Nup88 expression is associated with myometrial invasion in endometrial carcinoma.

BACKGROUND: The nuclear pore complex protein Nup88 has been shown in previous studies to be overexpressed in tumor cells and to be associated in breast cancer with all clinical and biological features defining a more aggressive phenotype. METHODS: In this pilot study, Nup88-mRNA expression was studied in a series of 29 endometrial carcinomas, of which 27 belonged to the endometrioid variety, the remaining 2 being papillary serous tumors, to verify if Nup88 plays a similar role in endometrial carcinoma as the one described in breast cancer. The tumor samples were obtained directly at the operating suite and fresh frozen at the time of hysterectomy. Nup88-mRNA expression was studied in them by means of differential reverse transcriptase-polymerase chain reaction. Nup88-mRNA expression was correlated with the clinical features of the tumors. RESULTS: A significant correlation (r = 0.41, P = 0.027) was found between growing levels of Nup88-mRNA expression and depth of myometrial invasion. There was no correlation between Nup88-mRNA expression and the other 2 available clinical parameters, that is, tumor grade (r = 0.05, P = 0.79) and surgical stage (r = -0.18, P = 0.34). CONCLUSIONS: From these results, it is concluded that Nup88 expression seems indeed to be associated with a distinct feature of tumor aggressiveness (myometrial invasion) in endometrial carcinoma and that larger studies are therefore probably worthwhile.

Bcl-2 expression in breast cancer: a comparative study at the mRNA and protein level.

BACKGROUND: Bcl-2 is one of the most important antiapoptotic genes. Although it facilitates the survival of tumor cells, its expression has been consistently associated with a better prognosis for breast cancer. Virtually all studies on Bcl-2 conducted in breast cancer have been carried out by means of immunohistochemistry. The aim of this study was to examine for the first time the expression of Bcl-2 in a series of human breast cancer, both at the mRNA and protein level. MATERIALS AND METHODS: One hundred samples from previously untreated human breast cancers were used; Bcl-2 expression was determined both by means of immunohistochemistry using the bcl-2/100/D5 monoclonal antibody and differential RT-PCR. Additionally, the expression of hormone receptors (ER and PR), c-erb-B2, p53 and the proliferation-associated Ki-67 antigen were also studied by means of immunohistochemistry as part of the standard pathological workup. RESULTS: Any degree of immunohistochemical staining correlated significantly and inversely with c-erb-B2 expression (p = 0.0008), nuclear grade 3 (p = 0.0015), a Ki-67 labeling index > 10% (p = 0.02) and tumor size (p = 0.048), and in a direct fashion with estrogen (p = 0.0003) and progesterone receptor expression (p = 0.0002). mRNA expression of the Bcl-2 gene showed a significant inverse correlation with c-erb-B2 (p = 0.016) and p53 (p = 0.014) expression, as well as with a nuclear grade 3 (p = 0.006), and a direct correlation with estrogen (p = 0.0004) and progesterone (p = 0.001) receptor expression, as well as with nodal invasion (p = 0.04). CONCLUSION: The study of Bcl-2 expression in breast cancer by means of either immunohistochemistry or RT-PCR yields very similar results. In spite of its role opposing tumor cell death, Bcl-2 is associated with biological features of the tumors which define a better intrinsic prognosis, such as hormone receptor expression, low proliferation and absence of c-erb-B2 and mutant p53 expression. This may in great part explain why Bcl-2 expression has been invariably found to correlate with a better prognosis of breast cancer.

Positive correlation between the expression of X-chromosome RBM genes (RBMX, RBM3, RBM10) and the proapoptotic Bax gene in human breast cancer.

In a recent report, it has been postulated that the ubiquitous RBM proteins might constitute a novel family of apoptosis modulators. We measured the expression of the X-chromosome RBM genes (RBMX, RBM3, and RBM10) in 122 breast cancers by means of differential RT-PCR. Using the same method, we also studied the expression of the apoptosis-related genes Bcl-2 and Bax. Markers of hormone dependence (estrogen and progesterone receptors), proliferation (Ki67 and DNA-ploidy), angiogenesis (VEGF and CD105), as well as oncogene (c-erb-B2), and tumor suppressor gene (p53) expression were also analyzed. The expression of all X-chromosome RBM genes was significantly associated with the expression of the proapoptotic Bax gene (RBMX, P=0.039; RBM3, P<0.001; RBM10 large variant, P<0.001; RBM10 small variant, P<0.001). Furthermore, the expression of both RBM10 variants was significantly associated with the expression of the VEGF gene (large variant, P=0.004; small variant, P=0.003). We also found an association of borderline significance (P=0.05) between the expression of RBM3, the large variant of RBM10 and wild-type p53. Expression of the small RBM10 variant, finally, was associated with high proliferation of the tumors (Ki67>or=20%; P=0.037). The expression of both RBM10 variants seems to be interdependent to a significant degree (r=0.26, P=0.006). From these results, it seems that the X-chromosome, through its RBM genes, plays a formerly unknown role in the regulation of programmed cell death (apoptosis) in breast cancer.

Proliferation measurement in breast cancer by two different methods.

BACKGROUND: Different studies show that proliferation measurement in breast cancer may have an independent prognostic value. In the present study, tumor proliferation in breast cancer was analyzed by two radically different methods according to the technique used (immunohistochemistry and flow cytometry), associated costs and necessary equipment. The aim was to evaluate which method discriminates better between tumors with high and low proliferation in relation to all other available clinical and biological parameters. MATERIALS AND METHODS: Two hundred and eighty breast cancers (231 ductal infiltrating, 30 lobular, 19 or less frequent varieties) were studied. The post-surgical staging was as follows: 164 pT1, 87 pT2, 7 pT3, the remaining 22 were multifocal, diffuse tumors. Axillary nodal invasion was found in 99 cases (35.4%). Proliferation was studied by means of flow cytometry (DNA index and S-phase) in fresh tumor tissue and immunohistochemistry (Ki67) in paraffin-embedded tissue. Furthermore, hormone receptor (estrogen receptor, ER; progesterone receptor, PR), c-erb-B2 and p53 expressions were studied using the same method. Finally, histological and nuclear grade, tumor size and axillary nodal invasion were also included as variables of the study. RESULTS: A DNA index >1 (aneuploidy) correlated significantly with histological grade 3 (p = 0.01), nuclear grade 3 (p < 0.0001), nodal invasion (p = 0.007), absence of ER (p = 0.006) and of PR (p = 0.002), c-erb-B2 expression (p = 0.008), p53 expression (p = 0.007) and tumor size (p = 0.01). An expression of Ki67 in 20% or more of tumor cell nuclei, on the other hand, correlated significantly with histological grade 3 (p < 0.0001), nuclear grade 3 (p < 0.0001), absence of ER (p < 0.0001) and of PR (p < 0.0001), c-erb-B2 expression (p < 0.0001), p53 expression (p < 0.0001) and tumor size (p = 0.0005), but not with nodal invasion. CONCLUSION: Although flow cytometry provides additional data (association with nodal invasion), the study of Ki67 expression emerges from this study as a simple, inexpensive and reliable method to study the proliferation rate of breast cancer.

Characterization of a human Bid homologue protein from Gallus gallus.

Bid protein, a member of the "BH3-only" subgroup of Bcl-2 family, plays a critical role in mammalian apoptosis regulation. In this study, we have cloned the chicken Bid gene, which encodes a 193 amino acid protein and shares 40% homology with human and mouse Bid proteins. Bid sequence comparison emphasises the conservation of both the functional domain BH3 and the proteolytic cleavage sites. An induction of apoptosis by chicken Bid and the cleavage of the protein, after TNFalpha treatment, were also demonstrated. In addition, mRNA Bid expression was detected along all embryo stages and tissues examined, suggesting a role for this protein in the developmental process. This is the first report demonstrating the functionality of a "BH3-only" protein in chicken.

Circulating hormone levels in breast cancer patients. correlation with serum tumor markers and the clinical and biological features of the tumors.

BACKGROUND: Breast cancer grows in a hormone-rich environment which influences its biological features and thus, ultimately, its clinical behavior. Circulating hormone levels were measured in premenopausal and postmenopausal breast cancer patients prior to surgery, and correlated with all available clinical and biological features of the tumors and with serum tumor marker levels. MATERIALS AND METHODS: FSH, LH, 17-beta-estradiol, progesterone and prolactin were measured in 112 previously untreated breast cancer patients (54 premenopausal and 58 postmenopausal). Serum tumor markers (CEA, CA 15.3, CA-125 and Ca19.9) were measured at the same time. All tumors were studied after surgery for hormone receptor (ER and PR), Ki67, c-erb-B2 and p53 expression by means of immunohistochemistry, and for DNA-ploidy by means of flow cytometry. RESULTS: In premenopausal patients, high gonadotropin levels correlated directly with c-erb-B2 overexpression by the tumors (FSH: p=0.02; LH: p=0.05). High estradiol levels correlated inversely (p=0.009) with Ki67 expression. In postmenopausal patients, high estradiol levels were inversely related to c-erb-B2 expression by the tumors (p=0.03), and high progesterone levels were also inversely related to Ki67 expression by the tumors (p=0.05). FSH levels correlated inversely (p = 0.02) with circulating carcinoembryonic antigen (CEA) levels. CONCLUSION: Circulating estradiol levels seem to be associated with a less proliferative breast cancer phenotype. FSH and LH levels, on the other hand, seem to exert dual actions in premenopausal and postmenopausal breast cancer patients.

Developing chick embryos express a protein which shares homology with the nuclear pore complex protein Nup88 present in human tumors.

Nup88 is a nuclear pore complex protein which is overexpressed in a variety of human tumors of the stomach, colon, liver, pancreas, breast, lung, ovary, uterus, prostate and kidney. A monoclonal antibody crossreacting with the yeast Candida albicans and Nup88 was used to investigate the expression of cross-reactive antigens in chick embryos, in an attempt to identify an experimental model for studying the role played by Nup88 during cell development and differentiation. All cells in the trilaminar embryo were labeled with the antibody, but as development advanced and organogenesis was completed, expression of the corresponding antigen became more restricted. Thus, some structures continued to be intensely labeled (skin epithelium, oropharyngeal endothelium, perichondral mesenchymal tissue), whereas others ( muscular tissue, vascular endothelium, respiratory endothelium, digestive tract mucosa, peripheral nerves, medullary white matter and the retinal axons) were more moderately stained. No immunoreactivity was observed in the medullary grey matter or cartilage. A specific band of 53 kDa observed by Western blotting of chick embryo extracts suggested that the chicken antigen recognized by the monoclonal antibody is the homologue of human Nup88, which is associated with the high proliferation and low differentiation of tumor cells. The present results indicate that the role of Nup88 in cell differentiation and organ development could be fruitfully investigated using the developing chick embryo as an experimental model.

mRNA expression of the angiogenesis markers VEGF and CD105 (endoglin) in human breast cancer.

BACKGROUND: Both VEGF and CD105 (endoglin) have been identified as markers of tumor angiogenesis and prognosis in breast cancer. They have always been studied in this kind of tumor by means of immunological methods. OBJECTIVE: To study, by means of reverse-transcription polymerase chain reaction (RT-PCR), the expression of VEGF and CD105 (endoglin) at the mRNA level in a series of breast cancers and to correlate the results obtained with all available clinical and biological features of the tumors. MATERIALS AND METHODS: Fresh tumor tissue from 103 previously untreated breast cancer patients was studied for VEGF and CD105 (endoglin) expression. In addition, the following parameters were determined in all tumors: DNA ploidy by means of flow cytometry; hormone receptor (ER & PR), Ki67, c-erb-B2 and p53 expression by means of immunohistochemistry; and h-MAM (mammaglobin) expression by means of RT-PCR. Classical prognostic parameters of the tumors, such as histological and nuclear grade or axillary lymph node invasion, were also included into the statistical analysis. RESULTS: VEGF mRNA expression levels above the 25th percentile were significantly (p<0.05) associated with high proliferation (Ki67>10%) and aneuploidy of the tumors and inversely with estrogen receptor expression (p<0.01). CD105 (endoglin) mRNA expression levels above the 25th percentile only correlated significantly with nuclear grade 3 (p<0.05). The expression of both genes did not correlate with each other. CONCLUSION: VEGF mRNA expression levels seem to be directly associated with VEGF functions at the protein level, whereas this seems not to be the case for CD105 (endoglin) mRNA levels.

Nup88 mRNA overexpression is associated with high aggressiveness of breast cancer.

The nuclear pore complex protein Nup88 is overexpressed in tumor cells. Immunohistochemical studies have shown that this overexpression is linked to higher aggressiveness of colorectal carcinoma and to enhanced metastatic potential of melanoma cells. However, the antibodies so far developed against Nup88 have the drawback of recognizing a number of other, up to now unspecified antigens besides Nup88. For this reason, we devised the present study on Nup88 expression at the mRNA level. RNA was extracted from fresh tumor tissue corresponding to 122 breast cancer patients. Nup88 mRNA expression was measured by means of differential RT-PCR, standardizing against a constitutive internal control gene (beta-actin). The results were dichotomized into "high" and "low" expression levels, using the median value as cut-off. High Nup88 mRNA expression levels correlated significantly with ductal and tubular histology (p = 0.012), histologic and nuclear grade 3 of tumors (p < 0.001), absence of hormone receptor expression (p < 0.001), expression of the c-erb-B2 oncogene (p < 0.001), expression of mutant p53 protein (p < 0.001), high proliferation (defined by Ki67 labeling index >20%, p < 0.001), DNA aneuploidy (p < 0.001) as well as the most important ominous clinical prognostic factor, axillary node invasion (p < 0.001). We also found an inverse correlation (p < 0.001) with expression of the H-MAM (mammaglobin) gene, a marker of low biologic and clinical aggressiveness of breast cancer. All of these factors, without exception, define a highly aggressive tumor phenotype. These findings appear to be specific to Nup88 and not to nuclear pore proteins in general. Indeed, analysis of Nup107 (which is a limiting component of the nuclear pore complex) under the same conditions in the same tumors did not yield comparable results.