A case of gastric cancer following living donor liver transplantation.

Only a few cases of de novo malignancy, especially gastric cancer after living donor liver transplantation (LDLT), have been reported. We report a case of gastric cancer following LDLT, after which immunosuppressants were minimized in accordance with the results of the mixed lymphocyte reaction (MLR) assay. A 65-year-old woman had previously undergone LDLT for hepatocellular carcinoma associated with hepatitis B virus infection. The liver graft had been donated by her son. During the course of postoperative surveillance with the MLR assay in order to minimize immunosuppressants, she was incidentally found to have gastric cancer during an endoscopic examination, 8 years after the liver transplantation. She underwent total gastrectomy with lymph node dissection. In this case, gastric cancer was detected 8 years after LDLT, which is longer than previously reported intervals between LDLT and malignancy detection. The number of patients undergoing LDLT is increasing, and the prognosis after liver transplantation has improved. Therefore, endoscopic surveillance programs are important for detecting malignancies in the early stages in liver transplant recipients.

A novel anti-MUC1 antibody against the MUC1 cytoplasmic tail domain: use in sensitive identification of poorly differentiated cells in adenocarcinoma of the stomach.

BACKGROUND: Isolated cancer cells of non-solid type poorly differentiated adenocarcinoma (por2) or signet-ring cell carcinoma (sig) are frequently seen in scirrhous gastric cancers with a very poor prognosis. These cells are often scattered in granulation tissue or desmoplastic fibrotic tissue and tend to be overlooked in routine pathological examination. We aimed to raise a novel antibody that can identify the isolated cancer cells easily. METHODS: Because the MUC1 cytoplasmic tail domain (CTD) has many biological roles including tumor progression and cell adhesion disturbance and is expected to be expressed in isolated cancer cells, we raised a novel monoclonal antibody (MAb) MUC1-014E against an intracellular nonrepeating 19-amino-acid sequence (RYVPPSSTDRSPYEKVSAG: N-1217-1235-C) of the MUC1 CTD, using a synthetic peptide including the 7-amino-acid epitope (STDRSPY: N-1223-1229-C). RESULTS: In the immunohistochemical staining of 107 gastrectomy specimens including 48 por2 and 31 sig lesions, the MAb MUC1-014E showed high rates of positive staining (≥5% of carcinoma cells stained) for por2 (100%) and sig (97%), and of the highest intensity staining (4+, ≥75% of carcinoma cells stained) for por2 (100%) and sig (90%). In the 89 biopsy specimens including 82 por2 and 38 sig lesions, the MAb MUC1-014E showed high rates of positive staining for por2 (100%) and sig (100%) and of 4+ staining for por2 (87%) and sig (84%). All the rates were significantly higher than those with cytokeratins (AE1/AE3 or CAM5.2). CONCLUSIONS: The MAb MUC1-014E is very useful for accurate detection of isolated cancer cells in scirrhous gastric cancers.

Identification of HLA-A*2402-restricted epitope peptide derived from ERas oncogene expressed in human scirrhous gastric cancer.

ERas is a recently identified oncogene involved in the tumorgenic growth of embryonic stem cells. We examined the significance of ERas expression in scirrhous gastric carcinoma, and the possibility of ERas as a tumor-associated antigen of gastric cancer for developing a cancer vaccine. ERas expression was determined in scirrhous gastric carcinoma specimens by immunohistochemical staining. To assess the possibility of the ERas protein as an anticancer vaccine target, we examined whether ERas for HLA-A-restricted epitope peptides were capable of eliciting cytotoxic T lymphocyte activity. Immunohistochemical analysis identified ERas protein in the nucleus and cytoplasm of cancer cells, yet ERas was not expressed in normal gastric epithelium. By western blotting, lysates of the scirrhous gastric cancer cell lines, OCUM-8, OCUM-2MD3 and OCUM-2M were shown to contain a 25-kDa band of ERas protein. ERas mRNA was detected in these cell lines by RT-PCR. To investigate cytotoxicity, we successfully established cytotoxic T lymphocyte clones stimulated by HLA-A*2402-restricted ERas peptides (FALDDPSSL). These peptides have specific cytotoxicity against corresponding HLA-A*2402-positive target cells pulsed with the candidate peptide. We found that the cytotoxic T lymphocyte clones demonstrated cytotoxic activity against OCUM-8 cells that endogenously express ERas. Our results suggest that ERas is a novel tumor-associated antigen with the potential application to be a vaccine against scirrhous gastric cancer.

Cellular and stromal characteristics in the scirrhous hepatocellular carcinoma: comparison with hepatocellular carcinomas and intrahepatic cholangiocarcinomas.

Scirrhous hepatocellular carcinoma (SHCC) is a rare variation of HCC, for which characteristics of tumor cells and the fibrotic stroma have not been clarified in detail. The present study was therefore carried out to elucidate cytological features of tumor and stromal cells and components of the stromal extracellular matrix in 15 SHCC patients undergoing hepatectomy without preoperative transarterial embolization. Diagnosis was on the basis of a scirrhous histological pattern exceeding 50% of the tumor area. Expression of cytoplasmic and extracellular matrix proteins was compared among SHCC, HCC and intrahepatic cholangiocarcinoma (ICC) cases with immunohistochemical staining. The lesions could be histologically divided into radiating and sinusoidal types. Common stromal components of SHCC and ICC were collagen types I and III. There was no expression of laminin-5 in the stroma of SHCC, but it was present in almost all ICC cases. Tenascin-C expression was significantly lower in the SHCC cases and its distribution differed between SHCC and ICC. Matrix metalloproteinase-7 (MMP-7) expression was significantly higher in SHCC compared with HCC. Almost all stromal cells were alpha-smooth muscle actin-positive both in SHCC and ICC, whereas glial fibrillary acid protein (GFAP)-positive stromal cells were significantly more increased in ICC than in SHCC. SHCC clearly differed from HCC with respect to collagen types I, III and MMP-7 expression, and from ICC with regard to stromal components including laminin-5, tenascin-C and GFAP(+) stromal cells.

Prognostic value of tumor-infiltrating dendritic cells expressing CD83 in human breast carcinomas.

DCs are the most potent antigen-presenting cells that play a major role in initiating the antitumor immune response. Although the clinical significance of TIDCs has been investigated in a variety of human cancers, few studies have focused on the in situ maturation status of DCs. We have analyzed the maturation-specific significance of TIDCs in the prognosis of patients with breast carcinoma. We evaluated 130 breast carcinomas for the presence of TIDCs using immunohistochemistry with an anti-CD1a antibody for immature DCs and an anti-CD83 antibody for mature DCs. Intratumoral expression of immunosuppressive cytokines was also examined. All samples contained CD1a(+) TIDCs, and 82 (63.1%) samples contained CD83(+) TIDCs. The number of CD83(+) TIDCs was inversely correlated with lymph node metastasis and with tissue expression of VEGF and TGF-beta, whereas the number of CD1a(+) TIDCs was not. Kaplan-Meier analysis (log rank statistics) revealed a significant association of increasing number of CD83(+) TIDCs with longer relapse-free (p = 0.002) and overall (p < 0.001) survival. Furthermore, among patients with lymph node metastasis, the survival rate of those with larger numbers of CD83(+) TIDCs was significantly better than that of patients with fewer CD83(+) TIDCs. Multivariate analysis revealed that CD83(+) TIDCs had independent prognostic relevance in breast carcinomas. The infiltration of tumors by mature DCs expressing CD83 may be of great importance in initiating the primary antitumor immune response and is confirmed as an independent, immunologic prognostic parameter for survival in patients with breast cancer.

Infiltrating dendritic/Langerhans cells in primary breast cancer.

It is fully anticipated that dendritic cells (DCs) will become a mainstay for inclusion in biological therapies for patients with cancer including breast cancer. To elucidate the cellular composition of DCs infiltrating human breast cancers, we investigated the correlations between the density of infiltrating DCs and some clinicopathological factors of breast cancer patients, examined cytokine expression on cancer cells and finally, assessed the numbers of CD45RO+ tumor infiltrating lymphocytes (TIL). Tissues adjacent to cancer nests contained significantly more S-100 protein+ and S-100 protein+ CD1a- DCs, but less CD1a+ DCs, than the nests. In invasive ductal carcinomas infiltration by S-100 protein+ DCs within and adjacent to nests, CD1a+ DCs within nests and S-100 protein+ CD1a- DCs adjacent to nests was denser than that in non-invasive carcinomas. With respect to the histological subtypes, there were fewer DCs in scirrhous carcinomas. Patients with stage IV disease had significantly fewer DCs of primary lesions than at other clinical stages. There were good correlations between infiltration by S-100 protein+ DCs and expression of the cytokines GM-CSF, IL-1alpha and TNF-alpha on cancer cells and between GM-CSF expression and S-100 protein+ CD1a- DCs. There was a close correlation between CD45RO+ TIL and S-100 protein+ DC densities both within and adjacent to the cancer nests and the S-100 protein+ CD1a- DC density adjacent to the cancer nests. Despite extensive immunoelectron microscopic observation, CD1a+ DCs within cancer nests contained only few Birbeck's granule-like structure. These data indicate that cancer nests are infiltrated predominantly by CD1a+ DCs, whereas S-100 protein+ CD1a- DCs predominate in surrounding tissues, and a infiltration by DCs may require cytokine expression on cancer cells and simultaneous lymphocyte infiltration. The findings of this clinicopathological study indicate the importance of evaluating simultaneously the types and localizations of infiltrating DCs in cancer tissues.

Establishment of lymph node metastatic model for human gastric cancer in nude mice and analysis of factors associated with metastasis.

The actual mechanisms responsible for lymph node metastasis in gastric cancer are still unclear. To investigate the mechanisms of lymph node metastasis in gastric cancer, we established a lymph node metastatic model for human scirrhous gastric carcinoma. Lymph node metastasis had frequently developed after orthotopic implantation of OCUM-2M LN derived from a scirrhous gastric cancer cell line, OCUM-2M, which had low capacity for lymph node metastasis. We elucidated the different characteristics including binding ability, migratory capacity and immunoresponses induced by the cell surface molecules of these two cell lines. The binding ability to Matrigel and migratory capacity of OCUM-2M LN cells were significantly greater than those of OCUM-2M cells. On flow cytometric analysis, both OCUM-2M and OCUM-2M LN cells strongly expressed HLA-I (99.5 and 97.1%) and LFA-3 (76.6 and 99.2%) in level of expression between the two cell lines, but neither cell line expressed HLA-II (0.0 and 0.0%), B7-1 (0.0 and 0.0%) or B7-2 (0.4 and 0.3%). ICAM-1 expression in OCUM-2M LN cells was weaker (0.7%) than that in OCUM-2M cells (36.8%). Strong adhesiveness and cytotoxicity of mononuclear lymphocytes for OCUM-2M cells were observed in adhesion and cytotoxic assays, both of which were significantly decreased by the addition of anti-ICAM-1 antibodies. On the other hand, the adhesiveness and cytotoxicity of OCUM-2M LN cells were significantly less than those of OCUM-2M cells, and were not affected by the addition of anti-ICAM-1 antibodies. These findings suggest that decreased ICAM-1 expression in a new gastric cancer cell line with a high rate of lymph node metastasis may in turn decrease immune responses mediated through LFA-1-dependent effector cell adhesion, and that this escape from the immunosurveillance system may be one of the factors inducing lymph node metastasis. In conclusion, we established a gastric cancer cell line, OCUM-2M LN, with a high rate of lymph node metastasis. An in vivo lymph node-metastatic model with this cell line should be useful for analysing the mechanism and therapeutic approach of lymph node metastasis.

A case report of immunotherapy on a patient with advanced gastric cancer by adoptive transfer of OK-432-reactive HLA-matched allogeneic lymphocytes.

Adoptive immunotherapy (AIT) for non-hematological malignancies, using HLA-matched donor lymphocytes, has been rarely reported. For a 35-year-old male patient with peritoneal disseminated advanced gastric cancer, we performed AIT using lymphocytes from his HLA-matched 37-year-old brother and a streptococcal preparation, OK-432, as an antigen. After the donor had been immunized by intradermal administration of OK-432, OK-432-reactive lymphocytes were induced in vitro and transferred to the patient intravenously with OK-432. Low-dose systemic immunochemotherapy, using interleukin-2, 5-fluorouracil and cyclophosphamide, was concurrently administered with AIT. As a result, the Schnitzler metastasis in the patient reduced in size without any significant graft-versus-host-related complications. One of the effector mechanisms of therapeutic benefit was suggested to be cytokine release from the transferred OK-432-reactive lymphocytes. Our findings suggest the safety and efficacy of AIT using lymphocytes from an HLA-matched sibling and OK-432 as an antigen. Further studies to investigate the use of tumor-associated antigen and an HLA-matched sibling's lymphocytes for AIT of advanced cancer are warranted.

Establishment of two new scirrhous gastric cancer cell lines: analysis of factors associated with disseminated metastasis.

Determination of the differences between cell lines which are derived from a primary tumour and a disseminated metastatic lesion from the same patient may aid in elucidating the factors associated with disseminated metastases. We report on the establishment and characterisation of two new scirrhous gastric cancer cell lines, designated OCUM-2M and OCUM-2D, derived from a 49-year-old female. OCUM-2M was derived from a primary gastric tumour, and OCUM-2D was derived from a sample of disseminated metastasis. The two cell lines were derived from the same patient. We investigated biological differences between the two cell lines to study mechanisms involved in disseminated metastasis. The growth activity of OCUM-2D cells as determined by doubling time and tumorigenicity was greater than that of OCUM-2M cells. The level of epidermal growth factor receptor (EGFR) expression in OCUM-2D cells was about twice that of OCUM-2M cells and the growth of OCUM-2D cells was stimulated more by epidermal growth factor (EGF) than that of OCUM-2M cells. The invasive activity of OCUM-2D cells was higher than that of OCUM-2M cells and was increased after addition of transforming growth factor-beta 1 (TGF-beta 1). An increase in the number of attached and spreading cells was found following the addition of 10 ng ml-1 TGF-beta 1. These findings suggest that high growth and invasive activity may play an important role in disseminated metastasis and that EGF and TGF-beta 1, which affect the growth and invasive activity of OCUM-2D cells, might be factors associated with metastasis in scirrhous gastric carcinoma. The two cell lines OCUM-2M and OCUM-2D should be beneficial for analysing mechanisms of tumour progression.

Mucin antigens expression and Ki-67 labeling in breast cancer: the peculiarity in scirrhous carcinoma.

The expression of mucin-related antigens (Tn, T, Sialosyl-Tn [STn], DF3 [mammary-type apomucin related antigen], and intestinal-MRP [intestinal-type apomucin related antigen]) as well as Ki-67 labeling was examined in 58 mammary invasive ductal carcinomas (IDC) divided into 26 scirrhous subtype (SC) and 32 non-scirrhous subtype comprising papillotubular carcinoma and solid-tubular carcinoma (PT-ST). These data were analyzed in connection with the various pathological prognostic factors such as nodal status, tumor size, estrogen receptor status and histological grading of carcinomas. The results were as follows: (a) in SC, the expression rate of Tn was significantly higher in the cases with positive lymph node metastasis or with large tumor size (> 2 cm) than in those with negative lymph node metastasis or with small tumor size (< or = 2 cm); (b) in PT-ST, the expression rate of STn was higher in the cases with positive lymph node metastasis or with large tumor size than in those with negative lymph node metastasis or with small tumor size; (c) in SC, Ki-67 labeling was significantly higher in the cases with positive lymph node metastasis than in those with negative lymph node metastasis; and (d) in PT-ST, Ki-67 labeling was lower in the cases with positive lymph node metastasis than in those with negative lymph node metastasis. In conclusion, Tn antigen expression was correlated with pathological prognostic factors in SC but not in PT-ST, whereas STn antigen expression was correlated with pathological prognostic factors in PT-ST but not in SC.(ABSTRACT TRUNCATED AT 250 WORDS)

Induction of cytolytic activity of lymphocytes from carcinomatous pleural effusion by IL-2 and autologous tumor cells.

We established a cell line (STKM-1) from tumor cells obtained from carcinomatous pleural effusion of a gastric cancer patient. The lymphocytes separated from her peripheral blood or pleural effusion were cryopreserved and immunological experiments were performed after the establishment of the cell line. They were treated with IL-2 or with both IL-2 and mitomycin C (MMC)-treated autologous STKM-1 cells. The cytolytic activity against STKM-1 cells was elevated in lymphocytes cultured with IL-2, and was more prominently augmented in lymphocytes cultured with both IL-2 and MMC-treated STKM-1 cells. The elevation in cytolytic activity was more marked with pleural effusion lymphocytes than with the peripheral blood lymphocytes. The results suggest that the lymphocytes obtained from the pleural effusion would be an excellent source for adoptive immunotherapy.

Stability and localization of biotin-labeled murine monoclonal antibody to human gastric cancer in normal mice and nude mice-human tumor models.

We examined the tissue localization of biotin-labeled murine monoclonal antibody (MAb) S202 directed against the human scirrhous gastric carcinoma cell line MK-01 in normal and tumor-bearing mice after intravenous (IV) administration. The biotin-labeled MAb proved to be stable in vivo under normal conditions, antibody titer being 1:256 at 4 hr after IV injection. At 24 hr after injection, the tumor was stained by the avidin-biotin-peroxidase complex (ABC) method. Biotin-labeled MAb was found to be suitable for detection of the xenografted tumor of nude mice. This study provides new information concerning the dynamics of the distribution of biotin-labeled MAb in vivo.

A tumor-associated antigen in the scirrhous gastric carcinoma cell line MK-01 defined by monoclonal antibody S202.

A monoclonal antibody (MAb) S202, with an IgG1 isotype, that reacted strongly with the scirrhous gastric carcinoma cell line MK-01 was established. MAb S202 reacted with the colonic cancer cell line, SW1116, and the pancreatic cancer cell line, PK-1, when tested by indirect immunofluorescence. The S202 reactive antigen was expressed in the majority of acetone-fixed fresh frozen cancer tissues. Eighty to 100 per cent of the paraffin-embedded sections of stomach, colon and pancreatic adenocarcinoma were positive for the S202 antigen, with diffuse cytoplasmic staining, whereas esophageal and breast cancers demonstrated markedly less immunostaining. Supplemented serum-free medium collected from 7 day old tumor cell cultures were assayed for the presence of antibody-defined antigens. Antigens detected by MAb S202 were released by the cell lines SW1116 and PK-1. The binding of MAb S202 to the colonic adenocarcinoma sections was reduced after treatment with sodium periodate which suggests that respective antigenic determinants are of carbohydrate nature without sialic acid residues.

[Immunomodulation by leukocytolysis. On the significance of lectin-polysaccharide complexes]. / Immunmodulation durch Leukozytolyse. Zur Bedeutung von Lektin-Polysaccharid-Komplexen.

Polymorphonuclear granulocytes (PMNs) are thought to function only as phagocytes. The ability of PMNs for a physiologically lysis has not been noticed so far. Therefore, PMNs are not only a clamp between the specific and unspecific immune system but an important regulator of the homoeostasis. All immune modifiers therapeutically used seem to stimulate the physiologically lysis of PMNs. This has been shown with mistletoe lectins. Most remarkable are findings that show lectins bound to polysaccharides drastically rise the ability of PMNs to undergo lysis.

[Breast cancer associated with pregnancy].

Two cases of breast cancer associated with pregnancy were reported and the immunological assessment was performed in the second case. Case 1: A 36 year-old woman in the first trimester of pregnancy and having a large axillary mass and breast lump was admitted. She was diagnosed as having Stage IIIb breast cancer. After interruption of pregnancy, she received an extended radical mastectomy and radiotherapy. However, she died nine months after surgery for brain and liver metastases. Case 2: A 30 year-old woman in her second trimester of pregnancy was admitted for a large breast mass. She was diagnosed as having Stage IIIa breast cancer. Interruption of pregnancy was again necessary to perform surgery followed by chemotherapy. After an extended radical mastectomy she was placed on a combination chemotherapy regimen with cyclophosphamide, adriamycin and 5-FU. In both cases tumors began to develop rapidly at pregnancy. Immunological studies in Case 2 showed a depression of T-lymphocyte function and NK cell activity. Our study suggests that the depression of cell-mediated immunity during pregnancy may promote tumor growth.

Localization of antigen defined by anti-scirrhous gastric carcinoma monoclonal antibody S202 in fixed human cancer tissues.

Monoclonal antibody S202 reactive with scirrhous gastric carcinoma cell line MK-01 was used in an immunoperoxidase assay to characterize the antigenic profile of a variety of fixed samples. Representative sections of malignant tissue from the stomach, colon, esophagus, pancreas, bile duct, gallbladder and breast were examined by the avidin-biotin complex immunoperoxidase technique and compared. The antibody reacted with 90 out of 98 stomach adenocarcinomas, and with 30 out of 32 colon adenocarcinomas with diffuse cytoplasmic staining, whereas it reacted with only 19 out of 37 breast carcinomas, and 5 out of 10 esophageal squamous cell carcinomas. The antigenic determinants appeared to be a carbohydrate, since it was highly sensitive to sodium periodate.

[The immunohistochemical localization of CA 15-3 in human breasts and their tumors].

The tissue distribution of CA 15-3, a breast carcinoma associated antigenic determinant, has been investigated by an avidin-biotin immunoperoxidase technique in the human breasts and in their tumors. In cases of scirrhous carcinoma, positive stainings for CA 15-3 were observed in almost all carcinoma cells. In cases of solid-tubular carcinomas and papillotubular carcinomas, the reaction was various and there was no correlation between the distribution of CA 15-3 and the clinical stage. Apical staining was recognized in both solid-tubular and papillotubular carcinomas, in benign breast tumors, and in normal breast tissues. CA 15-3 appears to be related to the degree of differentiation of the breast tissue.

Localization of radiolabeled monoclonal antibody S74 in human scirrhous type gastric cancer xenografts in nude mice.

A mouse monoclonal antibody (MoAb), S74 against a human scirrhous gastric carcinoma cell line was developed. The radioiodinated antibody bound preferentially in vivo to xenografted tumors rather than to normal tissue, whereas tumor uptake of normal mouse IgG did not occur. Thus this MoAb was able to selectively localize a human tumor in an animal model, and clinically it may enable immunotherapy against scirrhous carcinoma of the stomach.

Monoclonal antibodies against human scirrhous carcinoma of the stomach.

We established three monoclonal antibodies (MoAbs) S43, S74 and S202 which reacted strongly with scirrhous carcinomas of the stomach. MoAb S43 reacted with culture supernatants of the scirrhous gastric cancer cell line MK-01, and thus may be useful for in vivo detection of markers for this tumor. MoAb S74 reacted with the majority of gastric cancer tissues, but showed no reaction with normal tissues. It may be useful as a drug carrier in targeting chemotherapy against scirrhous gastric cancer, because of its tumor specificity. MoAb S202 reacted strongly with 100% of scirrhous gastric cancer samples and with the culture supernatants of colonic and pancreatic cancers.

Class distribution of immunoglobulin-containing plasma cells in the stroma of medullary carcinoma of breast.

A class distribution of plasma cells associated with the stroma in twenty-eight cases of medullary carcinoma of the breast was investigated by an unlabeled immunoperoxidase method. The stroma of the medullary carcinomas tested was found to contain predominantly IgG plasma cells except in two cases. Stroma of the other types of breast carcinoma, including ten cases of papillo-tubular carcinoma, five cases of scirrhous carcinoma, and six cases of medullary tubular carcinoma, contained predominantly IgG plasma cells, although few plasma cells were associated with carcinoma tissues in the latter group. Plasma cells associated with control specimens, including normal breast, fibroadenoma, cystic disease, and intraductal papilloma, were found to be predominantly of IgA type. Few carcinomatous epithelial cells contained secretory components in the cytoplasm, while a number of cells positive for secretory components were observed in acinar and ductular epithelia of normal breast tissues and in benign proliferative lesions of the breast. It is suggested that the lymphoid cells infiltrating the stroma of medullary carcinoma represent a sign of host immune response against the carcinoma cells which is related to the well-known favorable prognosis associated with this tumor.