Noriaki Tezuka

A comparison of epidermal growth factor receptor levels and other prognostic parameters in non-small cell lung cancer

Epidermal growth factor receptor (EGFR) was measured using a competitive radioligand binding assay in membrane preparations from 74 primary human non-small cell lung cancer (NSCLC) tissues and 20 pathologically normal peripheral lung tissues. The mean EGFR level in tumours was 30.38 fmol/mg (+/-41.95 S.D.) of membrane protein (mg.p), significantly higher (P = 0.00016) than in normal tissues (mean, 10.26 +/- 10.02 fmol/mg.p). The mean EGFR concentration was also significantly higher in pathological stage IV tissue than in stages I (P = 0.049) and II (P = 0.040), and the mean EGFR concentration was significantly higher in cases with mediastinal involvement than in cases without it (P = 0.029). The mean EGFR level was higher in DNA aneuploid and multiploid cases than in DNA diploid cases, but there was no significant difference. No significant relationships were found to exist between receptor concentrations and pathological tumour size or histological type, or patient gender or age. From the above findings, a possible prognostic role for EGFR in primary NSCLC should be investigated.

Induction of Glandular Stomach Cancers in C3H Mice Treated with N‐Methyl‐N‐nitrosourea in the Drinking Water

Establishment of an animal model of stomach carcinogenesis in mice was attempted using N‐methyl‐N‐nitrosourea (MNU) in the drinking water. One hundred and forty‐eight male 6‐week‐old C3H mice were given MNU in their drinking water at a concentration of 120 ppm (group 1), 60 ppm (group 2), 30 ppm (group 3) or 0 ppm (group 4) for 30 weeks. At the end of this time, dose‐related induction of adenomatous hyperplasias was found. From weeks 31 to 54 adenocarcinomas developed in a dose‐dependent manner in groups 1, 2 and 3. In total, 6 well differentiated and 5 poorly differentiated adenocarcinomas as well as 6 signet ring cell carcinomas arose in 15 stomach cancer‐bearing animals in group 1, 4 well differentiated and 2 poorly differentiated adenocarcinomas with one signet ring cell carcinoma in 5 mice of group 2 and one well differentiated adenocarcinoma in group 3. In the forestomach, only one squamous cell carcinoma was found at week 54 in group 1 along with a single well differentiated adenocarcinoma in the duodenum. Thus, MNU in the drinking water selectively induced neoplastic lesions in the glandular stomach epithelium of mice.

Expression and Immunogenicity of a Tumor- Associated Antigen, 90K/Mac-2 Binding Protein, in Lung Carcinoma The Possibility of Its Clinical Use as a Tumor Marker and a Target Antigen in Cancer Immunotherapy

The authors attempted to obtain shared proteins among lung carcinoma cells by column chromatographies. A glycoprotein with approximately 500 kDa isolated from QG56 cells showed an identical amino acid sequence to 90K/Mac‐2 binding protein (M2BP). This protein has been reported to be highly expressed and to modulate the expression of surface molecules involved in immune responses on cultured cancer cells. Therefore, it would be beneficial for M2BP to be targeted in cancer immunotherapy.

Novel vaccination protocol consisting of injecting MUC1 DNA and nonprimed dendritic cells at the same region greatly enhanced MUC1-specific antitumor immunity in a murine model

In order to induce specific antitumor immunity in mice, we attempted to immunize C57BL/6 mice with DNA vaccine encoding MUC1 polypeptide. When the mice immunized with MUC1 DNA were challenged with EL4-muc, MUC1-transfected syngeneic lymphoma cells, they completely rejected tumors. When DNA vaccine was given to the EL4-muc tumor-bearing mice, this vaccination was insufficient to suppress tumor growth in the mice. However, activated, but nonprimed dendritic cells (DCs) obtained from syngeneic mice and MUC1 DNA vaccine were given simultaneously to the same site of EL4-muc tumor-bearing mice, tumor growth was markedly suppressed accompanying prolongation of survival time. MUC1 antigen was detected on the DCs at the vaccination site and in regional nodes in the mice which received MUC1 DNA vaccine and DCs. These mice showed markedly enhanced cellular immune responses specific for MUC1 compared to those in mice vaccinated with MUC1 DNA alone. No significant difference in titers of antibodies to MUC1 between the two groups was observed. These results suggest that nonprimed DCs inoculated at the DNA vaccine site are essential for eliciting strong antitumor cellular immunity to suppress tumor growth efficiently in DNA-vaccinated mice. This animal model is useful for developing DNA vaccine for anti-cancer immunotherapy.

Inhibition of Transforming Growth Factor-β–Mediated Immunosuppression in Tumor-Draining Lymph Nodes Augments Antitumor Responses by Various Immunologic Cell Types

Tumor-draining lymph nodes (DLN) are the most important priming sites for generation of antitumor immune responses. They are also the location where an immunosuppressive cytokine, transforming growth factor-beta (TGF-beta), plays a critical role in suppressing these antitumor immune responses. We focused on TGF-beta-mediated immunosuppression in DLNs and examined whether local inhibition of TGF-beta augmented antitumor immune responses systemically in tumor-bearing mice models. For inhibition of TGF-beta-mediated immunosuppression in DLNs, C57BL/6 mice subcutaneously bearing E.G7 tumors were administered plasmid DNA encoding the extracellular domain of TGF-beta type II receptor fused to the human IgG heavy chain (TGFR DNA) i.m. near the established tumor. In DLNs, inhibition of TGF-beta suppressed the proliferation of regulatory T cells and increased the number of tumor antigen-specific CD4(+) or CD8(+) cells producing IFN-gamma. Enhancement of antitumor immune responses in DLNs were associated with augmented tumor antigen-specific cytotoxic and natural killer activity in spleen as well as elevated levels of tumor-specific antibody in sera. The growth of the established metastatic as well as primary tumors was effectively suppressed via augmented antitumor immune responses. Inhibition of TGF-beta-mediated immunosuppression in DLNs is significantly associated with augmented antitumor responses by various immunocompetent cell types. This animal model provides a novel rationale for molecular cancer therapeutics targeting TGF-beta.

Role of transforming growth factor-β1 and decorin in development of central fibrosis in pulmonary adenocarcinoma

Transforming growth factor-beta1 (TGF-beta1) is known as the growth factor that stimulates the synthesis of extracellular matrix. Recently, TGF-beta has been found to control the growth of cancer cells. Small chondroitin-dermatan sulfate (decorin) is an abundant extracellular matrix component. TGF-beta1 stimulates the synthesis of decorin, and decorin is considered to bind TGF-beta1. The activity of decorin in neutralizing TGF-beta1 activity suggests that decorin serves as a negative-feedback regulator of TGF-beta1 activity. To investigate the role and relationship of TGF-beta1 and decorin in the formation of central fibrosis in pulmonary adenocarcinoma, we performed an immunohistochemical study of TGF-beta1 and decorin in 61 cases of T1 pulmonary adenocarcinoma. Positive stainings for TGF-beta1 were shown in 40 cases and negative in 21 cases. Twenty-seven of 32 cases with central fibrosis were positive for TGF-beta1. Positive staining for TGF-beta1 was significantly related to the appearance of central fibrosis in pulmonary adenocarcinoma. When central fibrosis was composed of proliferative connective tissue with loose staining for decorin, cancer cells showed intense staining for TGF-beta1. When central fibrosis was composed of old fibrotic tissue with dense staining for decorin, cancer cells showed weak staining for TGF-beta1. Our results suggest that TGF-beta1 has an important role in the formation of central fibrosis in pulmonary adenocarcinoma, and decorin may play a role as a negative feedback regulator in the production of TGF-beta1 in pulmonary adenocarcinoma.

Angiomatoid fibrous histiocytoma in mediastinum

Angiomatoid fibrous histiocytoma (AFH) is a rare tumor of soft tissue with low-grade malignancy that occurs most commonly in the soft tissues of the extremities or trunk. We present a case of AFH of the mediastinum, which is a very unusual site for this tumor. The patient has survived with no recurrence of the disease for 60 months after surgery and adjuvant radiotherapy.

Encapsulated pericardial fat necrosis treated by video-assisted thoracic surgery: report of a case.

A 55-year-old moderately obese man who was admitted to a local hospital following a traffic accident reported having experienced an episode of sharp and sudden pleuritic pain in the left anterior lower chest 2 days earlier. A computed tomographic scan on admission demonstrated a nonhomogeneous mass in the anterior left side of the chest, abutting the left cardiac margin, and a left-sided pleural effusion. As a mediastinal tumor was suspected, he was referred to our hospital for investigation and treatment. An exploratory thoracotomy was performed by video-assisted thoracic surgery (VATS) about 3 weeks later, which revealed a firm, yellowish mass on the oral side of the pericardial fat pad, adhering to the anterior chest wall. The mass was easily removed. The resected specimen consisted of a lobulated fragment of adipose tissue measuring 5.0 × 3.5 × 2.0cm, and the final pathologic diagnosis was pericardial fat necrosis. The patient had an uneventful postoperative recovery and has remained free of symptoms for 10 months since his operation. Pericardial fat necrosis remains a rare clinical entity. Surgical excision by VATS achieves symptomatic cure and probably continues to be the treatment of choice because of the need to exclude a neoplasm in the differential diagnosis.

Sequential observation of micrometastasis formation by bacterial lacZ gene-tagged Lewis lung carcinoma cells

Sequential events in micrometastasis formation including entry into the blood circulation and arrest, extravasation and initial growth in the lung was investigated using bacterial lacZ gene-tagged Lewis lung carcinoma cells (4A1-1). Micrometastases in the lung could thereby be specifically detected at the single cell level by X-Gal staining. After intravenous injection, X-Gal positive tumor cells appeared to extravasate within hours, but most cells then degenerated or died in the alveolar space by 2-3 days postinjection. A decreased BrdU labeling index to a negligible level at 2 days postinjection and reduction of X-Gal positive foci to a basal level (less than 0.1% of injected cells) by 4 days are in line with rapid clearance of tumor cells from the lung. The size and BrdU labeling indices of the persisting X-Gal positive foci, however, started to increase from 4 days postinjection. Type IV collagen immunostaining demonstrated loss of pre-existing basement membranes with growth of micrometastases: When 4A1-1 cells were inoculated subcutaneously, lung micrometastases from resulting tumors were detected as single or small numbers of X-Gal positive cells at 2 weeks postinjection. Progressive development of micrometastasis to macroscopic metastasis was noted by 4-5 weeks postinjection. The results indicate that micrometastasis formation by Lewis lung carcinoma cells involves a sequence of events starting with rapid extravasation after arrest in the lung within 1 day, followed by death of most cells at 2-3 days and subsequent new growth and expansion of persisting tumor cells from 4 days postinjection.

Cancer‐associated fibroblast‐targeted strategy enhances antitumor immune responses in dendritic cell‐based vaccine

Given the close interaction between tumor cells and stromal cells in the tumor microenvironment (TME), TME‐targeted strategies would be promising for developing integrated cancer immunotherapy. Cancer‐associated fibroblasts (CAFs) are the dominant stromal component, playing critical roles in generation of the pro‐tumorigenic TME. We focused on the immunosuppressive trait of CAFs, and systematically explored the alteration of tumor‐associated immune responses by CAF‐targeted therapy. C57BL/6 mice s.c. bearing syngeneic E.G7 lymphoma, LLC1 Lewis lung cancer, or B16F1 melanoma were treated with an anti‐fibrotic agent, tranilast, to inhibit CAF function. The infiltration of immune suppressor cell types, including regulatory T cells and myeloid‐derived suppressor cells, in the TME was effectively decreased through reduction of stromal cell‐derived factor‐1, prostaglandin E2, and transforming growth factor‐β. In tumor‐draining lymph nodes, these immune suppressor cell types were significantly decreased, leading to activation of tumor‐associated antigen‐specific CD8+ T cells. In addition, CAF‐targeted therapy synergistically enhanced multiple types of systemic antitumor immune responses such as the cytotoxic CD8+ T cell response, natural killer activity, and antitumor humoral immunity in combination with dendritic cell‐based vaccines; however, the suppressive effect on tumor growth was not observed in tumor‐bearing SCID mice. These data indicate that systemic antitumor immune responses by various immunologic cell types are required to bring out the efficacy of CAF‐targeted therapy, and these effects are enhanced when combined with effector‐stimulatory immunotherapy such as dendritic cell‐based vaccines. Our mouse model provides a novel rationale with TME‐targeted strategy for the development of cell‐based cancer immunotherapy.