Yoshito Yamashita

Antitumor Effect of Trastuzumab for Pancreatic Cancer with High HER-2 Expression and Enhancement of Effect by Combined Therapy with Gemcitabine

Purpose: The purpose of the present study was to evaluate whether trastuzumab has antitumor effect against pancreatic cancer and whether this effect is concordant with levels of HER-2, which is reportedly overexpressed in pancreatic cancer. We also investigated whether the effect is potentiated in combined therapy with gemcitabine. Experimental Design: Using immunohistochemistry and FACScan, we analyzed HER-2 expression in 16 pancreatic cancer cell lines. The in vitro antiproliferative effect of trastuzumab, alone and in combination with gemcitabine, was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The in vitro antibody-dependent cell-mediated cytotoxicity of trastuzumab was investigated by 51Cr release assay. The in vivo antitumor effect of trastuzumab, alone and in combination with gemcitabine, was evaluated in nude mouse xenograft growth. The survival benefit was evaluated in a Capan-1 orthotopic implanted nude mouse model. Results: HER-2 expression of 2+ or more was observed in 10 and of 3+ in 2 of the 16 cell lines. No in vitro growth-inhibitory effect of trastuzumab was found in any cell line, but trastuzumab induced antibody-dependent cell-mediated cytotoxicity in proportion to HER-2 expression level. Trastuzumab inhibited tumor growth in Capan-1 (HER-2: 3+) xenografts and prolonged survival in the orthotopic model. These effects were increased by combined therapy with gemcitabine. In contrast, trastuzumab exhibited no antitumor effect against PANC-1 (HER-2: 1+) or SW1990 (HER-2: 2+) xenografts. Conclusions: The antitumor effect of trastuzumab in pancreatic cancer with high HER-2 expression was shown in vitro and in vivo. Clinical application of trastuzumab is expected in pancreatic cancer with 3+ HER-2 expression.

Predictive value of Bcl-2 and Bax protein expression for chemotherapeutic effect in gastric cancer : A pilot study

We investigated tissue staining for Bcl-2 and Bax proteins, which regulate apoptosis, as indicators of chemotherapeutic effect in patients with gastric cancer. In 23 patients with gastric carcinoma biopsy specimens were obtained endoscopically prior to chemotherapy and stained immunohistochemically with anti-Bcl-2 and anti-Bax antibodies. Patients then were treated with continuous infusion of 5-FU and cisplatin. No correlation was seen between chemotherapeutic effect and Bcl-2 or Bax alone. However, among the Bax-positive cases, the patients with Bcl-2-positive tumors were significantly more chemoresistant (p = 0.036) and had worse prognoses (p = 0.008) than Bcl-2-negative cases. Therefore, immunohistochemical staining for Bcl-2 protein may predict chemotherapeutic efficacy or guide specific therapeutic choices in treating Bax-positive tumors.

p53 protein overexpression as a predictor of the response to chemotherapy in gastric cancer.

This study was performed to evaluate p53 overexpression as a predictor of the response to chemotherapy of patients with gastric cancer. The subjects comprised 20 patients with Stage IV gastric cancer and three with locally recurrent lesions, all of whom were treated with 5-fluorouracil (5-FU) plus cisplatin (CDDP) for 4 weeks. Of the total 23 patients there were 10 responders; 2 showing complete response (CR) and 8, partial response (PR). Specimens obtained by endoscopic biopsy were immunohistochemically stained using anti-p53 protein and bcl-2 protein antibody. Of the 10 responders, 7 demonstrated negative p53 staining, and of the 13 nonresponders, 11 demonstrated positive p53 staining (P=0.013). Tissue from 3 of the responders and 7 of the nonresponders that stained for bcl-2 were positive prior to chemotherapy; however, there was no association between bcl-2 staining and chemotherapeutic effect. In conclusion, immunohistochemical identification of p53 in pretreatment tissue may represent a useful predictor for chemotherapeutic outcome in patients with gastric cancer.

Mucoepidermoid Carcinoma of the Pancreas : Report of a Case

We present herein the case of a 64-year-old man diagnosed as having a mucoepidermoid carcinoma of the pancreas. The tumor originated in the tail of the pancreas and invaded the spleen, left adrenal gland, left kidney, and transverse colon. Liver and peritoneal metastases were also noted. Despite surgical treatment and adjuvant chemotherapy, the disease progressed rapidly and the patient died of cachexia 4 months after his initial diagnosis. Mucoepidermoid carcinoma of the pancreas is a rare entity, and is believed to be a form of adenosquamous carcinoma known as adenoacanthoma. However, in this patient, no differentiated squamous cell component could be detected. In fact, the tumor was composed of mucin-producing cells, epidermoid cells, and intermediate cells. Immunohistochemical staining for the carcinoembryonic antigen, CA19-9, and SPan-1 demonstrated a production of cancerous mucin in the epidermoid cells, suggesting that mucoepidermoid carcinoma may arise from the squamoid metaplasia of an adenocarcinoma.

A novel orthotopic implantation model of human esophageal carcinoma in nude rats: CD44H mediates cancer cell invasion in vitro and in vivo

A new orthotopic esophageal cancer model was developed by implanting fragments of xenografts of T.T human esophageal squamous carcinoma cells into the cervical esophagus of athymic rats. The rats had symptoms analogous to the human clinical course such as respiratory distress, dysphagia, vomiting of blood, or Horner syndrome, followed by death resulting from suffocation. Microscopic metastases of lymph node were observed around the tumor in 3 of 18 rats. A new cell line (T.T‐1) was established from these metastases. Flow cytometry showed that T.T‐1 and T.T parental cells had nearly the same surface levels of β1‐integrin, α2‐integrin, α3‐integrin and E‐cadherin, and no expression of CD44v3, CD44v6 and α5‐integrin. T.T‐1 cells had a higher level of CD44H, however, and a greater binding efficiency to the extracellular matrix components; laminin, type IV collagen, hyaluronic acid, and fibronectin than T.T cells. Anti‐CD44H antibody significantly decreased the binding efficiency of T.T‐1 cells. T.T‐1 cells were also significantly more invasive than T.T cells through all the extracellular matrix components except hyaluronic acid. After orthotopic implantation histological examination showed that T.T‐1 tumors invaded beyond the esophageal mucosa and tracheal muscle layer and obstructed the esophagus and trachea. No invasion was observed with T.T tumors. Rats with T.T‐1 or T.T tumors survived an average of 32.0 and 50.7 days, respectively (p < 0.01). In addition T.T‐1 tumors expressed higher levels of CD44H mRNA than T.T tumors. In summary, our newly developed orthotopic implantation model is a valid model of esophageal cancer because it followed the same clinical course experienced by humans. Moreover, using cells derived from this model, we were able to demonstrate that CD44H is involved in esophageal cancer cell invasion.

Prognostic significance of T antigen expression in patients with gastric carcinoma

Thomsen‐Freidenreich (T) antigen, the immediate precursor antigen of the human blood MN system, has been detected in malignant cells, but not in most normal cells in which it is cryptic but can be unmasked by desialylation. In this study, we determined the prognostic significance of T antigen in specimens with gastric carcinoma.

Intranodal Lymphangiogenesis Precedes Development of Lymph Node Metastasis and Accelerates Progression of Gastric Cancer

IntroductionPeritumoral lymphangiogenesis is significantly correlated with lymph node metastasis and poor prognosis in various cancers. However, there are few reports concerning the role of intranodal lymphangiogenesis in lymphatic metastasis. The aim of this study was to examine the association of lymphangiogenesis in regional lymph nodes with the progression of gastric cancer.Materials and MethodsLymphatic vessel density (LVD) was immunohistochemically estimated in 1,596 lymph nodes from 52 patients who underwent gastrectomy.ResultsIntranodal LVD was significantly correlated with the size of metastasis and the progression of cancer. Patients in the high LVD group had significantly poorer prognosis relative to patients in the low LVD group. Furthermore, expression of VEGF-C mRNA was significantly up-regulated in lymph nodes of pathological node positive patients compared to node negative patients.DiscussionThus, intranodal lymphangiogenesis was correlated with nodal metastasis and poor prognosis of patients with gastric cancer. Lymphangiogenesis in regional lymph nodes plays an early role in spreading of tumor cells through the lymphatic system to distant organs in gastric cancer.

Antibody‐dependent Cytotoxicity Mediated by Chimeric Monoclonal Antibody Nd2 and Experimental Immunotherapy for Pancreatic Cancer

In a previous study, mouse monoclonal antibody (MoAb) Nd2 (m‐Nd2, mouse IgGl) labeled with 131 I exhibited efficacy in in vivo radioimmunotherapy against pancreatic cancer. In this study we prepared mouse/human chimeric antibody Nd2 (c‐Nd2, human IgG1) for clinical use and examined whether c‐Nd2 induced antibody‐dependent cell‐mediated cytotoxicity (ADCC). Cytotoxicity to pancreatic cancer (PC) cell lines, including Nd2 antigen‐positive (SW1990, RWP‐1, Capan‐1) and Nd2 antigen‐negative (Panc‐1, MiaPaca‐2, Capan‐2) lines, was evaluated by mixed human leukocyte and tumor cell culture (MLTC) at an effector cell to target cell (E/T) ratio of 50 with or without Nd2. Cytotoxicities to SW1990 with no antibody, m‐Nd2 and c‐Nd2 (1 μg/ml) were 26.7%, 38.0% and 55%, respectively; to RWP‐1, 28%, 41% and 70%; to Capan‐1, 26%, 30% and 52%; to Panc‐1, 24%, 28% and 30%; to MiaPaca‐2, 18%, 20% and 27% and to Capan‐2, 29.7%, 35.0% and 40.6%. Cytotoxic capacity during MLTC with c‐Nd2 was significantly higher than during MLTC with m‐Nd2 or with no antibody. These findings indicated that cytotoxicity to Nd2‐positive PC cells during MLTC is induced by ADCC. Intraperitoneal injection of c‐Nd2 inhibited the tumor growth of SW1990 xenografted subcutaneously in nude mice and prolonged the survival of nude mice in which SW1990 tumor was transplanted orthotopically at the tail of the pancreas. These findings suggested that, because of its ability to induce ADCC, c‐Nd2 may be clinically useful for the immunotherapeutic treatment of pancreatic cancer.

Preoperative clinical radioimmunodetection of pancreatic cancer by 111In-labeled chimeric monoclonal antibody Nd2

The present study was carried out with the purpose of evaluating the clinical usefulness of radioimmunodetection (RAID) with 111In‐labeled murine/human chimeric monoclonal antibody, Nd2 (c‐Nd2) in patients with pancreatic cancer. Nineteen patients suspected to have pancreatic cancer were administered intravenously 74 MBq/2 mg 111In‐labeled c‐Nd2 in 100 ml of saline containing 2% albumin over 30 min. A scintigram was obtained on the 3rd day after infusion by using single photon emission computed tomography (SPECT) imaging. Of the 14 patients finally diagnosed as having pancreatic cancer on the basis of surgical specimens or progress of disease, specific focal uptake at the site of the tumor was detected in 12 (true positive cases), representing a sensitivity of 85.7% (12/14), and liver metastasis was found in one case with metastasis. Of the 5 patients diagnosed with tumor‐forming pancreatitis (TFP), 4 patients demonstrated true negative imaging, but one patient whose tumor demonstrated interesting findings in histology and immunostaining, showed false positive imaging. Of patients investigated for human anti‐chimeric antibody (HACA) response, none showed HACA response, and no allergic reaction was seen in any of the patients administered c‐Nd2. These results suggest that RAID with 111In‐labeled c‐Nd2 is useful for differential preoperative diagnosis between invasive pancreatic cancer and TFP.

Intratumoral Injection of an Adriamycin Immunoconjugate against Human Pancreatic Cancer Xenografts

We have evaluated the effect of an adriamycin conjugate of monoclonal antibody Nd2 (ADM‐Nd2) on the growth rate of SW1990 xenografts grown subcutaneously in athymic nude mice. Intravenous or intraperitoneal administration of radiolabeled Nd2 resulted in a maximum tumor accumulation of approximately 45% of the initial dose/g of tumor 3‐7 days after administration. However, administration into the tumor produced retention of 1200%ID/g 1 day after, with 50% of this high value remaining even at 7 days after administration. In contrast, intratumoral administration of a nonspecific immunoglobulin showed a lower initial retention and rapid loss of label. Both intravenously and intratumorally administered ADM‐Nd2 reduced the growth rate of SW1990 xenografts. While a single intravenous administration arrested growth for about two weeks, a single intratumoral injection prevented any increase in tumor size even 45 days after administration. Xenografts treated with ADM‐Nd2 showed degenerative changes at the histological level. Neither Nd2 alone nor Adriamycin alone inhibited growth when administered at the same dose as the conjugate.