Stainability of lung cancer cells with Leu-7 and OKT-9 monoclonal antibodies

One hundred and one cases of lung cancer were immunohistochemically studied for stainability with Leu-7 (anti-myelin fiber associated glycoprotein) and OKT-9 (anti-transferrin receptor) monoclonal antibodies. All small cell carcinomas and carcinoid tumors were positively stained by Leu-7, and 22 of 41 differentiated adenocarcinomas were also positively stained, especially well-differentiated Clara cell-type adenocarcinoma, (11/14 cases). However, only one of 26 squamous cell carcinomas, one of six large cell carcinomas, one of six adenosquamous carcinomas and none of 16 poorly differentiated adenocarcinomas were stained by Leu-7. On the other hand, all squamous cell carcinomas (26/26 cases), 10 of the 16 poorly differentiated adenocarcinomas, four of the six large cell carcinomas showed positive membranous staining with OKT-9. However, only one of 41 differentiated adenocarcinomas and no small cell carcinomas nor carcinoid tumors were stained by OKT-9. The stainability of lung cancer by these antibodies is discussed.     

免疫标志物及AB／PAS黏液染色在子宫颈癌分型中的应用研究

目的：探讨免疫标志物细胞角蛋白7（cytokeratin 7,CK7）、低分子质量细胞角蛋白CAM5．2、癌胚抗原（carcino-embryonic antigen,CEA）、癌基因p63、高分子质量细胞角蛋白34βE12及阿利新蓝-过碘酸雪夫氏（alcian blue-periodic acid schiff,AB／PAS）黏液染色在子宫颈癌分型中的意义。方法采用免疫组织化学EnVision法检测CK7、CAM5．2、CEA、p63、34βE12在59例子宫颈癌中的表达,结合AB／PAS黏液染色,分析上述免疫标志物及特殊染色与子宫颈癌分型的关系。结果 CK7、CAM5．2、CEA在子宫颈腺癌（6例）、腺鳞癌（4例）的腺癌细胞质中均有100．0％的表达,低分化鳞癌（34例）中CK7、CAM5．2阳性率分别为52．9％、32．4％,而中及高分化鳞癌（15例）中不表达。CEA在鳞癌中有不同程度表达,中及高分化鳞癌仅在角化区域表达,低分化鳞癌中阳性率44．1％;p63在腺癌中不表达,在鳞癌中100．0％表达;34βE12在腺癌及鳞癌中均有100．0％表达,鳞癌中强度高,腺癌中较弱;AB／PAS在子宫颈腺癌、腺鳞癌、低分化鳞癌中发生率分别为83．3％（5／6）、75．0％（3／4）、5．9％（2／34）,但表达程度很弱。结论 CK7、CAM5．2、p63、34βE12联合检测对子宫颈癌的分型有重要意义。AB／PAS黏液染色对子宫颈癌有一定的鉴别作用。     

人肺癌CD44蛋白和基因的异常表达与淋巴结转移的相关性

目的：研究CD44蛋白和基因在人肺癌组织中的异常表达,以及这种异常表达与淋巴结转移的相关性。方法：应用SP免疫组织化学染色方法和RT－PCR／Southern方法,研究人肺癌组织中,CD44蛋白和基因的表达,并应用QTM970计算机图像分析仪,对免疫组织化学染色结果进行定量分析。结果：SP免疫组织化学染色和计算机图像定量分析结果显示,非小细胞肺癌（non-small cell lung carcinomas,NSCLCs)组织比癌旁组织,有淋巴结转移比无淋巴结转移的非小细胞肺癌组织染色强度深,差异有极显著意义（F＝16．67,P＜0．01）;RT－PCR／Southern杂交结果显示,有淋巴结转移较无淋巴结转移的非小细胞肺癌表达更多的CD44mRNA异常转录子（X^2=12.13,P＜0．01）。结论：CD44蛋白和基因在人非小细胞肺癌中表达中出现异常表达,这种异常表达在人非小细胞肺癌的发展和淋巴结转移中起重要作用。检测CD44蛋白和基因的表达,可能对判断非小细胞肺癌病人的预后、预测淋巴结的转移趋势,具有重要的临床意义。     

Clinical and biological characteristics of clear cell carcinomas of the ovary in FIGO stages I-II

Clear cell carcinoma of the ovary is considered to be a specific subtype among the epithelial ovarian malignancies. To characterize clear cell carcinomas in early FIGO stages (I-II) with regard to clinical and biological properties, a retrospective study was performed to compare these tumors with other histological subtypes. From a complete series of 226 patients with epithelial ovarian carcinomas in FIGO stages I-II, 28 patients with clear cell carcinomas were selected and the clinical and biological characteristics of these tumors were compared with the remaining non-clear cell carcinomas. All patients underwent primary staging laparotomy followed by adjuvant radiotherapy or chemotherapy. The apoptosis regulators p53, bcl-2 and bax, and the growth factor receptors EGFR and HER-2/neu were analyzed by immunohistochemical techniques and DNA analysis was performed by flow cytometry. Clear cell carcinomas stained negative for p53 significantly more often than other histological subtypes. Positive EGFR staining was seen more frequently in serous carcinomas than in the clear cell carcinomas. Aneuploid DNA status was seen more frequently in clear cell carcinomas than in other histological subtypes and tetraploid tumors made up 50% of the non-diploid tumors. Clear cell tumors were frequently (64%) found in FIGO stages IC and IIC and this was more common than for non-clear cell tumors. No difference was found in the rate of tumor recurrences or survival for patients with clear cell and non-clear cell carcinomas. Clear cell carcinomas of the ovary should be regarded as a separate entity among the epithelial ovarian carcinomas and they differ with regard to both clinical and biological characteristics when compared with non-clear cell tumors.     

Transplantation of human tumors in nude mice.

Ninety-one human tumors, including various common carcinomas, low-grade malignant tumors, and benign tumors, were transplanted into athymic nude mice. Tumor take was confirmed histologically for 22 neoplasms at the initial transplantation, and 14 serially transplantable tumors were established, including some hitherto unestablished or unreported, such as lung and hepatic cell carcinomas. Among the 91 tumors were 21, 14, and 13 carcinomas of the lung, stomach, and breast, respectively. Transplantability was highest in lung carcinomas (10/21), followed by gastric carcinomas (2/14) and breast carcinomas (1/13). Morphology of original tumors was retained well in most transplanted tumors, but desmoplastic or scirrhous tumors, such as gastric and breast carcinomas, tended to become medullary with a decrease in amount of tumor stroma. The ability to produce mucin in gastric carcinomas or melanin in malignant melanoma was maintained in serially transplantable tumors. In addition, ectopic production of adrenocorticotropin and beta melanocyte-stimulating hormone continued in a transplanted small cell carcinoma of the lung. Preliminary results were obtained on hormone dependency of the transplantable breast carcinoma and on alpha1-fetoprotein in the transplantable hepatic cell carcinoma.     

Gene amplification and overexpression of EGF receptor in squamous cell carcinomas of the head and neck

Tumours of the head and neck were examined for gene amplification and expression of the epidermal growth factor (EGF) receptor by Southern blot and Western blot analyses. The EGF receptor gene was found to be amplified in four (19%) of 21 squamous cell carcinomas. The EGF receptor was overexpressed in eight (53%) of 15 squamous cell carcinomas examined, including all four tumours showing gene amplification. No amplification or overexpression of the EGF receptor gene was detected in any of nine malignant or eight benign tumours of other types of the head and neck. The tumours showing amplification and/or overexpression of the EGF receptor gene (8/15) were all identified histologically as well differentiated squamous cell carcinomas, whereas none of the histologically less differentiated squamous cell carcinomas (0/9) showed amplification and/or overexpression of the EGF receptor gene. Within our sample set, no correlation was evident between amplification and/or overexpression and the clinical stage or tumour site. Our results support the possible involvement of gene amplification and overexpression of the EGF receptor in a subclass of squamous cell carcinomas of the head and neck.     

Analysis of human cancer DNA for DNA sequences of human adenovirus type 4.

We investigated whether human adenovirus type 4 (Ad4) might cause cancer in humans. Cell culture and animal model studies indicated that Ad4-induced human tumors should contain Ad4 DNA sequences. Thus Ad4 DNA was labeled in vitro (sp act, approximately 10(7) 3H counts/min/microgram) and hybridized in liquid phase with DNA extracted from human tumors. No viral sequences were detected in 31 squamous cell carcinomas of the lung, 5 adenocarcinomas of the lung, 4 oat cell carcinomas of the lung, 4 carcinomas of the stomach, 10 carcinomas of the colon, 3 carcinomas of the kidney, 3 carcinomas of the breast, 2 carcinomas of the ovary, and 6 Hodgkin's lymphomas. Reconstruction experiments with added Ad4 DNA indicated that the probe detected about 1 copy of 5% of the Ad4 genome per tumor cell. Therefore, these data were strong evidence (but did not prove) that none of these particular tumors were induced by Ad4.     

Determination of HER-2/neu amplification and expression in tumor tissue and cultured cells using a simple, phenol free method for nucleic acid isolation.

A rapid, simple and non-toxic procedure for the simultaneous isolation of DNA and RNA from tumor tissue and cells grown in vitro is described. Guanidinium isothiocyanate was used for homogenization of tumor tissue and for cell lysis. Separation of proteins, DNA and RNA was carried out by isopycnic centrifugation in cesium trifluoroacetate. DNA was further purified by salting out residual protein. Nucleic acids prepared by this method from 47 primary human carcinomas and 17 human cell lines were analysed for amplification and expression of the HER-2/neu proto-oncogene. 2- to 10-fold amplification of HER-2/neu was noted in 7/22 mammary carcinomas (32%) and in 4/14 ovarian carcinomas (28%). No amplification of the proto-oncogene was found in 4 laryngeal carcinomas, 1 pharyngeal carcinoma, 2 retrolingual carcinomas, 3 gastric carcinomas and 1 kidney carcinoma. HER-2/neu overexpression was observed in 6/22 of mammary carcinomas (27%) and 7/14 of ovarian carcinomas (50%). No overexpression was found in all other carcinomas studied. Concordance between amplification and overexpression was noted in 3 mammary and 4 ovarian carcinomas, respectively. 3 mammary and 3 ovarian carcinomas showed overexpression without amplification. 5 human mammary carcinoma cell lines showed both amplification and overexpression of HER-2/neu. In two mammary carcinoma cell lines (MDA MB-453 and ZR 75-1) overexpression was noted without amplification of the proto-oncogene. These data combine to suggest that mechanisms other than gene amplification may also lead to overexpression of the HER-2/neu protooncogene in cancer cells.     

Lectin bindings in non-neoplastic esophageal epithelium and esophageal carcinomas

Lectin binding was examined histochemically in 22 cases of primary esophageal carcinomas (10 well differentiated, 8 moderately differentiated and 3 poorly differentiated squamous cell carcinomas, and 1 undifferentiated carcinoma) and was compared with the adjacent non-neoplastic epithelium by means of a panel of 10 different lectins (RCA-I, WGA, Con A, LCA, SEA, UEA-I, HPA, PNA, DBA and GS-I) on formalin-fixed paraffin-embedded sections. In the non-neoplastic epithelium, RCA-I and WGA showed basal/parabasal binding, Con A, LCA, SEA, UEA-I, HPA and PNA revealed prickle cell binding, while DBA and GS-I only stained the surface cells of the squamous cell layer. In squamous cell carcinomas, no clear difference was evident regarding the grade of differentiation. However, basal/parabasal specific lectins were expressed in all the cases, the prickle cell-specific lectins were expressed less frequently, whereas lectins expressed at the surface cells of the squamous cell layer were only infrequently expressed. Therefore, basal/parabasal cell specific lectins were widely preserved in squamous cell carcinomas. One case of undifferentiated cancer tested was devoid of all the lectins.     

Atypical cysts and carcinomas of the kidneys in the phacomatoses. A quantitative DNA study using static and flow cytometry

Reported are the pathologic features of atypical cysts and/or renal cell carcinomas found in the kidneys of four patients having either tuberous sclerosis or Hippel-Lindau disease. In addition, cellular DNA contents of the cells lining the atypical cysts and comprising the carcinomas were quantitated using both static and flow cytometric techniques. These studies showed that cysts lined by atypical epithelial cells are frequently present in renal parenchyma adjacent to the renal carcinomas, and that the cytologic features of atypical cells lining the cysts were essentially the same as the cytologic features found in the adjacent well-differentiated, renal cell carcinomas. DNA quantitative studies revealed that both the renal cell carcinomas and the atypical cyst lining cells had the same DNA indices and were essentially DNA euploid. In this patient group these findings are consistent with the hypothesis that the atypical cyst lining cells evolve into the renal cell carcinomas; however, they do not prove this proposed but likely sequence.     

Carcinogenicity of subcutaneously injected N-nitrosoheptamethyleneimine in European hamsters.

Male and female European hamsters (45 of each sex) recieved sc injections once weekly for life of N-nitrosoheptamethyleneimine (NHMI) at one-fifth the median lethal dose (LD50) (females: 44 mg/kg body wt; males: 66 mg/kg body wt), one-tenth the LD50 (females: 22 mg/kg body wt; males: 33 mg/kg body wt), or one-twentieth the LD50 (females: 11 mg/kg body wt; males: 16.5 mg/kg body wt). Survival times for both males and females were dependent on the dose of NHMI. Pulmonary neoplasms were induced in almost all the treated animals. They were histologically diagnosed as adenocarcinomas, squamous cell carcinomas, and mixed cell carcinomas. In addition, nasal cavity tumors developed in all hamsters of all treatment groups; these were papillomas, squamous cell carcinomas, and a few adenocarcinomas. Only 1 tumor of the larynx and 1 tumor of the trachea were observed. Several papillomas and a few carcinomas were also detected in the forestomach. The results were discussed with reference to previous findings in rats and Syrian golden hamsters.     

PD-L1 expression in small cell neuroendocrine carcinomas

Small cell lung cancer and extrapulmonary small cell carcinomas are the most aggressive type of neuroendocrine carcinomas. Clinical treatment relies on conventional chemotherapy and radiotherapy; relapses are frequent. The PD-1/PD-L1/PD-L2 pathway is a major target of anti-tumour immunotherapy. Aberrant PD-L1 or PD-L2 expression may cause local immune-suppression. Here we investigated expression of PD-1 and its ligands by immunohistochemistry and RNA-seq in small cell carcinomas.PD-L1 and PD-1 protein expression were analysed in 94 clinical cases of small cell carcinomas (61 pulmonary, 33 extrapulmonary) by immunohistochemistry using two different monoclonal antibodies (5H1, E1L3N). RNA expression was profiled by RNA-seq in 43 clinical cases.None of the small cell carcinomas showed PD-L1 protein expression in tumour cells. PD-L1 and PD-1 expression was noticed in the stroma: Using immunohistochemistry, 18.5% of cases (17/92) showed PD-L1 expression in tumour-infiltrating macrophages and 48% showed PD-1 positive lymphocytes (45/94). RNA-seq showed moderate PD-L1 gene expression in 37.2% (16/43). PD-L1 was correlated with macrophage and T-cell markers. The second PD-1 ligand PD-L2 was expressed in 27.9% (12/43) and showed similar correlations.Thus, the PD-1/PD-L1 pathway seems activated in a fraction of small cell carcinomas. The carcinoma cells were negative in all cases, PD-L1 was expressed in tumour-infiltrating macrophages and was correlated with tumour-infiltrating lymphocytes. Patients with stromal PD-L1/PD-L2 expression may respond to anti-PD-1 treatment. Thus, evaluation of the composition of the tumour microenvironment should be included in clinical trials. Besides conventional immunohistochemistry, RNA-seq seems suitable for detection of PD-L1/PD-L2 expression and might prove to be more sensitive.     

A phase II study of CPT-11, a camptothecin derivative, in patients with primary lung cancer. CPT-11 Cooperative Study Group

A Phase II study of CPT-11, a new camptothecin, was performed in patients with primary lung cancer. Patients with previously untreated non-small cell carcinomas (group A), or previously treated non-small cell carcinomas (group B), and with small cell carcinomas (group C), were enrolled in this study. CPT-11 was given at a dose of 100 mg/m2 i.v. infusion once a week for three weeks or more. Out of 153 patients enrolled, 128 (A: 67; B: 26; C: 35) were assessed to be evaluable for response by an extramural review committee. Response rates were 34.3% (23/67) for A, 0% (0/26) for B and 37.1% (13/35) for C. The response rate was 50% for previously untreated patients (4/8), and 33.3% for previously treated patients (9/27) including 2 complete responses in the group C. Major toxicities were leukopenia, nausea/vomiting, diarrhea, anorexia and alopecia. Leukopenia and diarrhea were considered to be dose limiting toxicities, but they were reversible. It was, however, suggested that some patients should be monitored carefully for severe reactions and delay in recovery. The results showed that CPT-11 was highly effective against non-small cell and small cell carcinomas of the lung.     

Induction of pancreatic carcinomas in rats with N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine: histopathology

The carcinogenicity of N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine (HPOP) for rat pancreas was evaluated. Two-week-old male LEW rats were given a single ip injection of HPOP, 160 mg/kg body weight; the rats were autopsied 4, 6, or 12 months later. Histologic examination showed that the pancreata contained multiple foci of atypical acinar cells and nodules of atypical acinar cells (AACN), acinar cell adenomas, localized carcinomas, and carcinomas. The incidence of carcinomas was 77%. The carcinomas were composed of poorly differentiated acinar cells and ductlike structures. Pancreatic ducts were unaffected. The prominence of AACN, the histologic type of the neoplasms, and the absence of hyperplastic changes in ductal epithelium suggest that the pancreatic carcinomas were derived from acinar cells. The incidence of liver cell carcinomas and pulmonary adenomas was similar to that of localized pancreatic carcinomas. Neoplasms of other organs were less frequent. HPOP has been shown to induce pancreatic carcinomas in hamsters but has not previously been reported to be a pancreatic carcinogen in rats.     

C-jun oncogene expression in nonsmall cell lung-cancer

Primary lung cancer specimens of the non-small cell (NSC) types were examined for expression of the c-jun gene at the protein level by immunocytochemistry using the polyclonal antibody c-jun 890. The results obtained revealed that the c-jun p39 protein was expressed in 82% (18/22) of squamous cell carcinomas, 77% (10/15) of adenocarcinomas and 40% (2/5) of large cell anaplastic carcinomas, but no staining was seen in the non-cancer cases. Positive immunostaining was also seen in 16.7% (3/18) of the nonneoplastic bronchial cells of positively stained squamous cell carcinomas and in 40% (4/10) of adenocarcinomas. We suggest that elevated expression df c-jun p39 protein which was seen in a total of 75% (30/40) of the NSCLC, plays a role in lung cancer.     

Expression of p53 in premalignant and malignant squamous epithelium.

Using three antibodies (JG8, CM-1 and 1081) directed to the p53 protein, strong positivity was found in 16/47 (34.0%) of mucosal squamous cell carcinomas of the head and neck and in two squamous carcinoma cell lines (LICR-LON- HN5 and HN6Rr). The presence of the mutant p53 was confirmed in the cell lines as substitutions in exon 7 (codon 238, TGT greater than AGT) and exon 5 (codon 152, CCG greater than CTG) respectively. Positive staining was seen only in the undifferentiated cells and progressively lost as the cells keratinized, both in the tumour specimens and in the cell lines. Similar results were seen in areas of dysplasia, well removed from the site of the primary tumour. Staining of epidermal lesions showed positivity in 2/12 (16.6%) cases of Bowen's disease, 0/12 (0.0%) cases of solar keratosis, 0/10 (0.0%) basal cell carcinomas and in 3/20 (15.0%) squamous cell carcinomas. These results are discussed in relation to the multifocal origin of squamous cell carcinomas, the role of p53 mutations in squamous cell carcinomas from different sites and the significance of the 'basal' distribution of p53 as a normal growth regulator. The possible significance of the distribution of p53 in squamous epithelium as it relates to papilloma virus infection is also considered.     

Gastric and intestinal phenotypic expressions of human signet ring cell carcinomas revealed by their biochemistry, mucin histochemistry, and ultrastructure

Gastric and intestinal phenotypic expression in 37 surgically obtained primary signet ring cell carcinomas, five of their metastases to lymph nodes, and three signet ring cell carcinomas transplanted into nude mice were determined by biochemical, mucin, histochemical, and ultrastructural studies. Crude extracts of cancer tissues were used for measurements of pepsinogen isozymes, sucrase, aminopeptidase (microsomal), and alkaline phosphatase. Histochemical staining of mucin by paradoxical concanavalin A, the galactose oxidase-Schiff sequence and sialidase-galactose oxidase-Schiff, and the periodate-borohydride technique/potassium hydroxide/periodic acid-Schiff procedure was performed. The procedures allowed clear definition of pyloric gland, surface mucous, small and large intestinal goblet, and intestinal absorptive cell types. Of 40 specimens examined, 19 consisted entirely of gastric-type cells, and three entirely of intestinal-type cells. The others consisted of mixtures of gastric and intestinal-type cells. The observed high incidence of intestinal-type cells in signet ring cell carcinomas suggested that intestinal-type cells develop independently from intestinal metaplasia within signet ring cell carcinomas (diffuse-type gastric cancers), which probably originate from nonmetaplastic gastric mucosa.     

Lung carcinoma-associated atypical adenomatoid hyperplasia, squamous cell dysplasia, and chromosome alterations in non-neoplastic bronchial mucosa

This article analyzes phenotype and genotype alterations of the lung in association with lung cancer. The frequency of phenotype preneoplastic lesions (atypical adenomatoid hyperplasia (AAH) and squamous cell dysplasia (SCD)) was analyzed at distinct distances from the tumor boundary in 150 lung carcinomas. AAH was noted in 19/150 (13%) cases and more frequently seen in adeno carcinomas, squamous cell dysplasia was noted in 46/150 (31%) cases and more frequently seen in squamous cell carcinomas. The degree of cellular atypia decreased with increasing distance from tumor boundary in both AAH and SCD. At similar distances, genotype (chromosome) alterations of surrounding bronchial mucosa were studied in additional 55 primary and secondary lung tumors by karyotype analysis. Numerical chromosome aberrations occur frequently in primary lung carcinomas and adjacent bronchial mucosa, and affect at average 4.5/10 metaphases in primary lung cancer and 2/10 metaphases in metastases. Most abnormal metaphases were induced by chromosome losses, only few by additional copies, i.e. trisomy, etc. Losses of y chromosome were seen in both malignancy and adjacent bronchial mucosa, and interpreted as "tumor related", losses of chromosome 21 in adjacent bronchial mucosa were non-tumor related in adenocarcinoma and metastases, losses of chromosome 19 in adjacent bronchial mucosa occurred independently in squamous cell and large cell carcinomas. The data suggest the hypothesis that preneoplastic lesions in the lung might be partly induced by the tumor itself.