Hajime Sunakawa

Mode of invasion, bleomycin sensitivity, and clinical course in squamous cell carcinoma of the oral cavity

Forty patients with squamous cell carcinoma of the oral cavity were treated with bleomycin prior to undergoing surgery. The degree of the clinical effect of bleomycin and the postoperative clinical course of each case were estimated from the viewpoint of correlation with the mode of invasion. A strong correlation was found among the mode of invasion, bleomycin sensitivity, and clinical course. A slight effect of bleomycin and poor prognosis existed in the group with a diffuse invasion of mode of invasion, while the greatest effect of bleomycin and good clinical course were achieved in the group with a welldefined tumor‐host borderline.

Multivariate analysis of occult lymph node metastasis as a prognostic indicator for patients with squamous cell carcinoma of the oral cavity

The biologic aggressiveness of squamous cell carcinoma of the oral cavity is reflected in its ability to metastasize to regional cervical lymph nodes. Patients with clinically negative cervical lymph nodes are believed to have a good prognosis; however, the prognosis of patients with lymph node metastasis occurring after excision or radiotherapy of the primary tumor is poor.

Efficient Cross-Presentation by Heat Shock Protein 90-Peptide Complex-Loaded Dendritic Cells via an Endosomal Pathway

It is well-established that heat shock proteins (HSPs)-peptides complexes elicit antitumor responses in prophylactic and therapeutic immunization protocols. HSPs such as gp96 and Hsp70 have been demonstrated to undergo receptor-mediated uptake by APCs with subsequent representation of the HSP-associated peptides to MHC class I molecules on APCs, facilitating efficient cross-presentation. On the contrary, despite its abundant expression among HSPs in the cytosol, the role of Hsp90 for the cross-presentation remains unknown. We show here that exogenous Hsp90-peptide complexes can gain access to the MHC class I presentation pathway and cause cross-presentation by bone marrow-derived dendritic cells. Interestingly, this presentation is TAP independent, and followed chloroquine, leupeptin-sensitive, as well as cathepsin S-dependent endosomal pathways. In addition, we show that Hsp90-chaperoned precursor peptides are processed and transferred onto MHC class I molecules in the endosomal compartment. Furthermore, we demonstrate that immunization with Hsp90-peptide complexes induce Ag-specific CD8+ T cell responses and strong antitumor immunity in vivo. These findings have significant implications for the design of T cell-based cancer immunotherapy.

Human papillomavirus DNA in adenosquamous carcinoma of the lung.

AIM: To investigate the presence of human papillomavirus (HPV) DNA in adenosquamous carcinoma of the lung--which is relatively common in Okinawa but not in mainland Japan--and examine its histological features. METHODS: Of 207 cases where primary lung cancers were surgically removed between January 1995 and June 1997 in Okinawa, 23 were adenosquamous carcinoma. HPV was detected by non-isotopic in situ hybridisation (NISH) and polymerase chain reaction (PCR) amplification with primers specific for E6 and E7 regions of the HPV genome. PCR products were analysed by Southern blotting. Immunohistochemical determination of high molecular weight cytokeratin (HMC) and involucrin was also carried out. RESULTS: 18 cases were positive for HPV DNA by PCR and NISH. HPV types 6, 11, 16, and 18 were found. Seven cases were dual positive for different types of HPV. Using NISH, HPV was also found in the squamous cell components and in neighbouring enlarged adenocarcinoma cells. The HMC and involucrin were demonstrated immunohistochemically in the same areas. CONCLUSIONS: HPV DNA was found in a high proportion (78.3%) of adenosquamous carcinomas in Okinawa, a region where HPV has previously been shown to be prevalent in squamous cell carcinoma of the lung. The adenocarcinoma cells adjacent to the squamous cell carcinoma component were enlarged and positive for HPV, HMC, and involucrin. This is thought to indicate the transition from adenocarcinoma to squamous cell carcinoma.

Tumor-derived heat shock protein 70-pulsed dendritic cells elicit tumor-specific cytotoxic T lymphocytes (CTLs) and tumor immunity

Vaccination with autologous tumor‐derived heat shock proteins (Hsp), such as Hsp70, Hsp90 and gp96, has been demonstrated to elicit specific immune responses against the tumor from which the Hsps were isolated. The effect of Hsp immunization is wholly dependent on the presence of functional antigen‐presenting cells (APCs) in the immunized host, and Hsp receptors on APCs have recently been identified. Here we show that bone marrow‐derived dendritic cells (DCs) are able to internalize HSP‐peptide complex and that peptides are re‐presented by DCs via the major histocompatibility complex (MHC) class I presentation pathway. In addition, immunization with tumor‐derived HSP‐pulsed DCs induces strong cytotoxic T cell (CTL) responses against multiple antigenic peptides in a transporter‐associated antigen processing (TAP)‐dependent manner. The results of the present study provide strong evidence of an efficient cross‐priming activity of Hsp70, which could be exploited in the development of new and more effective immunotherapeutic strategies for cancer patients. (Cancer Sci 2004; 95: 248–253)

INTRAOPERATIVE RADIATION THERAPY (IORT) FOR HEAD AND NECK CANCER

PURPOSE To determine the efficacy of intraoperative radiation therapy (IORT) for patients with advanced or recurrent head and neck cancer. METHODS AND MATERIALS Intraoperative radiation therapy was given at 30 sites in 25 patients using a 6-18 MeV electron beam with or without conventional external beam irradiation. A single dose of 10-30 Gy was delivered after surgical resection. Sites treated with IORT were classified into three types after surgical resection: gross residual disease (GR, n = 7), microscopic residual disease (MR, n = 12), and close margin (CM, n = 11). Local control rate, patterns of recurrence, survival rate, and complications were analyzed. RESULTS The 2-year cumulative local control rate within the IORT port was 54.1% for all cases, 0% for GR, 54.5% for MR, and 81.8% for CM. There were significant differences between GR and MR (p < 0.05), and GR and CM (p < 0.01). The majority of the failures inside the IORT port were associated with recurrence outside the port. Distant metastases occurred in five patients. Four of these had GR. The 2-year cumulative survival rate was 45.1% for all, 0% for GR, 33.0% for MR, and 70.0% for CM. Five patients (22%) experienced late complications. The 2-year cumulative complication rate was 32.8%. Four sites developed osteoradionecrosis and three developed carotid artery blowout. Incidence of complications increased when patients received over 20 Gy with a single dose of IORT. CONCLUSIONS Considering both therapeutic ratio and patterns of failure, it is not suitable to treat patients with gross residual disease with IORT. We could not firmly determine the therapeutic value of IORT for patients with microscopic residual disease and close margin. For this subset, further study of moderate dose (less than 20 Gy) IORT combined with adequate postoperative irradiation is needed.

Novel function of glutathione transferase in rat liver mitochondrial membrane: role for cytochrome c release from mitochondria.

Microsomal glutathione transferase (MGST1) is activated by oxidative stress. Although MGST1 is found in mitochondrial membranes (mtMGST1), there is no information about the oxidative activation of mtMGST1. In the present study, we aimed to determine whether mtMGST1 also undergoes activation and about its function. When rats were treated with galactosamine/lipopolysaccharide (GalN/LPS), mtMGST1 activity was significantly increased, and the increased activity was reduced by the disulfide reducing agent dithiothreitol. In mitochondria from GalN/LPS-treated rats, disulfide-linked mtMGST1 dimer and mixed protein glutathione disulfides (glutathionylation) were detected. In addition, cytochrome c release from mitochondria isolated from GalN/LPS-treated rats was observed, and the release was inhibited by anti-MGST1 antibodies. Incubation of mitochondria from control rats with diamide and diamide plus GSH in vitro resulted in dimer- and mixed disulfide bond-mediated activation of mtMGST1, respectively. The activation of mtMGST1 by diamide plus GSH caused cytochrome c release from the mitochondria, and the release was prevented by treatment with anti-MGST1 antibodies. In addition, diamide plus GSH treatment caused mitochondrial swelling accompanied by cytochrome c release, which was inhibited by cyclosporin A (CsA) and bongkrekic acid (BKA), inhibitors of the mitochondrial permeability transition (MPT) pore. Furthermore, mtMGST1 activity was also inhibited by CsA and BKA. These results indicate that mtMGST1 is activated through mixed disulfide bond formation that contributes to cytochrome c release from mitochondria through the MPT pore.

Epstein–Barr virus (EBV)-related oral squamous cell carcinoma in Okinawa, a subtropical island, in southern Japan—simultaneously infected with human papillomavirus (HPV)

Up to now, many authors have reported on the EBV infection and its carcinogenic importance in undifferentiated nasopharyngeal carcinoma (WHO classification, type III), but the infection of the virus in well differentiated oral squamous cell carcinoma has not been well described. We introduce the EBV-related well differentiated oral squamous cell carcinomas in Okinawa, a subtropical island in the southernmost part of Japan. This study aimed to clarify the pathogenesis of this malignancy in this area by carrying out analysis of the histology and the Epstein-Barr (EBV) and human papillomavirus (HPV) infection. In the Department of Oral Surgery, Ryukyu University Hospital Okinawa, 188 cases of oral malignant tumours were encountered from 1996 to 2000. The histopathological examination and the sequence analysis of LMP-1 carboxy terminal region and EBNA2 region of EBV were carried out, as were the analysis of virus subtypes, A and B, BamHI-F and f, and C and D. Additionally, HPV infection in the squamous cell carcinomas were demonstrated using E6 and E7 region primer sets by PCR method. In Okinawa, 94% (177/188) of the cases were squamous cell carcinomas. A surprisingly large number of EBV (72%) and HPV (78%) infections in the oral squamous cell carcinomas were demonstrated. EBV type B virus infection was found in 36% of EBV-related oral squamous cell carcinoma in Okinawa, but in only 2-5% of the mainland cases. In both regions the incidence of the BamHI- f variant infection was very low. The infected virus in 79 out of 80 (39 Okinawan and 41 mainland) cases was BamHI- F type. In Okinawa, the numbers of C and D variants were almost equal, whereas in the mainland the D variant was rare. Further, a 30 bp deletion in LMP-1 gene was frequently demonstrated in Okinawan and mainland cases of type A virus, but not in type B virus. Lastly, single nucleotide mutations in EBNA2 region of type A virus when compared with B95-8 strain were demonstrated in Okinawan cases. The prognosis for (mostly EBV/HPV infected) squamous cell carcinomas in Okinawa was better than that in the mainland where most cases were negative for EBV and/or HPV.

Clinical course of diffuse invasive carcinoma of the tongue excised after bleomycin treatment

According to the mode of invasion across the tumour-host boundary which was described and graded by Jacobsson et al. (1973), the clinical course of diffuse invasive squamous cell carcinoma of the tongue (grade 4) was investigated comparing it with the other grade groups (grades 1-3). Metastasis to regional lymph nodes and local recurrence after surgery were extremely common in grade 4. These local recurrences led to death in most grade 4 cases while a better clinical outcome was achieved in the other grades. Grade 4 was further subclassified into a cord-like type (grade 4 C) and a diffuse type (grade 4 D) from the histological features of the tumour-host interface, with the latter being more malignant than the former.

A modified version of galectin-9 induces cell cycle arrest and apoptosis of Burkitt and Hodgkin lymphoma cells

The identification of galectin‐9 as a ligand for T‐cell immunoglobulin‐ and mucin‐domain‐containing molecule‐3 (Tim‐3), expressed on T‐helper type‐1 (Th1) cells, has established the Tim‐3‐galectin‐9 pathway as a regulator of Th1 immunity. Whereas there is compelling evidence for the effects of galectin‐9 on T‐cell fate, limited information is available on the impact of galectin‐9 on B lymphocytes. We found that protease‐resistant galectin‐9, hG9NC(null), but not galectin‐1 or ‐8, prevented cell growth of malignant B cells, such as Burkitt lymphoma (BL) and Hodgkin lymphoma (HL). β‐galactoside binding was essential for galectin‐9‐induced cell growth suppression. hG9NC(null) induced cell cycle arrest by reducing the expression of cyclin D1, D2, B1, Cdk4, Cdc25C and c‐Myc, and apoptosis by reducing the expression of XIAP, c‐IAP2 and survivin. Most of the genes that encode these proteins are regulated by nuclear factor‐κB (NF‐κB), and constitutive activation of NF‐κΒ is a common characteristic of both types of malignancies. hG9NC(null) inhibited IκBα phosphorylation, resulting in suppression of NF‐κB. AP‐1 has also been implicated in the control of cell survival. hG9NC(null) inhibited the expression of JunD, resulting in the suppression of AP‐1. Our results suggest that hG9NC(null) is a potentially suitable agent for the management of BL and HL.