Tumor microenvironment in triple-negative breast cancer: the correlation of tumor-associated macrophages and tumor-infiltrating lymphocytes.

PURPOSE: Immune cells such as cytotoxic T cells, helper T cells, B cells or tumor-associated macrophages (TAMs) contribute to the anti-tumor response or pro-tumorigenic effect in triple negative breast cancer (TNBC). The interrelation of TAMs, T and B tumor-infiltrating lymphocytes (TILs) in TNBC has not been fully elucidated. METHODS: We evaluated the association of tumor-associated macrophages, T and B TILs in TNBC. RESULTS: TNBCs with a high CD68+, CD163+ TAMs and low CD4+, CD8+, CD20+ TILs had a significantly shorter relapse-free survival (RFS) and overall survival (OS) than those with low CD68+, CD163+ TAMs and high CD4+, CD8+, CD20+ TILs. TNBCs with high CD68+ TAMs/low CD8+ TILs showed a significantly shorter RFS and OS and a significantly poorer prognosis than those with high CD68+ TAMs/high CD8+ TILs, low CD68+ TAMs/high CD8+ TILs, and low CD68+/low CD8+. TNBCs with high CD163+ TAMs/low CD8+, low CD20 + TILs showed a significantly shorter RFS and OS and a significantly poorer prognosis than those with high CD163+ TAMs/high CD8+ TILs and high CD163+ TAMs /high CD20+ TILs. CONCLUSIONS: Our study suggests that TAMs further create an optimal tumor microenvironment (TME) for growth and invasion of cancer cells when evasion of immunoreactions due to T and B TILs occurs. In TNBCs, all these events combine to affect prognosis. The process of TME is highly complex in TNBCs and for an improved understanding, larger validation studies are necessary to confirm these findings.

Improvement in bronchiolitis obliterans organizing pneumonia in a child after allogeneic bone marrow transplantation by a combination of oral prednisolone and low dose erythromycin.

We report a 13-year-old boy who developed dyspnea at rest 1 year after the occurrence of cGVHD following an allogeneic bone marrow transplant (BMT). Pulmonary function data, imaging studies, lung biopsy, and bronchoalveolar lavage were consistent with the diagnosis of bronchiolitis obliterans organizing pneumonia (BOOP). Although reports suggest that oral methylprednisolone or methylprednisolone pulse therapies improve BOOP after BMT, we treated our patient with a combination of oral prednisolone (1 mg/kg) and low dose erythromycin (10 mg/kg) to avoid the side-effects of high-dose steroids. With this therapy, our patient showed clinical and radiological improvements within 1 week. The steroids were tapered off 12 months later and erythromycin was given for 14 months. We conclude that therapy consisting of a combination of oral prednisolone and low-dose erythromycin for BOOP after BMT may minimize the dose and duration of steroid use.

Progress reports on immune gene therapy for stage IV renal cell cancer using lethally irradiated granulocyte-macrophage colony-stimulating factor-transduced autologous renal cancer cells.

There is no effective treatment for patients with stage IV renal cell cancer (RCC), although the introduction of new therapy is imminent. Cancer gene therapy is currently considered to be one of the most promising therapeutic modalities in the field of cancer treatment. Based on the results of animal studies, vaccination using autologous granulocyte-macrophage colony-stimulating factor-transduced renal cancer cells appears promising. Before initiating a clinical study using an ex vivo gene-transduced autologous cell vaccine-based immunogene therapy for RCC in Japan, in 1992 we initially planned a Japanese version of a clinical protocol in collaboration with a US group. In 1993, the original protocol was refined. We performed five preclinical qualification studies using RCC nephrectomy specimens from patients in 1997, and the results showed that preparation of RCC cells for autologous vaccines at the Clinical Cell Technology Facility, Research Hospital of the Institute of Medical Science, University of Tokyo, was feasible. Subsequently in August 1998, the Ministry of Health and Welfare and the Ministry of Education, Science, Culture, and Sport approved our clinical protocol. We have recruited two patients with stage IV RCC to our study so far. Here we report the background to the initiation of cancer gene therapy in Japan.

Massive pericardial and pleural effusion with anasarca following allogeneic bone marrow transplantation.

A 10-year-old girl presented with massive pericardial/pleural effusion with anasarca 216 days after an allogeneic bone marrow transplantation from her HLA-matched sibling for relapsed acute lymphoblastic leukemia. She did not show any other symptoms of chronic graft-versus-host disease (GVHD). The antinucleolar antibody was elevated in the blood and the pleural fluid. The lymphocytes in the fluid were mostly CD8+/HLA-DR+, and a majority of CD8+ cells in the blood expressed CD57. These data suggested that she had chronic GVHD. Immunosuppressive therapy including prednisolone, cyclosporin A, high-dose methylprednisolone, tacrolimus (FK506), and methotrexate had no effect, and the patient died of Aspergillus pneumonia 183 days after the presentation of the disease. Although it has not been described before, isolated serositis with edema should be recognized as a clinical feature of chronic GVHD.

TCR-Vbeta repertoire analysis with RT-PCR was useful for the early detection of pulmonary relapsed T-cell lymphoma after autologous peripheral blood stem cell transplantation.

Pulmonary recurrence of malignant lymphoma is a rare event after stem cell transplantation. We report here a 45-year-old male who was successfully diagnosed with relapsed pulmonary T-cell lymphoma using an RT-PCR method. Clonal expansion of T cells expressing identical TCR V-D-J junction size (Vbeta5-Jbeta1.5) was demonstrated in lymphocyte groups obtained from both bronchoalveolar lavage fluid at relapse, and paraffin embedded lymph node samples resected when he was first diagnosed with angioimmunoblastic T-cell lymphoma. This method provided evidence to diagnose relapsed pulmonary angioimmunoblastic T-cell lymphoma in its early phase.

Secondary infections of AIDS autopsy cases in Japan with special emphasis on Mycobacterium avium-intracellulare complex infection.

In order to study the frequency of secondary infections of AIDS autopsy cases in Japan, especially the frequency of Mycobacterium aviumintracellulare complex (MAC) infection, retrospective autopsy study was conducted between 1986 and 1997 at the affiliated hospital of Institute of Medical Sciences, University of Tokyo. Secondary infections of various organs from 43 AIDS autopsy cases were examined using histopathology, genetic diagnosis of tuberculosis, Ziehl-Neelsen stain for acid-fast bacilli and immunohistochemistry. Nontuberculous mycobacterial infection (Mycobacterium avium) was observed in 17 cases (40%) out of 43 using polymerase chain reaction (PCR), but M. tuberculosis infection was not observed. Ziehl-Neelsen staining showed a positive reaction in lung and spleen tissues of 7 AIDS autopsy cases. Immunohistochemistry using anti-BCG antibody revealed positivity in 7 AIDS autopsy cases. CD4 counts of 17 AIDS patients with mycobacterial infection were less than 18.7/microl. Other opportunistic infections were also examined by histopathology. Secondary infections were present in every case, and these included cytomegalovirus infection (32 cases), Pneumocystis carinii (15 cases), Candida (16 cases), Aspergillus (12 cases), Cryptococcus (6 cases), Toxoplasma (6 cases), methicillin-resistant Staphylococcus aureus (3 cases), herpes virus (1 case) and Entamoeba histolytica (1 case). Malignant lymphoma was recognized in 14 cases and Kaposi's sarcoma in 6. This is the systemic report on secondary infections of AIDS autopsy cases in Japan. In diagnosis of mycobacterial infections, PCR was more useful than staining for acid-fast bacilli and immunohistochemistry. Secondary infections (especially mycobacterial infection) were closely associated with the low CD4 count.

Role of the cytomegalovirus (CMV)-antigenemia assay as a predictive and follow-up detection tool for CMV disease in AIDS patients.

Forty-two patients were evaluated to determine the value of the CMV antigenemia (CMV-Ag) test as a follow-up marker as well as a prediction marker of CMV disease. Twenty patients were positive for at least one positive CMV-Ag assay and 9 of them developed CMV retinitis. With the threshold value (10 positive cells), sensitivity was 56% and specificity was 94%. The CMV-Ag assay, with the threshold value, produced high specificity, positive predictive value and negative predictive value but relatively poor sensitivity. Eight patients experienced CMV disease relapse a total of 16 times. At relapse, 8 of the 16 times showed negative for CMV-Ag assay; 7 underwent systemic maintenance while 1 underwent local maintenance. It is inferred that the CMV-Ag test is a poor follow-up marker to detect the relapse of CMV disease, particularly in patients undergoing systemic maintenance.

Human herpesvirus type 8 and epstein-barr virus-associated cutaneous lymphoma taking anaplastic large cell morphology in a man with HIV infection.

Human herpesvirus type 8 (HHV-8, Kaposi's sarcoma-associated herpesvirus)-positive lymphoma taking anaplastic large cell morphology in the skin is described in a 46-year-old man with AIDS. Multiple erythematous nodules appeared on the trunk and extremities during the treatment of AIDS. Histological examination of cutaneous nodules showed dense infiltration of CD30 + atypical lymphoid cells in the deep dermis. Immunoglobulin JH gene rearrangement was detected in these lymphoma cells. Both Epstein-Barr virus-encoded small RNA and HHV-8 mRNA (T1.1/nut-1) were detected in these lymphoma cells by in situ hybridization. Remarkable retention of the pericardial fluid was observed at the same time that cutaneous lesions grew, and lymphoma cells in the pericardial fluid showed the same phenotype as the cutaneous lymphoma. Chemotherapy with cyclophosphamide, doxorubicin, vincristine and prednisone effectively reduced both the cutaneous nodules and pericardial fluid. However, the patient died 4 months after diagnosis because of cytomegalovirus infection. As far as we know, this is the first report of an HHV-8-positive cutaneous lymphoma taking anaplastic large cell morphology. This case suggests the association of AIDS-related anaplastic large cell lymphoma with HHV-8.

Diagnosis of measles viral pneumonia in a patient with Hodgkin's disease by reverse transcription-polymerase chain reaction of serum.

We report a case of fatal measles viral pneumonia in a patient with Hodgkin's disease who had no rash. The measles viral cDNA was detected in autopsy tissue from the lung by reverse transcription-polymerase chain reaction. This method was then applied successfully to stored serum. The diagnosis of measles viral pneumonia may be improved by the application of RT-PCR using peripheral blood. Sequence analysis of amplified cDNA suggested the virus infecting this patient was a recent strain, predominantly isolated after 1980. The fatal outcome may have been due to a lack of immune response to the newer strain.

Primary uterine T-cell lymphoma.

A uterine CD8-positive, HTLV-1-negative T-cell lymphoma associated with atypical endometrial hyperplasia was found in a 63-year-old Japanese woman. Primary T-cell lymphoma of the uterus has not been previously reported.

Follicular dendritic cell tumor with histiocytic characteristics and fibroblastic antigen.

A report is presented of a follicular dendritic cell (FDC) tumor arising in the lymph nodes and inguen of a 55-year-old Japanese female, who had suffered from schizophrenia for 25 years. The left submandibular lymph nodes had completely lost their normal architecture, except for the capsule, due to tumor cell infiltration. Occasional nodular structures resembling epithelioid granulomas, attributable, at least in part, to follicular involvement of tumor cells, were observed. These nodules were composed of epithelioid- or fibroblast-like tumor cells forming interwoven fascicles, to which small lymphocytes were attached. Tumor cells were also scattered in the internodular areas. For more atypical tumor cells, arranged in a sheet-like structure, were present in the inguinal specimen, the tumor cells of which expressed Ki-M4p, CD21, CD35 and other antigens known to be expressed on FDC. Furthermore, they also expressed the monocyte/macrophage antigens, alpha 1-antitrypsin, alpha 1-antichymotrypsin, lysozyme, CD14, CD33, CD68 and Mac387 and fibroblastic antigen. Ultrastructural studies demonstrated lysosomal granules as well as a few desmosomes, indicating the tumor cells possessed fibrohistiocytic and FDC characteristics.

Cytokeratin expression in normal human thymus at different ages.

Subsets of thymic epithelial cells were examined immunohistochemically to determine whether or not their phenotypes change during thymic growth and at early involution in terms of cytokeratin (CK) expression. Five monoclonal antibodies specific for CK4, CK8, CK13, CK18 and CK19 were used and applied to 16 neonatal, three infantile and one adult thymus specimen, which had been obtained at autopsy, that were normal macroscopically and microscopically. CK4, CK8, CK13, CK18 and CK19 were expressed simultaneously in the cortex, medulla and subcapsular area with the exception of CK4, which showed expression on the adult thymus. Light and electron microscopy showed that CK8 and CK19 expression was overlapped. Thus, it was thought that CK8 and CK19 formed complexes in the cytoplasm of thymic epithelial cells. The immunoreactivity to CK4, CK13 and CK18 were attenuated or disappeared in the subcapsular area during the early involution stage. Interestingly, two patterns of CK18 expression were observed in the neonatal and infantile thymus tissues, which indicated that the thymic microenvironment was changeable even under normal conditions.

Preferential expression of the multidrug-resistance-associated protein (MRP) in adenocarcinoma of the lung.

The expression of multidrug-resistance-associated protein (MRP) was assessed in various types of untreated lung cancer using an immunohistochemical technique. MRP was abundantly expressed in 28 of 59 adenocarcinoma specimens (47%) and its expression was associated with the degree of glandular differentiation of the tumor. MRP expression in well-differentiated adenocarcinomas (56%) was higher than in poorly differentiated adenocarcinomas (22%) (p < 0.01). lower--20% in squamous-cell carcinomas, 20% in large-cell carcinomas and 0% in small-cell carcinomas and carcinoids. RT-PCR showed that the MRP-positive adenocarcinomas and squamous-cell carcinomas expressed mrp mRNA significantly. Immunoelectron microscopically, MRP was localized in the plasma membrane and rough endoplasmic reticulum. It is thus important to take MRP into account when considering chemotherapy for lung cancers because levels of mdr I gene product, another multidrug-resistance gene family, are low in untreated lung cancers.

Expression of multidrug resistance-associated protein (MRP) in thyroid cancers.

It was found that the mechanism of anti-cancer drug resistance in anaplastic carcinoma of the thyroid was not explicable only in terms of expression of mdr1 and its gene product, P-glycoprotein. The multidrug resistance-associated protein (MRP), another member of the mdr gene family, may be involved in anti-cancer drug resistance of this carcinoma. The MRP expression was examined immunohistochemically in 8 cell lines and 73 thyroid cancer tissues; its frequency in anaplastic carcinoma (52%) was significantly higher than that in other thyroid cancer types.

Preferential reduction of nm23-H1 gene product in metastatic tissues from papillary and follicular carcinomas of the thyroid.

The nm23 gene product is a possible mediator of cancer invasion and metastasis, and has been divided into two distinct gene products, NM23-H1 and NM23-H2. It has been shown previously that expression of nm23-H1 messenger RNA is reduced in metastatic lymph nodes from patients with papillary carcinoma of the thyroid (Am J Pathol 142:1938-1944, 1993). Since mAb H1-229 specific for NM23-H1 is now available, we further extended our study to examine the expression of nm23-H1 gene product in various types of thyroid cancer (37 cases of papillary carcinoma, 42 cases of follicular carcinoma, 17 cases of anaplastic carcinoma, and 3 cases of medullary carcinoma). NM23-H1 was expressed in primary papillary and follicular carcinomas (14/15 and 21/22), but weakly or hardly expressed in metastatic lymph nodes and metastatic bone marrow (6/23 and 1/4) (P < 0.01). Expression of NM23-H1 was low or absent in anaplastic and medullary carcinoma (3/17 and 0/3) (P < 0.01).

A case of encapsulated noninvasive thymoma (stage I) with myasthenia gravis showing metastasis after a 2-year dormancy.

We herein describe a case of thymoma in which metastasis to the left lower visceral and parietal pleura was noticed after a 2-year dormancy. A closer examination revealed no evidence that the metastatic tumors were of lymph node origin. The initial thymoma was well encapsulated, and thus it was thought that the development of metastasis might have reflected a sudden rapid growth of the thymoma cells after a 2-year period of inactivity.

Expression of multidrug resistance-associated protein (MRP) in anaplastic carcinoma of the thyroid.

Although the incidence is very low, the prognosis of anaplastic carcinoma of the thyroid is very poor regardless of the results of various therapeutic trials. We found that the mechanism of anti-cancer drug resistance in anaplastic carcinoma of the thyroid was not explicable only in terms of expression of the mdr1 and its gene product, P-glycoprotein. Therefore, expression of multidrug resistance-associated protein (MRP) mRNA was examined in 11 anaplastic thyroid carcinomas and eight anaplastic thyroid carcinoma cell lines. High MRP mRNA expression was recognized in 7/11 and 8/8, respectively. Our results may contribute to elucidation of the mechanism of anti-cancer drug resistance in this neoplasm.

A case of recurrent intramural uterine stromal tumor with epithelial differentiation effectively treated with oral low-dose administration of etoposide.

A 66-year-old woman was diagnosed as having myoma of the uterus and total hysterectomy with bilateral salpingo-oophorectomy was performed. The histopathological findings were puzzling, and the case was finally diagnosed as intramural uterine stromal tumor with epithelial differentiation. Three years after the operation, a firm tumor developed in the pelvic cavity. Laparotomy failed to remove the tumor and neither CAP (cyclophosphamide 500 mg, adriamycin 50 mg, CDDP 70 mg) therapy nor radiation therapy was effective. Oral administration of etoposide (25 mg/day), however, showed PR (50% decrease) as determined by CT scanning, and ultrasonography, and no metastatic lesions were found. This tumor was coincident with the endometrial stromal tumor with epithelial elements classified by Clement and Scully. The histological feature of the tumors and the efficacy of oral etoposide therapy are discussed in this study.