Cancer Cells, Biochemical and Biophysical Research Communications, Vol.273, No.3, 1168-1174, 2000.
(要約)
Activation of the transcription factor NF-kappaB results in protection against apoptosis, and the chemotherapeutic agent 5-Fluorouracil (5-FU) exerts its cytotoxic effect through the induction of apoptosis. Thus, we examined whether 5-FU could induce apoptosis through the suppression of NF-kappaB activity. We found that upon treatment of a human salivary gland cancer cell line (cl-1) with 5-FU, the NF-kappaB activity was suppressed in a time-dependent manner. This inhibition was mediated by a prevention of the degradation of the inhibitory IkappaB-alpha protein. In addition, the expression of TRAF-2 and cIAP-1, which are transcriptionally regulated by NF-kappaB and function as anti-apoptotic molecules through the interruption of caspase pathway, was also inhibited by 5-FU. Finally, the activity of caspase-8 and caspase-3 showed a significant increase in response to 5-FU. By flow cytometric analysis, 5-FU did not affect the expression level of Fas on the cell surface. Thus, our results suggest that one of the molecular mechanisms involved in 5-FU-induced apoptosis in cl-1 cells may be due to the suppression of NF-kappaB activity, resulting in the activation of the pro-apoptotic pathway.
Fumie Omotehara, Daisuke Uchida, Satoshi Hino, NM Begum, Hideo Yoshida, Mitsunobu Sato and Hitoshi Kawamata : In vivo enhancement of chemosensitivity of human salivary gland cancer cells by overexpression of TGF-beta stimulated clone-22, Oncology Reports, Vol.7, No.4, 737-740, 2000.
(要約)
We have isolated transforming growth factor-beta-stimulated clone-22 (TSC-22) cDNA as an anti-cancer drug-inducible gene in a human salivary gland cancer cell line, TYS. We have previously reported that TSC-22 negatively regulates the growth of TYS cells, and that overexpression of TSC-22 protein in TYS cells enhanced the in vitro chemosensitivity of the cells. In this study, we examined the in vivo chemosensitivity of TSC-22-expressing TYS cells. TSC-22-expressing TYS cells formed tumors in nude mice, but tumors formed by TSC-22-expressing TYS cells were significantly smaller than tumors formed by control cells (p<0.001, one way ANOVA). Furthermore, intraperitoneal injection of 5-fluorouracil (5-FU) markedly inhibited the growth of the TSC-22-expressing TYS tumors, but did not affect the growth of control tumors. It was found by TUNEL assay that TSC-22-expressing TYS tumors were induced to undergo apoptosis by 5-FU treatment. These findings suggest that overexpression of TSC-22 protein in TYS cells enhances the in vivo chemosensitivity of the cells to 5-FU via induction of apoptosis.
Daisuke Uchida, Hitoshi Kawamata, Fumie Omotehara, Yoshihiro Miwa, Satoshi Hino, Mila Nasima Begum, Hideo Yoshida and Mitsunobu Sato : Over-expression of TSC-22 (TGF-beta stimulated clone-22) markedly enhances 5-fluorouracil-induced apoptosis in a human salivary gland cancer cell line, Laboratory Investigation; a Journal of Technical Methods and Pathology, Vol.80, No.6, 955-963, 2000.
(要約)
We have recently isolated TSC-22 (transforming growth factor-beta-stimulated clone-22) cDNA as an anticancer, drug-inducible (with vesnarinone) gene in a human salivary gland cancer cell line, TYS. We have also reported that TSC-22 negatively regulates the growth of TYS cells and that down-regulation of TSC-22 in TYS cells plays a major role in salivary gland tumorigenesis (Nakashiro et al, 1998). In this study, we transfected TYS cells with an expression vector encoding the TSC-22-GFP (green fluorescent protein) fusion protein, and we established TSC-22-GFP-expressing TYS cell clones. Next, we examined (a) the subcellular localization of the fusion protein, (b) the sensitivity of the transfectants to several anticancer drugs (5-fluorouracil, cis-diaminedichloroplatinum, peplomycin), and (c) induction of apoptotic cell death in the transfectants by 5-fluorouracil treatment. The TSC-22-GFP fusion protein was clearly localized to the cytoplasm, but not to the nucleus. Over-expression of the TSC-22-GFP fusion protein did not affect cell growth, but significantly increased the sensitivity of the cells to the anticancer drugs (p < 0.01; one-way ANOVA). Furthermore, over-expression of the TSC-22-GFP fusion protein markedly enhanced 5-fluorouracil-induced apoptosis. These findings suggest that over-expression of TSC-22-GFP protein in TYS cells enhances the chemosensitivity of the cells via induction of apoptosis.
Hitoshi Kawamata, Daisuke Uchida, Ko-ichi Nakashiro, Satoshi Hino, Fumie Omotehara, Hideo Yoshida and Mitsunobu Sato : Haematogenous cytokeratin 20 mRNA as a predictive marker for recurrence in oral cancer patients, British Journal of Cancer, Vol.80, No.3-4, 448-452, 1999.
(要約)
We examined the expression of cytokeratin 20 (CK20) mRNA in the peripheral blood of oral squamous cell carcinoma (SCC) patients by reverse transcriptase polymerase chain reaction (RT-PCR). Eleven out of 12 oral SCC patients showed positive RT-PCR results. However, there is no clear relationship between the haematogenous CK20 mRNA and the metastasis. After initial treatment, all of the tumour-free survivors tested showed negative RT-PCR results. CK20 mRNA in peripheral blood can be used as a marker for tumour recurrence but not not for metastasis in oral SCC patients.
Hitoshi Kawamata, Daisuke Uchida, Hamano Hironori, Tetsuo Kimura-Yanagawa, Ko-ichi Nakashiro, Satoshi Hino, Fumie Omotehara, Hideo Yoshida and Mitsunobu Sato : Active-MMP2 in cancer cell nests of oral cancer patients: correlation with lymph node metastasis, International Journal of Oncology, Vol.13, No.4, 699-704, 1998.
(要約)
We examined the gelatinolytic activity in human oral squamous-cell carcinoma tissues in order to evaluate the capability of intravasation and extravasation of cancer cells. By a microdissection-zymography, we demonstrated separately the gelatinolytic activities in cancer cell nests and stroma adjacent to the cancer cells. The gelatinolytic activities, such as pro-matrix metalloproteinase (MMP)9 and active-MMP2 in most of cancer cell nests were much higher than those of normal gingival epithelium. Moreover, the activities of active-MMP2 in cancer cell nests of metastatic cancers were significantly higher than those of non-metastatic cancers (p<0.05). These results suggest that active-MMP2 in cancer cells can be a predictive marker for metastasis formation in oral squamous-cell carcinoma patients.
Koichi Nakashiro, Hitoshi Kawamata, Satoshi Hino, Daisuke Uchida, Yoshihiro Miwa, Hironori Hamano, Fumie Omotehara, Hideo Yoshida and Mitsunobu Sato : Down-regulation of TSC-22 (transforming growth factor beta-stimulated clone 22) markedly enhances the growth of a human salivary gland cancer cell line in vitro and in vivo, Cancer Research, Vol.58, No.3, 549-555, 1998.
(要約)
We have recently isolated TSC-22 (transforming growth factor beta-stimulated clone 22) cDNA as a new anticancer drug (Vesnarinone)-inducible gene in a human salivary gland cancer cell line, TYS. We conducted the present study to examine whether up-regulation or down-regulation of TSC-22 can affect the growth of TYS cells in vitro and in vivo. We constructed an expression vector containing sense- or antisense-oriented human TSC-22 cDNA under the transcriptional control of the SR alpha promoter. We cotransfected TYS cells with the sense or antisense expression vector and pSV2neo and obtained more than 200 G418-resistant colonies in each sense or antisense transfectant. Approximately 80% of representative G418-resistant clones expressed the transcripts from transfected sense or antisense TSC-22 cDNA. To avoid the clonal heterogeneity of the cells, we mixed all of the G418-resistant colonies together in each sense or antisense transfectant and examined the expression of TSC-22 protein, in vitro growth, and the tumorigenicity in nude mice. The expression of TSC-22 protein was examined by solid-phase ELISA using a specific antibody against recombinant TSC-22 protein. The expression of TSC-22 protein was up-regulated in the sense transfectants and down-regulated in the antisense transfectants. Contrary to our expectation, up-regulation of TSC-22 protein did not affect both in vitro and in vivo growth of TYS cells. However, down-regulation of TSC-22 markedly enhanced the growth of TYS cells in vitro and in vivo. Furthermore, we examined the expression of TSC-22 mRNA in several human salivary gland tumors. The mRNA expression of TSC-22 in benign and malignant salivary gland tumors was significantly decreased when compared to that in tumor-free salivary glands (P < 0.05; one-way ANOVA), and in some salivary gland tumors, the expression of TSC-22 mRNA was not detectable by reverse transcription-PCR. These results suggest that down-regulation of TSC-22 may play a major role on salivary gland tumorigenesis.
Hitoshi Kawamata, Koh-ichi Nakashiro, Daisuke Uchida, Satoshi Hino, Fumie Omotehara, Hideo Yoshida and Mitsunobu Sato : Induction of TSC-22 by treatment with a new anti-cancer drug, vesnarinone, in a human salivary gland cancer cell, British Journal of Cancer, Vol.77, No.1, 71-78, 1998.
(要約)
We undertook the present study to clarify the molecular mechanism of the effect of a new anti-cancer drug, vesnarinone, on a human salivary gland cancer cell line, TYS. We isolated TSC-22cDNA as avesnarinone-inducible gene from a cDNA library constructed from vesnarinone-treated TYS cells. TSC-22 was originally reported as a transforming growth factor (TGF)-beta-inducible gene. The expression of TSC-22 was up-regulated within a few hours after treatment with vesnarinone and was continued for 3 days. The level of TSC-22 mRNA in TYS cells was continuously increased until the cells reached confluency. Furthermore, the induction of TSC-22 by vesnarinone was inhibited by treatment with cycloheximide. When we treated the cells with an antisense oligonucleotide against TSC-22 mRNA under quiescent conditions, the antisense oligonucleotide stimulated the growth of TYS cells; however, under growing conditions the antisense oligonucleotide did not affect cell growth. Furthermore, the antisense oligonucleotide suppressed the antiproliferative effect of vesnarinone. These results suggest that TSC-22 may be a negative growth regulator and may play an important role in the antiproliferative effect of vesnarinone.
Masayuki Azuma, Katsumi Motegi, Keiko Aota, Yoshio Hayashi and Mitsunobu Sato : Role of cytokines in the destruction of acinar structure in Sjogren's syndrome salivary glands, Laboratory Investigation; a Journal of Technical Methods and Pathology, Vol.77, No.3, 269-280, 1997.
49.
Mitsunobu Sato, Koji Harada, Hideo Yoshida, Yura Yoshiaki, Masayuki Azuma, Hiroki Iga, Takashi Bando, Kawamata Hitoshi and Yoshihiro Takegawa : Therapy for oral squamous cell carcinoma by tegafur and streptococcal agent OK-432 in combination with radiotherapy : Association of the therapeutic effect with differentiation and apoptosis in the cancer cells, Apoptosis, Vol.2, No.2, 227-238, 1997.
(要約)
Twenty patients with oral squamous cell carcinoma having mainly stage II or III lesions without distant metastasis, were treated with tegafur and streptococcal agent, OK-432, in combination with radiotherapy. As a consequence, 16 cases among the treated 20 cases showed complete remission by this therapy alone. Especially, we have found that the squamous cell carcinoma arising in non-keratinizing oral epithelium rather than in keratinizing oral epithelium has better response to this therapy. Among the 16 cases with complete remission (CR) by the current therapy, 10 cases were histopathologically diagnosed as well-differentiated squamous cell carcinoma and six cases as moderately differentiated squamous cell carcinoma. When we examined immunohistochemically the expression of various antigens such as proliferating cell nuclear antigen (PCNA), p53 and LeY or the presence of DNA fragmentation by nick-end labelling in the biopsy materials taken at the first visit to our clinic from 20 patients treated with the current therapy, the CR group showed a significantly increased LeY expres-sion level ( p< 0.05) and DNA fragmentation rate (p< 0.05) as compared with the partial response (PR, n= 3) + no change (NC, n= 1) group. On the other hand, the CR group with respect to PCNA expression level was significantly decreased as compared with the PR + NC group ( p< 0.05). From these findings, it can be considered that the therapy for oral squamous cell carcinoma by UFT and OK-432 in combination with radiotherapy is very effective, which may be associated with differentiation or apoptosis in oral squamous carcinoma cells. In addition, we present the clinical findings and results of immunohistochemical staining for the biopsy materials obtained from four CR cases treated with the current therapeutic method.
Hitoshi Kawamata, Ko-ichi Nakashiro, Daisuke Uchida, Satoshi Hino, Koji Harada, Hideo Yoshida and Mitsunobu Sato : Possible contribution of active MMP2 to lymph-node metastasis and secreted cathepsin L to bone invasion of newly established human oral-squamous-cancer cell lines, International Journal of Cancer, Vol.70, No.1, 120-127, 1997.
(要約)
We have established human oral-squamous-cancer cell lines, BHY and HN, derived from non-metastatic cancer and metastatic cancer respectively. We examined the expression of matrix-degrading enzymes and their inhibitors in these cell lines. Both cell lines expressed pro-matrix metalloproteinase (MMP)1, proMMP2, proMMP9, membrane-type MMP and urokinase-type plasminogen activator. In addition to these enzymes, BHY cells secreted proMMP7 and procathepsin L, while HN cells secreted a large amount of active MMP2. BHY cells secreted a tissue inhibitor of matrix metalloproteinase, TIMP2, but only a trace level of TIMP1. Contrary to BHY cells, HN cells secreted TIMP1, but only a trace level of TIMP2. When we inoculated these cells into the masseter muscle of nude mice, both types of cell formed solid tumors, whose microscopic appearance was identical to that of the original tumors. BHY tumors were highly differentiated squamous-cell carcinomas, and invasive to the masseter muscle and the mandibular bone. Despite their local aggressiveness, BHY tumors did not metastasize to any distant organs. HN tumors were poorly differentiated squamous-cell carcinomas, weakly invasive to the muscle, but not to the mandibular bone. However, HN tumors frequently metastasized to cervical lymph nodes. These results suggest that the net activity of MMP2 (active MMP2/TIMP2) and cathepsin L secreted from cancer cells may contribute respectively to lymph-node metastasis and to bone invasion by oral cancer cells.
Masayuki Azuma, Katsumi Motegi, Keiko Aota, Yoshio Hayashi and Mitsunobu Sato : Role of cytokines of acinar structure in Sjogren's syndrome salivary glands, Laboratory Investigation; a Journal of Technical Methods and Pathology, Vol.77, No.3, 269-280, 1997.
Mitsunobu Sato, Hideo Yoshida, Kali Ryoli, Masato Okamoto, Hiroki Iga, Kawamata Hitoshi, Kobayasi Shosuke, Aladib Walid, Yanagawa Tetsuo and Yoshihiro Takegawa : Induction of Bone Formation in an Adenoid Cystic Carcinoma of the Maxillary Sinus by Adoptive Immunotherapy Involving Intra-arterial Injection of Lymphokine-Activated Killer Cells and Recombinant Interleukin-2 in Combination with Radiotherapy, Journal of Biological Response Modifiers, Vol.9, No.3, 329-334, 1990.
(要約)
A patient with an adenoid cystic carcinoma of the maxillary sinus was treated by adoptive immunotherapy involving intra-arterial injection of lymphokine-activated killer cells (a total number of 7 x 10(7) cells) and recombinant interleukin-2 (a total dose of 2.75 x 10(7) units) in combination with radiotherapy (60Co; total irradiation dose of 5,000 rads). Consequently, it was found that bone formation was induced in the treated tumor, which was then replaced completely by lamellar bone tissue with myxomatous stroma. This finding indicates that the tumor cells composing certain adenoid cystic carcinoma can be converted into normal-appearing, bone-forming cells by current therapy and this differentiation phenomenon leads to cure of the tumor.
(キーワード)
Biopsy / Bone Development / Carcinoma, Adenoid Cystic / Combined Modality Therapy / Drug Administration Schedule / Humans / Injections, Intra-Arterial / Interleukin-2 / Killer Cells, Lymphokine-Activated / Male / Maxillary Sinus Neoplasms / Middle Aged / Paranasal Sinus Neoplasms
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 2166140
Yoshio Hayashi, Tetsuo Yanagawa, Hideo Yoshida, Masayuki Azuma, Toshinobu Nishida, Yoshiaki Yura and Mitsunobu Sato : Expression of vasoactive intestinal polypeptide and amylase in a human parotid gland adenocarcinoma cell line in culture, Journal of the National Cancer Institute, Vol.79, No.5, 1025-1037, 1987.
57.
Yoshio Hayashi, Shinichi Nagamine, Tetsuo Yanagawa, Hideo Yoshida, Yoshiaki Yura, Masayuki Azuma and Mitsunobu Sato : Small cell undifferentiated carcinoma of the minor salivary gland containing exocrine, neuroendocrine, and squamous cells, Cancer, Vol.60, No.7, 1583-1588, 1987.
58.
Yoshio Hayashi, Hideo Yoshida, Shinichi Nagamine, Tetsuo Yanagawa, Yoshiaki Yura, Masayuki Azuma and Mitsunobu Sato : Induction of cells with acinar cell phenotype including presence of intracellular amylase, --- Treatment with 12-O-tetradecanoyl-phorbol-13-acetate in a neoplastic human salivary intercalated duct cell line grown in athymic nude mice ---, Cancer, Vol.60, No.5, 1000-1008, 1987.
59.
Yoshio Hayashi, Toshinobu Nishida, Hideo Yoshida, Tetsuo Yanagawa, Yoshiaki Yura and Mitsunobu Sato : Immunoreactive vasoactive intestinal polypeptide in acinic cell carcinoma of the parotid gland, Cancer, Vol.60, No.5, 962-968, 1987.
60.
Masayuki Azuma, Furumoto Nanayo, Kawamata Hitoshi, Hideo Yoshida, Yanagawa Tetsushi, Yura Yoshiaki, Yoshio Hayashi, Yoshihiro Takegawa and Mitsunobu Sato : The relation of ras oncogene product p21 expression to clinocopathological status criteria and clinical outcome in squamous cell head and neck cancer, The Cancer Journal, Vol.1, No.9, 375-380, 1987.
61.
Tetsuo Yanagawa, Yoshio Hayashi, Hideo Yoshida, Yoshiaki Yura, Shinichi Nagamine, Takashi Bando and Mitsunobu Sato : An adenoid squamous carcinoma-forming cell line established from an oral keratinizing squamous cell carcinoma expressing carcinoembryonic antigen, The American Journal of Pathology, Vol.124, No.3, 496-509, 1986.
Yoshio Hayashi, Haruhiko Saito, Shiro Saito, Tetsuo Yanagawa, Hideo Yoshida, Yoshiaki Yura and Mitsunobu Sato : Immunoreactive somatostatin in Warthin's tumor, The American Journal of Pathology, Vol.123, No.2, 250-255, 1986.
64.
Yoshio Hayashi, Yoshiaki Yura, Hideo Yoshida, Tetsuo Yanagawa and Mitsunobu Sato : Development of allergic sialadenitis in mice immunized with mumps virus-infected submandibular salivary gland, The American Journal of Pathology, Vol.123, No.2, 271-279, 1986.
65.
Yoshio Hayashi, Hideo Yoshida, Furumoto Nanako, Yanagawa Tetsuo, Yura Yoshiaki, Yoshihiro Takegawa and Mitsunobu Sato : Monoclonal antibody analysis of peripheral blood lymphocyte subpopulations in squamous cell head and neck cancer, The Cancer Journal, Vol.1, No.1, 25-29, 1986.
66.
Masayuki Azuma, Yoshio Hayashi, Hideo Yoshida, Tetsuo Yanagawa, Yoshiaki Yura, Akemichi Ueno and Mitsunobu Sato : Emergence of differentiated subclones from a human salivary adenocarcinoma cell clone in culture after treatment with sodium butyrate, Cancer Research, Vol.46, No.2, 770-777, 1986.
67.
Tetsuo Yanagawa, Yoshio Hayashi, Toshinobu Nishida, Hideo Yoshida, Yoshiaki Yura, Masayuki Azuma and Mitsunobu Sato : Immunohistochemical demonstration of carcinoembryonic antigen (CEA) on tissue sections from squamous cell head and neck cancer and plasma CEA levels of the patients, International Journal of Oral and Maxillofacial Surgery, Vol.15, No.3, 296-306, 1986.
M Urata, Toshinobu Nishida, Yoshio Hayashi, Hideo Yoshida, N Furumoto, T Yanagawa, Y Yura and Mitsunobu Sato : Immunoglobulin A circulating immune complexes in oral squamous cell cancer, The Cancer Journal, Vol.1, No.2, 68-71, 1986.
69.
Mitsunobu Sato, Hideo Yoshida, Yoshio Hayashi, Kenji Miyakami, Takashi Bando, Tetsuo Yanagawa, Yura Yura, Masayuki Azuma and Akemichi Ueno : Expression of epidermal growth factor and transforming growth factor-beta in human salivary gland adenocarcinoma cell line, Cancer Research, Vol.45, No.12 P 1, 6160-6167, 1985.
70.
Mitsunobu Sato, Yoshio Hayashi, Tetsuo Yanagawa, Hideo Yoshida, Yoshiaki Yura, Masayuki Azuma and Akemichi Ueno : Intermediate-sized filaments and specific markers in a human salivary gland adenocarcinoma cell line and its nude mouse tumors, Cancer Research, Vol.45, No.8, 3837-3790, 1985.
71.
Yoshio Hayashi, Tetsuo Yanagawa, Hideo Yoshida, Yoshiaki Yura, Toshiharu Nitta and Mitsunobu Sato : Induction of other differentiation stages in neoplastic epithelial duct and myoepithelial cells from the human salivary gland grown in athymic nude mice, Cancer, Vol.55, No.11, 2575-2583, 1985.
72.
Yoshio Hayashi, Mitsunobu Sato and Katsuiku Hirokawa : Induction of experimental allergic sialadenitis in mice, The American Journal of Pathology, Vol.118, No.3, 476-483, 1985.
Mitsunobu Sato, Hideo Yoshida, Yanagawa Tetsuo, Yura Yoshiaki, Urata Mitsuru, Masayuki Azuma, Furumoto Nanayo, Yoshio Hayashi and Yoshihiro Takegawa : Interferon Activity and Its Characterization in the Sera of Patients with Head and Neck Cancer, Cancer, Vol.54, No.1, 1239-1251, 1984.
Yoshio Hayashi, Toshinobu Nishida, Hideo Yoshida, Tetsuo Yanagawa, Yoshiaki Yura, Nanayo Furumoto and Mitsunobu Sato : Peripheral Tγ lymphocyte population in head and neck cancer, Cancer Immunology, Immunotherapy, Vol.17, No.3, 160-164, 1984.
(要約)
Peripheral T gamma lymphocytes were measured in head and neck cancer patients and controls. The percentage was significantly higher in the 59 cancer patients than in the 46 normal controls (P less than 0.001). The 12 patients with recurrent disease had elevated percentages of T gamma lymphocytes compared with the untreated group (n = 31; P less than 0.05) and the treated, disease-free group (n = 16; P less than 0.05). Moreover, the percentage of T gamma lymphocytes was significantly higher in the 31 patients with regional lymph node metastasis than in the node-negative group (n = 28; P less than 0.05). In a total of 37 patients with squamous cell carcinoma histologically graded I, II, and III, the absolute counts and percentages of T gamma lymphocytes in the grade I group (n = 13) showed significant decreases compared with those in the grade III group (P less than 0.05; n = 6). Moreover, postoperative serial determinations of the percentage of T gamma lymphocytes in the 14 treated, disease-free patients revealed a gradual decrease of T gamma lymphocytes, whereas the five patients with recurrent disease had a tendency to increases in the percentage of T gamma lymphocytes.
Yoshio Hayashi, Toshinobu Nishida, Hideo Yoshida, Tetsuo Yanagawa, Yoshiaki Yura, Toshiharu Nitta and Mitsunobu Sato : Peripheral K-lymphocyte population in head and neck cancer, Cancer, Vol.53, No.11, 2507-2514, 1984.
(要約)
Peripheral K-lymphocytes in head and neck cancer were measured by assay of plaque forming-cells. In the cancer patients, the percentage (4.37 +/- 0.87%, mean +/- SD, N = 42, P less than 0.01) and absolute counts (68 +/- 28/mm3, P less than 0.01) of K-lymphocytes were significantly lower than those in normal controls (8.04 +/- 1.41%; 175 +/- 53/mm3, N = 29). The untreated group showed decreased K-lymphocyte counts (63 +/- 22/mm3, N = 13) as compared with the treated, disease-free group (83 +/- 27/mm3, N = 13, P less than 0.05). There were significant correlations between absolute counts of K-lymphocytes and T-cells or B-cells within the untreated group (r = 0.79 in T-cells, P less than 0.01; r = 0.64 in B cells, P less than 0.01). Moreover, absolute counts and percentage of K-lymphocytes in the patients having regional lymph node metastasis (58 +/- 25/mm3; 4.14 +/- 0.77%, N = 22) were significantly lower than those in the negative node group (80 +/- 26/mm3, P less than 0.01; 4.63 +/- 0.91%, P less than 0.05; N = 20). In a total of 25 patients with squamous cell carcinoma who were grouped into grade I, II, and III according to classification of the histologic differentiation of the World Health Organization, the absolute counts and percentage of K-lymphocytes in the grade I group (93 +/- 33/mm3; 5.14 +/- 1.08%, N = 7) showed significant increases in comparison to those in the grade II group (62 +/- 19/mm3, P less than 0.02; 4.06 +/- 0.62%, P less than 0.02, N = 14). Moreover, the change of the K-lymphocyte population in the treated, disease-free eight patients revealed a gradual increase of K-lymphocytes. These results led us to suggest that the measurement of peripheral K-lymphocytes is a useful method of characterizing host defense in head and neck cancer.
Mitsunobu Sato, Yoshio Hayashi, Hideo Yoshida, Tetsuo Yanagawa and Yoshiaki Yura : A family with hereditary lack of T4+ inducer/helper T cell subsets in peripheral blood lymphocytes, The Journal of Immunology, Vol.132, No.3, 1071-1073, 1984.
Mitsunobu Sato, Hideo Yoshida, Yanagawa Tetsuo, Yura Yoshiaki, Urata Mitsuru, Atsumi Masahiro, Yoshio Hayashi and Yoshihiro Takegawa : Effects of intradermal administration of streptococcal preparation OK-432 on interferon and natural killer cell activities in patients with oral cancer, International Journal of Oral Surgery, Vol.13, No.1, 7-15, 1984.
Masato Okamoto, S Furuichi, Yasuhiko Nishioka, T Oshikawa, G Ohe, H Nishikawa, T Tano, U S Armed, Y Moriya, T Matsubara, M Saito, Saburo Sone and Mitsunobu Sato : Maturation and activation of human dendritic cells in vitro by lipoteichoic acid-related molecule isolated from OK-432, a streptococcal agent: Involvement of Toll-like receptor 4, American Association for Cancer Research (AAGR), USA, May 2002.
Yoshio Hayashi, Toshinobu Nishida, Tetsuo Yanagawa, Hideo Yoshida and Mitsunobu Sato : Expression of carcinoemryonic antigen in the neoplastic cells originated from human salivary gland, Tissue Culture in Dentistry, Vol.19, 27-28, 1982.
13.
Yoshio Hayashi, Toshinobu Nishida, Toshiharu Nitta, Tetsuo Yanagawa, Hideo Yoshida and Mitsunobu Sato : Peripheral k lymphocytes as an immunologic parameter in the patients with oral cancer, Tissure Culture in Dentistry, Vol.19, 25-26, 1982.