Tetsuhiko Itoh

Human papillomavirus-16 is integrated in lung carcinomas: a study in Chile

The human papillomavirus (HPV) was detected in 20 (29%) out of 69 lung carcinomas (LCs) in Chile, by PCR and Southern blot, and was more frequently detected in squamous cell carcinoma (SQC) than in adenocarcinomas (46 vs 9%, P=0.001). HPV-16, positive in 11 cases, was the most frequently detected HPV genotype determined by DNA sequencing. HPV-16 E2/E6 ratio, estimated from real-time PCR analysis, was much lower than the unity, suggesting that at least a partial HPV-16 genome was integrated in all but one HPV-16-positive SQCs. The remaining one case was suspected to have only episomal HPV-16. Although the viral load was low in most of the LCs, a case showed the HPV-16 copy number as high as 8479 per nanogram DNA, which was even a few times higher than the minimum viral load of seven cervical carcinomas (observed viral load: 3356–609 392 per nanogram DNA). The expression of the HPV-16/18 E6 protein was found in only two HPV-16-positive SQCs (13%) but not in the case with the highest viral load. Although the viral load was in general very low and HPV E6 expression is none or weak, further studies seem warranted to examine aetiological involvement of high-risk HPV in lung carcinogenesis.

Epstein-Barr virus-associated gastric carcinoma in Japanese Brazilians and non-japanese Brazilians in São Paulo

The proportion of Epstein‐Barr virus‐associated gastric carcinoma (EB V‐GC) was examined in 149 Japanese‐Brazilian and 151 non‐Japanese‐Brazilian gastric‐carcinoma cases using in situ hybridization (ISH) assay to detect EBV‐encoded small RNA (EBER), and the results were compared with our referent Japanese data. We found that 4.7% of cases in Japanese Brazilians were EBER‐positive. This frequency was slightly lower than that of the referent Japanese, among whom 6.2% of 2038 gastric‐carcinoma cases were EBER‐positive. On the other hand, the non‐Japanese‐Brazilian series showed a significantly higher proportion of EBV‐GC (11.2%) than the referent group did (P=0.01). Although EBV‐GC was predominant in males among non‐Japanese Brazilians (M/ F=3.6, P=0.047), as was the case in Japanese (M/F=2.7), Japanese Brazilians did not show such a male predominance. The sex‐ratio difference between the Japanese Brazilians and Japanese was statistically significant (P=0.005). In conclusion, the present study in Japanese Brazilians and Japanese yielded no evidence suggesting any change in the frequency of EBV‐GC caused by migration, except the absence of male predominance, which was observed both in Japanese and non‐Japanese Brazilians.

Expression of mucin antigens and Lewis X-related antigens in carcinomas and dysplasia of the pharynx and larynx

Recent studies have Identified that much antigens and Lewis X (Lex)‐related antigens behave like oncodevelopmental tumor‐asnoclated antigens in several human adenocarcino‐mas. However, the expression of these antigens In pharyn‐geal and latyngeal squamous cell carcinomas (SCC) and in the precursor lesion is not fully elucidated yet. In the present study, the expression of mucin core protein antigens associated with the MUC1 gene product (DF3 antigen, mammary‐type apomucin) and the MUC2 gene product (intestinal‐MRP antigen, Intestinal‐type apomucin) much carbohydrate antigens that are associated with the earliest steps in mucin glycosylation (Tn, sialyl‐Tn and T), and Lex ‐related antigens (Lex, Leγ and slayl Lex‐1) In biopsy or resected specimens from 26 normal squamous epithelia (NSE), 49 dysplastic squamous epithelia (DSE) and 51 SCC were examined. The DF3 antigen was not expressed In NSE (0%), whereas it was expressed in 20 DSE (41%) and in 31 SCC (61%). The intestinal‐MRP antigen showed no expression in NSE, DSE or SCC. The Tn antigen showed no expression in NSE, but showed low expression rates In DSE (14%) and in SCC (16%). The sialyl‐Tn and T antigens were expressed in NSE, as well as in DSE and SCC. The T antigen expression increased with progression from NSE to DSE to SCC, while the sialyl‐Tn antigen did not show such a tendency. Any of the three Lex‐related antigens showed no characteristic expression in DSE and SCC. In the eight antigens examined, only DF3 antigen was an effective marker for DSE and SCC in the pharyngeal and laryngeal region. Cytoplasmic expression of DF3 and slalyl‐ Tn antigens were more frequently seen In SCC than in DSE, and might be useful to differentiate SCC from DSE.

Malignant eccrine spiradenoma with smooth muscle cell differentiation: histological and immunohistochemical study.

Malignant transformation in a benign eccrlne eplradenoma Is rare. Since It was flrst reported In 1972, 31 cases have been reported. Of these, six cases had sarcomatous components, mainly osteosarcomatous, and one had rhabdomy‐obiastic differentiation. A case Is presented In which there were some small nodules of benlgn eccrine spiradenomas adjacent to a large nodule of malignant eccrine splradenoma with lelomyosarcomatous components, determined by his‐tologlc and lmmunohlstochemlcal studies.

Amplification and Overexpression of c-met Gene in Epstein-Barr Virus-Associated Gastric Carcinomas

To reveal the role of oncogenes in Epstein-Barr virus (EBV)-positive gastric carcinomas, the amplification and overexpression of the c-met gene were examined by a competitive polymerase chain reaction and immunohistochemistry, respectively. The proportion of c-met amplification and overexpression in EBV-positive and -negative carcinomas did not differ significantly. The amplification and overexpression of the c-met gene in EBV-negative gastric carcinomas were significantly associated with upper location, deeper invasion and lymphatic invasion, while in EBV-positive gastric carcinomas a significant correlation with c-met activation was observed only in deeper invasion. However, none of the observed associations of c-met amplification or overexpression with clinicopathological features in the EBV-positive and -negative carcinomas differed significantly in their strength or direction. These results suggest that the amplification and overexpression of c-met gene do not play a different role in the progression and metastasis of EBV-positive and EBV-negative gastric carcinomas.

Deletions and single-base mutations within the carboxy-terminal region of the latent membrane protein 1 oncogene in Epstein-Barr virus-related gastric cancers of Southern Japan

The 30‐base pair (bp) deletion of the cytoplasmic carboxy‐terminal domain of the latent membrane protein 1 (LMP‐1) gene was analyzed in 37 frozen tissues from patients with Epstein‐Barr virus (EBV)‐related gastric cancer and 18 throat washings from healthy adults in southern Japan. The 30‐bp deletion was identified in 33 (91.7%) of 36 specimens of EBV‐related gastric cancers and in 15 (83.3%) of 18 throat washings from healthy adults. In one case of gastric cancer, an additional 9‐bp deletion was identified downstream of the 30‐bp deletion. From the last transmembane domain to the end of the carboxy terminal of LMP‐1, mutations were examined in 37 cases of gastric cancers and in three cases of throat washings. Twenty‐eight nonsilent mutations were identified in this region of EBV‐related gastric cancer and throat washings. Five nonsilent mutations at positions 168,755, 168,746, 168,687, 168,357, and 168,355 were identified in all 30‐bp‐deleted cases of EBV‐related gastric cancers and throat washings. However, these nonsilent mutations were not identified in three patients without the 30‐bp deletion. Although the deletion and single‐base mutations of the LMP‐1 gene in gastric cancers and throat washings were similar to those of nasopharyngeal carcinoma in Taiwan and China, more single‐base mutations were found in southern Japan. These data indicate that high prevalence of the 30‐bp deletion of the LMP‐1 gene in gastric cancers may reflect the prevalence of the deletion variant in the normal population in southern Japan. J. Med. Virol. 57:152–158, 1999.

Nasal natural killer/T-cell lymphoma and its association with type “i”/XhoI loss strain Epstein–Barr virus in Chile

Background: Nasal T/natural killer (NK)-cell lymphoma is an aggressive type of non-Hodking’s lymphoma associated with Epstein–Barr virus (EBV) and striking geographical variations worldwide. Aim: To characterise nasal NK/T-cell lymphoma associated with genotypes of EBV in Chile, a Latin American country, where multiple strains of EBV, including two new recombinant strains, in healthy individuals were recently found. Methods: Cases with diagnosis of primary nasal lymphoma were selected for histological and immunohistochemical analysis (CD3, CD3e, CD4, CD8, CD79a, CD56, CD57 and TIA-1) and in-situ hybridisation, serology and genotyping analysis for EBV. Results: Out of 22 cases, 9 (41%) cases fulfilled the World Health Organization criteria for nasal NK/T-cell lymphoma; of these 7 (78%) cases were positive for EBV. Genotyping analysis revealed 6 cases of type 1 EBV and wildtype F at the BamHI-F region, 4 cases type “i” EBV at the BamHI-W1/I1 region; XhoI wild type was found in 2 and XhoI loss in 4 cases, respectively. Cosegregation analysis of the BamHI-W1/I1 region and XhoI restriction site showed the new recombinant strain type “i”/XhoI loss in 3 cases and type “i”/XhoI wild-type strain in 1 case. Most patients were treated with combined anthracycline-containing regimens. Half of the cases attained complete remission. Conclusion: Although nasal NK/T-cell lymphomas from Chile share similar clinicopathological features, high association with EBV and unfavourable prognosis with those described elsewhere, genotype analysis shows that the new recombinant type “i”/XhoI loss strain might contribute to explain the intermediate incidence of nasal NK/T-cell lymphomas in Latin America.

Loss of p16/CDKN2A protein in Epstein-Barr virus-associated gastric carcinoma.

We examined the expression of p16, the CDKN2A gene product, in EBV-associated gastric carcinomas (EBV-GCs). EBV-GCs were identified by detecting EBV-encoded small RNA (EBER) using an in situ hybridization assay of paraffin-embedded tissue. Two non-EBV-GC cases for each EBV-GC case were selected, matched for age, sex, tumor location, and depth of invasion. After excluding cases without sufficient tissue samples for immunohistochemical analysis, 54 EBV-GC and 117 non-EBV-GC cases were available for the present study. The loss of p16 expression was more frequently observed in EBV-GCs (89%) than non-EBV-GC cases (32%; p < 0.001). Among non-EBV-GC cases, the loss of p16 expression was more frequent in female cases (57%) than male cases (29%) (p = 0.042). Expression of p16 was not related to the location of tumor, clinical stage of tumor, age, or prognosis of the patients. In conclusion, the present study suggests that the loss of p16-related cell cycle regulation may be associated with the development of EBV-GC.

Expression of Epstein-Barr virus in malignant lymphoma.

In this report, we investigated 73 cases of Hodgkins disease (HD) and 752 consecutive cases of non-Hodgkins lymphoma (NHL) to demonstrate EBV in the tumor cells by EBER in situ hybridization. For EBV positive cases, expression of EBNA1, EBNA2, LMP1 and P53 gene products were examined immunohistochemically, EBV-serology and prognostic data were also analyzed. EBV was detected in 48 (65.8%) of Hodgkins disease in the tumor cells. EBV-positive HD showed expression of LMP1 and episomal monoclonality in all 5 cases examined. Among NHL, EBV was detected in 44 (12.0%) of 368 cases with ATL, 36 (31.6%) of 114 cases with T-ML, 18 (6.8%) of 263 cases with B-ML and none of 7 other types (p<0.00001). High prevalent expression of LMP1 (66%) and P53 gene products (80%) and low prevalent expression of EBNA1 and EBNA2 were observed in EBV-positive NHL cases with similar distribution of EBER ISH sporadic and diffuse positive cases. Episomal clonality of EBV were demonstrated in some cases examined. Elevated antibody titer particularly IgA antibodies against VCA and EA, and poor prognosis of EBV positive cases were observed.