Peripheral T cell lymphoma.

Thirty-one Chinese patients with peripheral T cell lymphoma (PTCL) were reviewed. Using the modified Japanese Lymphoma Group classification, there were nine (29%) of the pleomorphic type, 16 (52%) immunoblastic lymphadenopathy (IBL)-like, two (7%) T-zone lymphoma, and one (3%) Lennert's lymphoepithelioid type. Three (9%) were not classifiable. All were positive for T11 (E rosette receptor antigen). Fifty-four percent (15 of 28) were positive predominantly for T4 (helper T cell) and 46% (13/28) for T8 (suppressor T cell). The median age of the patients was 57 years. They usually presented with advanced disease, and while extranodal involvement was common, CNS disease was not seen. The IBL-like type was associated with a positive Coombs' test and polyclonal hypergammaglobulinemia. Five of the nine pleomorphic type were checked for antibody to HTLV-I virus and all were negative. PTCL was associated with poor prognosis, which was not influenced by the histologic subtypes and the T4/T8 phenotypes. The complete response rate of 13 consecutive patients who received the BACOP (bleomycin, doxorubicin, cyclophosphamide, vincristine, and prednisone) L17M regimen was significantly better than the 16 historic controls who received other less-intensive regiments, 84% v 19% (P less than .01). The relapse rate was also significantly lower, 9% v 100% (P less than .001). There appeared to be an improvement in the disease-free survival (DFS) (80% v 0% at 18 months), as well as the overall survival (60% v 36% at 18 months), but the differences did not reach statistical significance due to small sample sizes.

Phosphorothioate and normal oligodeoxyribonucleotides with 5'-linked acridine: characterization and preliminary kinetics of cellular uptake.

Certain phosphorothioate oligodeoxynucleotide (S-oligo) analogs, unlike their normal congeners, have been found to exhibit significant anti-HIV activity [Matsukura et al., Proc. Natl. Acad. Sci. USA 84 (1987) 7706-7710]. Here we report melting temperatures (Tm) of a series of S-oligos compared with those of the corresponding normal oligomers. The Tm's for AT base pairs of S-oligos are significantly depressed relative to normal oligos, while GC-containing S-oligos show much less Tm depression. The Tm's of S-dT oligomers with poly(rA) are reduced relative to the duplexes with normal dA oligomers. These results provide a rational basis for the S-d(CG) sequences as anti-message inhibitors of gene expression. We also describe an automated synthesis of 5'-acridine linked oligothymidylates using phosphoramidite-linked acridine. During this synthesis we noted the replacement of thiophenol for the 6-chloro substituent on acridine. We have measured the Tm's of the compounds with 3 and 5 methylene groups linked to normal and phosphorothioate dTn (with n = 3-40) on duplex formation with the equivalent dAn, and have found small increases of Tm for the 5-methylene-linked acridine derivative. We have monitored the uptake of these fluorescently labeled oligos into HL60 cells, and found that the shorter oligos are more rapidly taken up than the longer, and the normal oligos faster than the S-oligos. The temperature dependence of the cellular uptake suggests an energy-dependent process, and a possible membrane receptor for oligos. These results have significance for the potential use of such compounds as inhibitors of gene expression.

Clonal Epstein-Barr virus in lymphoepithelioma-like carcinoma of the lung.

Lymphoepithelioma of the nasopharynx is an undifferentiated carcinoma with a prominent lymphoid infiltrate. A consistent association with Epstein-Barr virus (EBV) has been demonstrated over the years by a variety of methods. More recently, undifferentiated carcinomas with a similar morphology have been described in other anatomical locations, often of foregut origin. However, for lymphoepithelioma-like carcinoma, the association with EBV has been more tenuous, especially in Western countries. Interestingly, these tumors have shown a geographic distribution similar to nasopharyngeal lymphoepithelioma, with relatively high frequency of EBV positive cases in Asian patients. We report five cases of lymphoepithelioma-like carcinoma arising in the lung. In situ hybridization revealed the presence of EBV in all cases with localization to the epithelial cells only. Southern-blot analysis from two cases revealed the presence of clonal episomal EBV in the tumor tissue. These findings further support the hypothesis that EBV is also associated with lymphoepithelioma-like lung carcinomas and suggest that EBV infection has preceded the clonal expansion of the tumor.

Pulmonary scar cancer. A pathologic reappraisal.

A total of 49 consecutive specimens of lung cancer were collected prospectively at surgical resection or autopsy from 40 men and nine women, aged 40-74 years. Of the 49 tumors, the gross appearance of 22 fitted the description of a scar cancer, i.e., a tumor with pleural puckering and central pigmentation. Nineteen of the "scar cancers" were peripheral (17 adenocarcinomas and two squamous cell carcinomas); three were central (one squamous cell carcinoma and two adenocarcinomas). In the 19 peripheral "scar cancers," elastic stains demonstrated the presence of collapsed, unfibrosed lung tissue at the center with traction of the overlying pleura toward it. Elsewhere in the tumor, the elastic framework was either destroyed or expanded by tumor filling the alveolar spaces. None of the "scar cancers" had a significant desmoplastic reaction that might otherwise explain the scarred appearance. It appeared that local atelectasis was solely responsible for the pleural puckering and central pigmentation. On the other hand, atelectatic lung tissue was not seen in the 27 cancers that did not have the appearance of a scar cancer. Tuberculosis was found in 10 of the 49 lung specimens. In only one specimen was the tuberculous lesion anatomically associated with the tumor. There was no evidence of pulmonary infarct in any of the specimens. The term "scar cancer" was considered inappropriate as there was no preformed fibrous tissue. The scarred appearance was thought to be the result of localized pulmonary atelectasis owing to small airways obstruction by tumor. Association with tuberculosis was considered incidental.

Polymorphic reticulosis and conventional lymphomas of the nose and upper aerodigestive tract: a clinicopathologic study of 70 cases, and immunophenotypic studies of 16 cases.

Seventy patients with malignant lymphomas, including the entity known as polymorphic reticulosis (PR), involving the nose, nasal sinuses, nasopharynx, oropharynx (excluding tonsil), and larynx were studied. There were 26 cases of PR, 19 cases of lymphoma with features of PR (ML[PR]) and 25 cases of conventional lymphomas. Fourteen of the 25 conventional lymphomas were due to dissemination from distant sites. For all histologic types of primary lymphoma, the presenting symptoms were similar, and the nasal cavity was more commonly involved than the nasopharynx. Patients with PR were younger, had a higher male:female ratio, and had a better overall survival rate than patients with conventional lymphomas. Cryostat section immunohistochemistry performed on 17 samples from 16 patients showed only one B lymphoma out of 11 primary lesions; the other 10 cases and three recurrent tumors at distant sites showed phenotypic markers of T lymphocytes and natural killer cells. All three secondary tumors were of B-cell type. Of eight patients with sequential biopsies, progression to a more malignant histopathologic type was found in six. In the PR and ML[PR] biopsies, angiocentricity was detected in 11%, and angioinvasion in 22%. We could not confirm identity of PR with other angiocentric immunoproliferative lesions.

Squamous cell carcinoma of the esophagus with mucin-secreting component. Mucoepidermoid carcinoma.

Among 1058 patients with cancer of the esophagus, 20 patients with mucoepidermoid or adenosquamous cell carcinoma of the esophagus and cardia, together defined as squamous cell carcinoma with a mucin-secreting component, were seen over a 10-year period. Their records were reviewed and appropriate comparisons were also made with the more common squamous cell carcinomas and adenocarcinomas. Squamous cell carcinoma with mucin-secreting component comprised 1.9% of all tumors encountered. Clinical features including age, male predominance, symptoms at presentation, length of tumor, and appearance of tumor did not differ from those of squamous cell and adenocarcinoma. The location of these tumors, however, followed that of squamous cell carcinomas, with 55% in the middle third and 25% in the lower third. Adenocarcinomas were found predominantly at the cardia (83%). Operability and resectability rates were higher than those of squamous cell and adenocarcinomas. Primary treatment consisted of resection in 19 of the 20 patients (95%); 18 of them had a one-stage resection and 1 patient had a two-stage resection. Postresection staging showed that 5% had stage I disease, 16% had stage II, and 79% had stage III disease. None of the patients who underwent resection died within 30 days of the operation. The mortality after 30 days was 10.5%. The 1 patient in whom intubation was the primary treatment had distant metastases at the time of presentation (stage IV). The overall median survival was 9.2 months. The median survival for patients who had their tumors resected was 9.5 months. The survival improved to 33 months for curative resection but was only 8.7 months for palliative resection. The 1-, 2-, and 5-year survivals were 46%, 39%, and 0%, respectively. This prognosis was not significantly different from that of patients with squamous cell carcinoma or adenocarcinoma.

Latent sites of Epstein-Barr virus infection.

Epstein-Barr virus (EBV) has been detected in a wide spectrum of tumors. This study investigates the detection rate of EBV-DNA by Southern blot hybridization analysis (SOBH) and polymerase chain reaction (PCR) in different tissues from persons without apparent EBV-related diseases. Of 20 tonsillectomy specimens studied, SOBH indicated positivity for EBV-DNA in 1 case, and PCR indicated positivity in 10. In autopsies performed on patients with no apparent evidence of EBV-related diseases, the viral DNA was only detected by PCR in the following: parotid gland (7/15), submandibular gland (8/20), nasopharynx (8/10), tonsil (8/10), larynx (5/6), lung (5/9), cervical lymph node (7/10), mediastinal lymph node (7/10), abdominal lymph node (4/10), spleen (6/10), thyroid (5/10), liver (1/10), pancreas (1/4), kidney (4/10), uterine cervix (1/4), ovary (1/5) and testis (1/3). These results provide a baseline for interpreting the role of EBV in carcinogenesis.

Disseminated zygomycosis masquerading as cerebral lupus erythematosus.

Zygomycosis often occurs in patients with an underlying disease, e.g., diabetes mellitus, leukemia, and lymphoma, or an immunocompromised state. This report discusses a case of a 21-year-old woman with systemic lupus erythematosus complicated by uremia, acidosis, steroid therapy, prolonged antimicrobial therapy, and disseminated zygomycosis.

Lack of correlation between expression of Epstein-Barr virus (EBV) latent membrane protein and bcl-2 oncoprotein in vivo.

AIMS: To evaluate whether there is any correlation between the expression of Epstein-Barr virus (EBV) latent membrane protein (LMP) and oncoprotein bcl-2 in the lymph node biopsy specimens of a Chinese patient with EBV-related reactive lymphoproliferation who later developed T cell lymphoma after a short period of time. METHODS: Immunohistochemistry, with a standard alkaline phosphatase antialkaline phosphatase (APAAP) method and New Fuchsin as a chromogen, was used for single staining of bcl-2 or LMP. Double immunostaining combining APAAP and indirect immunofluorescence was performed for dual labelling of LMP and bcl-2. RESULTS: bcl-2 was expressed in 10-30% of cells in the first lymph node biopsy specimen (EBV-associated lymphoproliferative disorder) and 30-50% of cells in the second lymph node biopsy specimen (T cell lymphoma). LMP was expressed in the first biopsy specimen but not in the second. Double immunostaining results showed that around 78% of LMP positive cells were bcl-2 negative and 94% bcl-2 positive cells were LMP negative. Among the very small fraction of LMP and bcl-2 double positive cells, the intensity of bcl-2 staining was heterogeneous and was not always stronger than that observed in LMP negative bcl-2 positive cells. CONCLUSIONS: The expression of bcl-2 protein is independent of LMP protein status in vivo. Several mechanisms may be involved in EBV associated lymphomagenesis, and bcl-2 induction may occur independently of LMP expression.

Improved silver technique for showing nucleolar organiser regions in paraffin wax sections.

An improved technique for the demonstration of nucleolar organiser regions (NORs) was devised. It has the advantage that any non-specific background deposits which mimic the NOR dots formed during staining are trapped by the celloidin film. The specificity of the silver staining is also increased by the postfixation treatment of sections in an acetic acid-ethanol mixture. Moreover, it is economical because of the low silver content and small volume of the incubation solution used.

Histochemistry of mucin secreting components in mucoepidermoid and adenosquamous carcinoma of the oesophagus.

AIMS: To determine the direction of differentiation of the mucin secreting components in a rare group of oesophageal tumours--oesophageal squamous cell carcinomas with prominent mucin secreting components (mucoepidermoid carcinomas and adenosquamous carcinomas). METHODS: In a review of 617 cases of primary carcinoma of the oesophagus, 16 cases of squamous cell carcinoma with prominent mucin secreting components were studied using a battery of histochemical techniques. RESULTS: The mucin produced by these tumours was mixed and included a variable content of enzyme labile sialomucin (positive for mucicarmine, periodic acid Schiff, and alcian blue, and sensitive to sialidase digestion and negative for high iron diamine-alcian blue). Retrospective analysis of endoscopic biopsy specimens taken from these tumours showed that mucin was present in five (42%) cases. CONCLUSIONS: The glandular component of this group of tumours histochemically differentiated in the direction of oesophageal glands: examination of the mucin secreting component in squamous cell carcinoma in resected specimens is therefore required for recording the true incidence of this type of tumour.

Immunohistochemical demonstration of c-erbB-2 oncoprotein in gastric adenocarcinoma: comparison of cryostat and paraffin wax sections and effect of fixation.

AIMS: To investigate the effects of fixation on the immunohistochemical demonstration of c-erbB-2 oncoprotein using paraffin wax and cryostat sections; to compare c-erbB-2 expression in non-neoplastic and neoplastic gastric tissues. METHODS: Adjacent blocks of tumour and non-neoplastic tissue from four gastrectomy specimens were put into a panel of 10 fixatives including acetone, B5, Bouin's fluid, Carnoy's fluid, buffered formalin, formol dichromate, zinc formalin, 4% paraformaldehyde, periodate-lysine-paraformaldehyde (PLP) and periodate-lysine-paraformaldehyde-dichromate (PLPD) before embedding in paraffin wax for sectioning. Similar tissue blocks were snap frozen and cryostat sections were postfixed in these fixatives, either alone or in combination, before immunostaining. RESULTS: In paraffin wax embedded sections the best fixative was PLP, and in frozen tissues the best results were obtained after fixation of cryostat sections in buffered formalin followed by cold methanol and acetone. Applying these fixatives to samples from a further 16 gastrectomy specimens, strong membrane staining of c-erbB-2 protein was found in the tumour in eight of 16 cases (50%) using paraffin wax sections, and staining was stronger in the better differentiated carcinomas. For frozen tissues, positive membrane staining was found in all gastric adenocarcinomas, but differential staining intensity associated with tumour differentiation could not be detected. CONCLUSIONS: These results indicate that fixation and paraffin wax embedding affect the results of immunohistochemical demonstration of c-erbB-2 in gastric cancer. The choice of fixative is critical in the demonstration and evaluation of c-erbB-2 protein expression by immunohistochemistry in gastric carcinomas. Staining results also vary depending on whether frozen or paraffin wax embedded tissues are studied.

Improved double immunohistochemical staining method for cryostat and paraffin wax sections, combining alkaline phosphatase anti-alkaline phosphatase and indirect immunofluorescence.

AIMS: To develop and immunohistochemical staining method for cryostat and paraffin wax sections so that two different antigens in the same section of tissues could be detected by combining immunoenzyme and immunofluorescence techniques. METHODS: This double immunohistochemical staining method combines alkaline phosphatase-anti-alkaline phosphatase (APAAP) using New Fuchsin as a chromogen and indirect immunofluorescence. RESULTS: APAAP staining for one antigen of this double immunohistochemical staining method was observed under bright field conditions alternating with immunofluorescence for another antigen under ultraviolet light. The double exposed photograph of both easily identified the two signals within the same cell. CONCLUSIONS: This double immunohistochemical staining method can overcome the disadvantages of any masking effect of the double immunoenzymatic methods and the background problems of double immunofluorescence method especially when applied to paraffin wax sections. It also permits good morphological identification of the doubly stained cells which may be of crucial importance in studies on pathology specimens.

Cytokeratin expression in non-neoplastic oesophageal epithelium and squamous cell carcinoma of the oesophagus.

The expression of cytokeratins (CK) 19, 8, 18, 13, 10 and 7 was examined in 35 cases of squamous cell carcinomas of the oesophagus (10 well-differentiated, 13 moderately-differentiated, and 12 poorly-differentiated) and the adjacent mucosa by means of a panel of monoclonal antibodies on frozen sections. The study was undertaken to assess the pattern of expression of these keratins in oesophageal tumours and its relation to the degree of differentiation. The normal oesophageal epithelia expressed CK19 in 86%, CK18 in 17% and CK13 in 14% of cases. CK8, CK10 and CK7 immunoreactivity was not observed. The tumours expressed CK19 in 86%, CK8 in 46%, CK18 in 97%, CK13 in 83%, CK10 in 34% and CK7 in 29% of cases. Thus, the so-called simple epithelial markers CK18 and CK19 occurred in the majority of oesophageal squamous cell carcinomas. CK13 (the so-called non-keratinizing squamous epithelial marker) was only infrequently demonstrated in the non-neoplastic oesophageal mucosa, and its expression was more frequent in carcinomas. CK10 was not demonstrated in non-neoplastic mucosa, but was mostly associated with well-differentiated carcinomas. We therefore conclude that the pattern of expression of cytokeratins in oesophageal carcinomas is different from that in normal oesophageal epithelia and varies with differentiation.

Epstein-Barr virus DNA in nasopharyngeal carcinomas from Chinese patients in Hong Kong.

AIMS: To investigate the presence of Epstein-Barr virus (EBV) in cases of nasopharyngeal carcinoma (NPC) in Chinese patients living in Hong Kong. METHODS: Nasopharyngeal biopsy specimens, formalin fixed and paraffin wax embedded, from 24 patients, eight with undifferentiated nasopharyngeal carcinoma, eight with well differentiated squamous carcinoma, and eight showing normal tissue histology, were analysed for the presence of Epstein-Barr virus (EBV) DNA by slot-blot hybridisation on extracted unamplified DNA, and also after amplification of EBV specific sequences by the polymerase chain reaction (PCR). RESULTS: DNA slot-blot analysis showed viral DNA in all the undifferentiated, five of the well differentiated tumours, and none of the normal biopsy specimens. PCR studies confirmed positivity in the eight undifferentiated tumours, but six of the well differentiated tumours and three of the normal biopsy specimens showed viral DNA by this method, illustrating its greater sensitivity. CONCLUSIONS: EBV genome is present in appreciable copy number in most cases of well differentiated NPC in Chinese patients in Hong Kong.

Human papillomavirus in oesophageal squamous cell carcinoma.

Thirty seven cases of oesophageal squamous cell carcinoma were studied by applying DNA slot blot analysis and in situ hybridisation using type specific probes for HPV 6, 11, 16 and 18. Cases of condyloma accuminata, cervical carcinoma, and laryngeal papilloma were used as controls. Blocks including areas of invasive carcinoma, intraepithelial neoplasia, and normal epithelium were studied in each case. No HPV genome was detectable in any of the oesophageal cases. It is concluded that these types of HPV do not have an association with oesophageal squamous cell carcinoma.

Localisation of CD10 to biliary canaliculi by immunoelectron microscopical examination.

Common acute lymphoblastic leukaemia antigen (CALLA) was first characterised in lymphoid leukaemic cells. The antigen is present in different stages of lymphoid cell differentiation as well as in subsets of myeloid cells, and further studies have also shown its presence in non-lymphoid tissues. The recent cloning and sequencing of the gene permitted deduction of its amino acid sequence which is identical with the human membrane-associated enzyme, neutral endopeptidase. Strong immunostaining for CALLA was detected in the human liver with a canalicular pattern. Immunoelectron microscopy also confirmed that the antigen was localised only in the area of the bile canaliculi. Although the function of neutral endopeptidase in the canaliculi is unknown, this antigen may prove useful in the study of biliary function and diseases.

Chemotherapy for early-stage gastrointestinal lymphoma.

A total of 176 patients with gastrointestinal lymphomas were reviewed. According to a modified staging classification, 51 of them had stage I/II disease and the remaining 125 had stage III/IV disease. In most cases (68%), the histology was intermediate-grade according to the NIH working formulation, and the B-cell immunophenotype was involved in 89% of the 45 cases with a known immunophenotype. The primary site was the stomach in 56% of cases and the bowel in 44%. A significantly higher proportion (P = 0.001) of those with bowel lymphomas had stage III/IV disease (88% vs 57%). The primary gastrointestinal lesion was resected in 122 patients, including all 51 cases of stage I/II disease. In all, 8 stage I/II patients were given radiotherapy alone following surgery and the other 43 underwent chemotherapy; of the latter, 19 received additional radiotherapy following chemotherapy. Chemotherapy was also given to 112 stage III/IV patients, 42 of whom underwent additional radiotherapy. Factors associated with a poorer prognosis included advanced disease, bowel lymphoma and advanced age. Although the complete response (CR) rate according to disease stage was similar, stage I/II patients receiving chemotherapy showed a significantly lower relapse rate, better disease-free survival following CR and improved survival as compared with those receiving radiotherapy alone. However, additional radiotherapy following chemotherapy did not further improve the clinical outcome.

Intensive chemotherapy for adult lymphoblastic lymphomas.

A total of 20 adults patients presenting with previously untreated lymphoblastic lymphoma underwent an intensive chemotherapy protocol. Either the BACOP or the m-BACOD regimen was used for induction. If the patients achieved a complete clinical remission (CR) after three courses, they were given intensive consolidation and maintenance chemotherapy based on a protocol that was modified from the L10/L17M regimen of the Memorial Sloan-Kettering group for acute lymphoblastic leukaemia and lymphoblastic lymphoma. Patients exhibiting localised areas of bulky disease were given additional involved-field radiotherapy. In all, 15 (75%) men and 5 (25%) women were entered in this study. Their median age was 28 years (mean, 30 years; range, 12-64 years). Overall, 3 (15%) had stage II disease, 3 (15%) had stage III disease and 14 (70%) had stage IV disease; 7 (35%) patients exhibited B symptoms and 4 (20%) had bulky disease. The overall (CR) rate was 10/20 (20%), and that following BACOP and m-BACOD therapy was 4/8 (50%) and 6/12 (50%), respectively. In all, 7 of the 10 complete responders (70%) relapsed. The disease-free survival of the ten who achieved a CR was 23% at 3 years. The overall survival of all 20 patients at 3 years was only 37%, and there were very few long-term survivors. More effective treatment for adult lymphoblastic lymphoma is required.

Expression of p53 in oesophageal squamous cell carcinoma in Hong Kong Chinese.

We studied p53 overexpression in a series of 99 primary oesophageal squamous cell carcinomas (28 well-differentiated, 42 moderately-differentiated and 29 poorly-differentiated squamous cell carcinomas) from Chinese patients using the p53 protein specific mouse monoclonal antibody DO-7 on paraffin sections. The p53 protein was detected in 30% (30 cases) of the tumours. A significantly higher positive rate was noted in the poorly-differentiated tumours (11% for the well-differentiated, 31% for the moderately-differentiated and 48% for the poorly-differentiated tumours). In addition, strong positive p53 staining was identified only in the less differentiated tumour cells in the periphery of the tumour cell nests in all the cases and the expression was weaker in the better differentiated foci. The central keratinizing areas and the immediately adjacent tumour cells were always negative for p53. The adjacent normal oesophageal mucosa was all negative for p53 protein but the non-invasive dysplastic epithelium next to the tumours could also be strongly positive for p53 protein (four out of 14 cases in which the dysplastic epithelium adjacent to the tumour was adequately sampled). In two out of these four cases, the dysplastic epithelium showed staining for p53; even the adjacent invasive tumour was negative for p53. It is concluded that there is a strong relationship between p53 overexpression and tumour cell differentiation in oesophageal squamous carcinoma and overexpression of p53 can occur in non-invasive tumour cells.