Yasuaki Harabuchi

Epstein-Barr virus in nasal T-cell lymphomas in patients with lethal midline granuloma

Five cases of lethal midline granuloma were identified histologically and phenotypically as peripheral T-cell lymphomas. Epstein-Barr virus (EBV) DNA was detected in the nasal tumour biopsy specimens by Southern blotting and in-vitro hybridisation with simultaneous detection of EBV-determined nuclear antigen (EBNA) and T-cell surface markers by two-colour immunofluorescence. Further immunofluorescence and northern blotting revealed that EBNA2 gene and also latent membrane protein gene were expressed in the nasal tumour cells. The patients had high titres of antibodies to EBV. These findings suggest that lethal midline granuloma is causally associated with EBV.

Expressions of Matrix Metalloproteinases in Early-Stage Oral Squamous Cell Carcinoma as Predictive Indicators for Tumor Metastases and Prognosis

Purpose: Matrix metalloproteinase (MMP)-2 and MMP-9 are considered to play an important role in the metastasis of malignant tumors. Membrane type 1-MMP (MT1-MMP) and tissue inhibitor of metalloproteinase 2 (TIMP-2) are essential factors for the activation of pro-MMP-2. There are some reports about expressions of MMP family in relationship to clinical features of head and neck squamous cell carcinoma (SCC), but the results were not uniform and the prognostic value of their expressions remains unclear. Experimental Design: The study group consisted of 53 Japanese patients with oral SCC of early stage (T1-2N0M0). Expressions of MMP-2, MMP-9, MT1-MMP, and TIMP-2 were examined using immunohistological methods on the sections of tumor biopsy samples. The intensity of MMP expression was categorized into four grades (score 0–3) by semiquantitative analysis using a computer with NIH image, and correlation between this grade and clinical aspects such as tumor recurrence, metastasis, and prognosis were examined. Results: The expression score of MMP-2 correlated with that of MMP-9 (r = 0.291; P = 0.036), MT1-MMP (r = 0.286; P = 0.039), and TIMP-2 (r = 0.257; P = 0.050). Patients who developed regional lymph node and/or distant metastasis showed significantly higher scores in the expressions of MMP-9 and TIMP-2 than patients without any tumor metastases (P = 0.036 and P = 0.043, respectively). Kaplan-Meier analyses as well as univariate analyses using the Cox proportional hazards model showed that expression of MMP-9 (P = 0.0143 and P = 0.0418, respectively) and marked expression of TIMP-2 (P < 0.0001 and P = 0.0004, respectively) correlated with worse-cause-specific survival. Multivariate analysis confirmed that marked expression of TIMP-2 was the only independent factor for cause-specific death (hazard ratio, 7.543; confidence interval, 1.693–33.610; P = 0.0080). Conclusions: Expressions of MMP-9 and TIMP-2 have predictive value for tumor metastases and cause-specific survival. High expression of TIMP-2 is the most independent factor for worse prognosis in early-stage oral SCC.

Nasal T-cell lymphoma causally associated with Epstein-Barr virus : Clinicopathologic, phenotypic, and genotypic studies

The authors have previously demonstrated nasal T‐cell lymphoma (NTL) associated with Epstein–Barr virus (EBV). The detailed clinical, phenotypic, and genotypic features and the role of EBV in lymphomagenesis remain to be clarified.

Prospective measurement of Epstein-Barr virus-DNA in plasma and peripheral blood mononuclear cells of extranodal NK/T-cell lymphoma, nasal type

Epstein-Barr virus (EBV)-DNA was prospectively analyzed in plasma and mononuclear cells (MNCs) from peripheral blood in patients with extranodal natural killer (NK)/T-cell lymphoma, nasal type, to evaluate the clinical significance for diagnosis, monitoring the tumor burden, and prognostication. Thirty-three patients were enrolled, and 32 were evaluable. Pretreatment plasma and MNC EBV-DNA was detectable in 14 (range, 50-71 000 copies/mL) and 6 patients (range, 20-780 copies/μg DNA), respectively, and both were well correlated (r = 0.8741, P < .0001). Detectable plasma EBV-DNA was associated with higher clinical stage (P = .02), presence of B symptoms (P = .02), worse performance status (P = .02), and higher serum soluble IL-2 receptor level (P < .0001). Twenty-two patients attained complete response. Plasma EBV-DNA level was significantly higher in nonresponders than in responders (mean, 16,472 vs 2,645 copies/mL; P = .02). Multivariate analysis showed clinical stage (hazard ratio, 9.0; 95% confidence interval, 1.8%-45.0%) and pretreatment plasma EBV-DNA (hazard ratio, 10.6; 95% confidence interval, 1.3%-87.0%) were significant prognostic factors. Three-year overall survival of plasma EBV-DNA positive and negative patients was 42.9% and 94.4%, respectively (P = .0009). Plasma was a preferable sample for this purpose in NK/T-cell lymphoma, nasal type, and EBV-DNA level was a good indicator for response and overall survival.

Non-Hodgkin's lymphoma of Waldeyer's ring and nasal cavity. Clinical and immunologic aspects

Twenty‐nine cases of non‐Hodgkins lymphoma of Waldeyers ring (W‐NHL) and nasal cavity or paranasal sinus (N‐NHL) were studied for tumor‐surface marker phenotype and histopathologic correlation with clinical features. Immunostaining procedures on tissue sections by using xenoantisera and monoclonal antibodies to human B‐ and T‐cells enabled the authors to demonstrate precise surface marker phenotypes of tumor cells and, moreover, the histologic localization of normal or neoplastic B‐ and T‐cells in preserving the original structure of lymphoid organs or tumor tissues. In 22 cases of W‐NHL, 19 (86%) had B‐cell markers and 3 (14%) had T‐cell markers, whereas 6 of 7 cases (86%) of N‐NHL had T‐cell markers. Tumor cells in T‐cell iymphomas in W‐NHL and N‐NHL reacted with antibodies to peripheral T‐cells except one case of W‐NHL. Rappaport “histiocytic” subtype was heterogeneous with respect to both surface marker characteristics and morphologic features, i.e., seven had B‐cell markers and four had T‐cell markers, and they were all subdivided into “large cell” or “large cell, immunoblastic” in Working Formulation and “large cell” or “pleomorpphic” in Lymphoma Study Group classification. The actuarial survival curve for all T‐cell lymphoma patients was characterized by a rapid initial decline and a subsequent plateau, which contained two of the long survivors. In contrast, the B‐cell lymphoma group had a more graded decline. The median and actuarial survivals of the T‐cell lymphoma group were far inferior to those for the lymphoma group that expressed B‐cell markers.

Tumour suppressive microRNA-874 regulates novel cancer networks in maxillary sinus squamous cell carcinoma

Background:On the basis of the microRNA (miRNA) expression signature of maxillary sinus squamous cell carcinoma (MSSCC), we found that miR-874 was significantly reduced in cancer cells. We focused on the functional significance of miR-874 in cancer cells and identification of miR-874-regulated novel cancer networks in MSSCC.Methods:We used PCR-based methods to investigate the downregulated miRNAs in clinical specimens of MSSCC. Our signature analyses identified 23 miRNAs that were significantly reduced in cancer cells, such as miR-874, miR-133a, miR-375, miR-204, and miR-1. We focused on miR-874 as the most downregulated novel miRNA in our analysis.Results:We found potential tumour suppressive functions such as inhibition of cancer cell proliferation and invasion. A molecular target search of miR-874 revealed that PPP1CA was directly regulated by miR-874. Overexpression of PPP1CA was observed in MSSCC clinical specimens. Silencing of the PPP1CA gene significantly inhibited cancer cell proliferation and invasion.Conclusion:The downregulation of miR-874 was a frequent event in MSSCC, which suggests that miR-874 functions as a tumour suppressive miRNA, directly regulating PPP1CA that has a potential role of an oncogene. The identification of novel miR-874-regulated cancer pathways could provide new insights into potential molecular mechanisms of MSSCC oncogenesis.

Prognostic value of p53 mutations, bax, and spontaneous apoptosis in maxillary sinus squamous cell carcinoma

Many researchers have attempted to correlate p53 mutation and spontaneous apoptosis with the effectiveness of radiochemotherapy and with prognosis in several malignancies.

Expression of CXCR4 and Its Down-Regulation by IFN-γ in Head and Neck Squamous Cell Carcinoma

Purpose: The functional expression of CXCR4, which plays roles in cell migration and proliferation in response to its unique ligand stromal cell–derived factor-1 (SDF-1), has been reported in variety of carcinomas. However, CXCR4 expression and its functional role in head and neck squamous cell carcinomas (HNSCC) remain unclear. In this study, we investigated CXCR4 expression and analyzed its functions in HNSCC cell lines. We also attempted to regulate CXCR4 expression using cytokines, such as interleukin-1β, tumor necrosis factor-α, and IFN-γ. Finally, we investigated correlation between CXCR4 expression and clinical features in patients with HNSCC. Experimental Design: Six HNSCC cell lines were used in this study. Reverse transcription-PCR and flow cytometry analysis were shown for CXCR4 expressions with or without stimulations of cytokines. SDF-1-mediated cell migration was assayed in Matrigel-coated chemotaxis chamber. The SDF-1-mediated cell proliferation was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The SDF-1-mediated signaling pathways were analyzed by Western blot analysis. Biopsy specimens from 56 patients with HNSCC were used for immunohistologic analysis. Results: The significant CXCR4 expression was found in HSQ-89, IMC-3, and Nakamura cells. The SDF-1-mediated cell migration and proliferation were observed in CXCR4-positive cells. SDF-1 also promoted rapid phosphorylation of extracellular signal-regulated kinase 1/2 and Akt signaling pathways in CXCR4-positive cells. The SDF-1-mediated cell migration and proliferation of CXCR4-positive cells were inhibited by neutralization of CXCR4. Among three cytokines tested, IFN-γ significantly reduced CXCR4 expression and SDF-1-induced cell migration and proliferation of CXCR4-positive cells. Immunohistologic analysis revealed that patients with advanced neck status and patients who developed distant metastases showed significantly higher CXCR4 expression, and the cause-specific survival of patients with CXCR4-expression was significantly shorter. Furthermore, multivariate analysis confirmed that CXCR4 positive was the independent factor for cause-specific death. Conclusion: Our results may provide an insight into future therapeutic agent that inhibits tumor metastasis and progression via down-regulating CXCR4 expression in patients with HNSCC.

Human milk secretory IgA antibody to nontypeable Haemophilus influenzae: possible protective effects against nasopharyngeal colonization.

Sixty-eight children fed human milk were followed prospectively from birth to 12 months of age to assess the effect of milk antibody on nasopharyngeal colonization. Human milk secretory IgA antibody to P6, a highly conserved outer membrane protein of nontypeable Haemophilus influenzae, was measured with the use of an enzyme-linked immunosorbent assay. Nasopharyngeal colonization with nontypeable H. influenzae and the occurrence of otitis media were determined. Nasopharyngeal colonization was found in 22 children (32%), and 39 children (57%) had otitis media. Frequency of isolation of nontypeable H. influenzae was directly related to episodes of otitis media (r = 0.35; p = 0.001). The level of human milk anti-P6 secretory IgA antibody was inversely related to frequency of isolation of the organism (r = -0.27; p = 0.026). The average antibody level, expressed as nanograms per 0.1 mg total secretory IgA, in human milk fed to children with no colonization of nontypeable H. influenzae was significantly higher than in milk fed to children in whom colonization occurred on multiple occasions (156 +/- 120 vs 69 +/- 50; p = 0.013). Prevention of colonization was most evident during breast-feeding. These data suggest that the protective effects of human milk against otitis media may be due in part to inhibition of nasopharyngeal colonization with nontypeable H. influenzae by specific secretory IgA antibody.

Frequent mutations of Fas gene in nasal NK/T cell lymphoma

Fas (Apo-1/CD95) is a cell-surface receptor involved in cell death signaling through binding of Fas ligand. Mutation of Fas gene in lymphoid cells results in accumulation of these cells, which might thus contribute to lymphomagenesis. We examined the open reading frame of Fas cDNA in 14 cases of nasal NK/T-cell lymphoma. Mutations of Fas gene were detected in seven (50%) of 14 cases which comprised four frameshift, two missense, and one silent mutations. Frameshift mutations were caused by insertion of 1 bp (A) at nucleotide 1095 in two cases and by deletion of 1 bp at nucleotide 597 and at 704, respectively, in one each. Mouse T-cell lymphoma cells transfected with two missense mutated genes and frameshift mutations caused by insertion of 1 bp (A) at nucleotide 1095 were resistant to apoptosis induced by the anti-Fas antibody. These findings suggested that accumulation of lymphoid cells with Fas mutations provides a basis for the development of nasal NK/T-cell lymphoma.