Agreement between specially trained and accredited optometrists and glaucoma specialist consultant ophthalmologists in their management of glaucoma patients.

AIMS: Optometrists are becoming increasingly involved in the co-management of glaucoma patients as the burden on the Hospital Eye Service continues to escalate. The aim of this study was to assess the agreement between specially trained optometrists and glaucoma-specialist consultant ophthalmologists in their management of glaucoma patients. METHODS: Four optometrists examined 23-25 patients each and the clinical findings, up to the point of dilation, were documented in the hospital records. The optometrist, and one of two consultant ophthalmologists, then independently examined and documented the optic-disc appearance before recording their decisions regarding the stability and management of the patient on a specially designed proforma. Percentage agreement was calculated together with kappa or weighted kappa statistics, where appropriate. RESULTS: Agreement between consultants and optometrists in evaluating glaucoma stability was 68.5% (kappa (κ)=0.42-0.50) for visual fields, 64.5% (weighted κ=0.17-0.31) for optic discs, and 84.5% (weighted κ=0.55-0.60) for intraocular pressures. Agreement regarding medical management was 96.5% (κ=0.73-0.81) and for other glaucoma management decisions, including timing of follow-up, referral to a consultant ophthalmologist, and discharge, was 72% (weighted κ=0.65). This agreement increased to 90% following a retrospective independent then consensus review between the two consultants and when qualified agreements were included. Of the 47 glaucoma and non-glaucoma queries generated during the study, 42 resulted in a change of management. CONCLUSION: Confirming the ability of optometrists to make appropriate decisions regarding the stability and management of glaucoma patients is essential if their involvement is to continue to develop to meet the demand of an aging population.

Epigenetic regulation of CD133 and tumorigenicity of CD133+ ovarian cancer cells.

The cancer stem cell hypothesis posits that malignant growth arises from a rare population of progenitor cells within a tumor that provide it with unlimited regenerative capacity. Such cells also possess increased resistance to chemotherapeutic agents. Resurgence of chemoresistant disease after primary therapy typifies epithelial ovarian cancer and may be attributable to residual cancer stem cells, or cancer-initiating cells, that survive initial treatment. As the cell surface marker CD133 identifies cancer-initiating cells in a number of other malignancies, we sought to determine the potential role of CD133+ cells in epithelial ovarian cancer. We detected CD133 on ovarian cancer cell lines, in primary cancers and on purified epithelial cells from ascitic fluid of ovarian cancer patients. We found CD133+ ovarian cancer cells generate both CD133+ and CD133- daughter cells, whereas CD133- cells divide symmetrically. CD133+ cells exhibit enhanced resistance to platinum-based therapy, drugs commonly used as first-line agents for the treatment of ovarian cancer. Sorted CD133+ ovarian cancer cells also form more aggressive tumor xenografts at a lower inoculum than their CD133- progeny. Epigenetic changes may be integral to the behavior of cancer progenitor cells and their progeny. In this regard, we found that CD133 transcription is controlled by both histone modifications and promoter methylation. Sorted CD133- ovarian cancer cells treated with DNA methyltransferase and histone deacetylase inhibitors show a synergistic increase in cell surface CD133 expression. Moreover, DNA methylation at the ovarian tissue active P2 promoter is inversely correlated with CD133 transcription. We also found that promoter methylation increases in CD133- progeny of CD133+ cells, with CD133+ cells retaining a less methylated or unmethylated state. Taken together, our results show that CD133 expression in ovarian cancer is directly regulated by epigenetic modifications and support the idea that CD133 demarcates an ovarian cancer-initiating cell population. The activity of these cells may be epigenetically detected and such cells might serve as pertinent chemotherapeutic targets for reducing disease recurrence.

Identification of genes associated with ovarian cancer metastasis using microarray expression analysis.

Although the transition from early- to advanced-stage ovarian cancer is a critical determinant of survival, little is known about the molecular underpinnings of ovarian metastasis. We hypothesize that microarray analysis of global gene expression patterns in primary ovarian cancer and metastatic omental implants can identify genes that underlie the metastatic process in epithelial ovarian cancer. We utilized Affymetrix U95Av2 microarrays to characterize the molecular alterations that underlie omental metastasis from 47 epithelial ovarian cancer samples collected from multiple sites in 20 patients undergoing primary surgical cytoreduction for advanced-stage (IIIC/IV) serous ovarian cancer. Fifty-six genes demonstrated differential expression between ovarian and omental samples (P < 0.01), and twenty of these 56 differentially expressed genes have previously been implicated in metastasis, cell motility, or cytoskeletal function. Ten of the 56 genes are involved in p53 gene pathways. A Bayesian statistical tree analysis was used to identify a 27-gene expression pattern that could accurately predict the site of tumor (ovary versus omentum). This predictive model was evaluated using an external data set. Nine of the 27 predictive genes have previously been shown to be involved in oncogenesis and/or metastasis, and 10/27 genes have been implicated in p53 pathways. Microarray findings were validated by real-time quantitative PCR. We conclude that gene expression patterns that distinguish omental metastasis from primary epithelial ovarian cancer can be identified and that many of the genes have functions that are biologically consistent with a role in oncogenesis, metastasis, and p53 gene networks.

High expression of insulin-like growth factor binding protein-2 messenger RNA in epithelial ovarian cancers produces elevated preoperative serum levels.

The molecular etiology of epithelial ovarian cancer remains unclear. Using microarray expression analysis, we recently reported that expression of the insulin-like growth factor binding protein-2 (IGFBP-2) gene is elevated in advanced epithelial ovarian cancers. The aim of this study was to further delineate the role of IGFBP-2 in the pathoetiology of epithelial ovarian cancer and determine if elevated ovarian cancer IGFBP-2 gene expression is reflected in serum. Relative IGFBP-2 expression was measured using quantitative real-time polymerase chain reaction in 113 epithelial ovarian cancers and 6 normal ovarian surface epithelial samples. Preoperative serum IGFBP-2 levels were measured by radioimmunoassay in 84 women (42 ovarian cancers, 26 benign gynecological conditions, and 10 healthy female controls). Ovarian cancers demonstrated 38-fold higher mean IGFBP-2 expression than normal ovarian epithelium (P < 0.01). Serum IGFBP-2 levels were elevated in women with early- and advanced-stage ovarian cancer compared to controls and patients with benign gynecological conditions (P = 0.05 and P < 0.01, respectively). Epithelial ovarian cancers express high levels of IGFBP-2 relative to normal ovarian epithelium, and this is associated with elevated serum IGFBP-2 levels compared to both normal controls and patients with benign gynecological disease. Our findings provide further support that the insulin-like growth factor pathway plays a significant role in epithelial ovarian cancer pathogenesis. Further, IGFBP-2 may represent an additional serum biomarker with utility in detection and monitoring of epithelial ovarian cancer.

Dhh1 regulates the G1/S-checkpoint following DNA damage or BRCA1 expression in yeast.

BACKGROUND: Heterologous expression of the tumor suppressor BRCA1 in the yeast Saccharomyces cerevisiae is lethal. To identify potential new BRCA1-interacting gene targets, we characterized highly conserved ionizing radiation (IR) sensitive gene deletions that suppress BRCA1-induced lethality in yeast. MATERIALS AND METHODS: Previously, we exposed an isogenic collection of yeast strains individually deleted for 4746 nonessential genes to IR and identified 199 radiation sensitive deletion strains. A subset (n = 130) of these were screened for those that suppressed the G1 arrest and lethality observed following galactose-induced expression from a GAL::BRCA1 plasmid in wild type yeast. RESULTS: We found that deletions of two core components of the highly conserved CCR4-NOT transcription complex (CCR4 or DHH1) rescued BRCA1-induced G1 arrest and lethality in yeast. This was not because of down regulation of the GAL promoter since both deletion strains produce large amounts of BRCA1 that is rapidly degraded. In addition, heterologous expression of BRCA1 results in increased transcription of the DNA damage-inducible reporter construct DIN::LacZ. Reduced viability following IR and nitrogen starvation was observed among strains deleted for CCR4 or DHH1 because of a defect in G1 to S phase checkpoint transition. Lethality following nitrogen starvation and IR was partially rescued in dhh1Delta strains by expressing the human ortholog of DHH1 (DDX6) which has been identified as a breakpoint oncogene.T CONCLUSIONS: hese results suggest that BRCA1 may promote genomic stability in human cells by interacting with the highly conserved ortholog of DHH1 (DDX6) to properly activate G1/S checkpoint arrest following DNA damage.

Loss of expression of the p16 tumor suppressor gene is more frequent in advanced ovarian cancers lacking p53 mutations.

OBJECTIVE: The aim of this study was to test the hypothesis that p53 mutations are less frequent in ovarian cancers with alterations in other genes that regulate G1 progression. METHODS: Expression of G1 stimulatory (cyclins D1 and E, cdk4, Ki67) and inhibitory (p16, Rb, p27, p14) genes was analyzed using Western blots in 84 primary ovarian cancers and seven cell lines of known p53 mutation status. Expression of p16 and Rb also was determined using immunohistochemistry and the p16 gene was examined for homozygous deletions and mutations. RESULTS: Loss of p16 protein was more frequent in ovarian cancers with wild-type p53. All four cell lines with wild-type p53 had lost p16 compared to only one of three with mutant p53 genes. p16 expression was absent in 34% (28/82) of primary ovarian cancers, and this was significantly more common in cases with wild-type p53 (14/28, 50%) compared to those with p53 mutations (14/54, 26%, P = 0.03). Homozygous deletion of the p16 gene was found in cell lines lacking p16, but not in any primary cancers. p16 loss was more common in serous (21/52, 40%) than nonserous cancers (4/23, 17%, P = 0.07). Cases that expressed p16 were more likely to express high levels of Rb (47/55, 85%) than p16-negative cases (12/28, 43%, P < 0.001). Loss of Rb occurred in 5/30 (17%) ovarian cancers lacking p53 mutations compared to 5/54 (9%) cases with p53 mutations (P = 0.48). Expression of G1 stimulatory proteins (cyclins D1 and E, cdk4, Ki67) did not correlate with p53 mutation status. CONCLUSIONS: Loss of expression of the p16 tumor suppressor occurs more often in ovarian cancers lacking p53 mutations. These data are consistent with the paradigm that inactivation of p53 is less of a requisite event in ovarian carcinogenesis when another G1 regulatory gene such as p16 already has been inactivated.

Predicting the clinical status of human breast cancer by using gene expression profiles.

Prognostic and predictive factors are indispensable tools in the treatment of patients with neoplastic disease. For the most part, such factors rely on a few specific cell surface, histological, or gross pathologic features. Gene expression assays have the potential to supplement what were previously a few distinct features with many thousands of features. We have developed Bayesian regression models that provide predictive capability based on gene expression data derived from DNA microarray analysis of a series of primary breast cancer samples. These patterns have the capacity to discriminate breast tumors on the basis of estrogen receptor status and also on the categorized lymph node status. Importantly, we assess the utility and validity of such models in predicting the status of tumors in crossvalidation determinations. The practical value of such approaches relies on the ability not only to assess relative probabilities of clinical outcomes for future samples but also to provide an honest assessment of the uncertainties associated with such predictive classifications on the basis of the selection of gene subsets for each validation analysis. This latter point is of critical importance in the ability to apply these methodologies to clinical assessment of tumor phenotype.

HIN-1, a putative cytokine highly expressed in normal but not cancerous mammary epithelial cells.

To identify molecular alterations implicated in the initiating steps of breast tumorogenesis, we compared the gene expression profiles of normal and ductal carcinoma in situ (DCIS) mammary epithelial cells by using serial analysis of gene expression (SAGE). Through the pair-wise comparison of normal and DCIS SAGE libraries, we identified several differentially expressed genes. Here, we report the characterization of one of these genes, HIN-1 (high in normal-1). HIN-1 expression is significantly down regulated in 94% of human breast carcinomas and in 95% of preinvasive lesions, such as ductal and lobular carcinoma in situ. This decrease in HIN-1 expression is accompanied by hypermethylation of its promoter in the majority of breast cancer cell lines (>90%) and primary tumors (74%). HIN-1 is a putative cytokine with no significant homology to known proteins. Reintroduction of HIN-1 into breast cancer cells inhibits cell growth. These results indicate that HIN-1 is a candidate tumor suppressor gene that is inactivated at high frequency in the earliest stages of breast tumorogenesis.

A SAGE (serial analysis of gene expression) view of breast tumor progression.

To identify molecular alterations involved in the initiation and progression of breast carcinomas, we analyzed the global gene expression profiles of normal mammary epithelial cells and in situ, invasive, and metastatic breast carcinomas using serial analysis of gene expression (SAGE). We identified sets of genes expressed only or most abundantly in a specific stage of breast tumorigenesis or in a certain subtype of tumors through the pair-wise comparison and by hierarchical clustering analysis of these eight SAGE libraries (two/stage). On the basis of these comparisons, we made the following observations: Normal mammary epithelial cells showed the most distinct and least variable gene expression profiles. Many of the genes highly expressed in normal mammary epithelium and lost in carcinomas encoded secreted proteins, cytokines, and chemokines, implicating abnormal paracrine and autocrine signaling in the initiation of breast tumorigenesis. Very few genes were universally up-regulated in all tumors regardless of their stage and histological grade, indicating a high degree of diversity at the molecular level that likely reflects the clinical heterogeneity characteristic of breast carcinomas. Tumors of different histology type and stage had very distinct gene expression patterns. No genes seemed to be specific for metastatic or for in situ carcinomas. We found that the most dramatic and consistent phenotypic change occurred at the normal-to-in situ carcinoma transition. This observation, combined with the fact that many of the genes involved encode secreted, cell-nonautonomous factors, implies that the normal epithelium-to-in situ carcinoma transition may be the most promising target for cancer prevention and treatment.

Prognostic significance of the number of lymph nodes examined in patients with lymph node-negative breast carcinoma.

BACKGROUND: A recent report suggested that the number of lymph nodes examined was a strong predictor of survival in patients with lymph node-negative breast carcinoma. Among women who had >or= 20 lymph nodes examined, the risk of dying from breast carcinoma within 5 years was increased nearly 4-fold compared with women who had fewer lymph nodes examined. Because these findings were based on a relatively small cohort of patients, corroborative studies with larger patient populations were needed. METHODS: The authors studied the relation between the number of lymph nodes examined and breast carcinoma survival among 911 women with lymph node-negative breast carcinoma with a median length of follow-up of 84 months. The association between other prognostic indicators and survival and the number of lymph nodes examined also was investigated. RESULTS: The number of lymph nodes examined was not found to be associated with either 5-year or long-term survival. The proportion of women dying from breast carcinoma was the same (8%) in both groups (those patients with >or= 20 lymph nodes examined vs. those in whom < 20 lymph nodes were examined) and the hazard ratio was 0.98 (95% confidence interval, 0.58-1.64). CONCLUSIONS: In this larger study population, the authors failed to confirm the findings of an earlier investigation in which having a larger number of lymph nodes examined was associated with poorer survival. This finding suggests that it is unlikely the number of lymph nodes examined is an important prognostic indicator in patients with lymph node-negative breast carcinoma.

Relationship between expression of coactivators and corepressors of hormone receptors and resistance of ovarian cancers to growth regulation by steroid hormones.

OBJECTIVE: To determine whether aberrant expression of hormone receptor corepressors or coactivators or defects in estrogen receptor-mediated transcription might underlie resistance of ovarian cancers to hormonal therapy. METHODS: Northern analysis, Western analysis, and polymerase chain reaction were used to examine expression of estrogen receptor (ER), progesterone receptor (PR), the nuclear receptor corepressors N-CoR and SMRT, and the steroid receptor coactivator BRG-1 in ovarian cancer cell lines and primary cancers. The effect of BRG-1 transfection on ER-mediated transcription was examined. We also determined the effect of estrogen and the pure estrogen antagonist ICI 182,780 on cell cycle profile and expression of ER. Finally, we examined the ability of estrogen to upregulate expression of known estrogen-responsive genes. RESULTS: Among primary ovarian cancers, 18 of 52 (35%) expressed N-CoR, and 37 of 52 (71%) expressed SMRT, but there was no correlation between expression of corepressors and hormone receptor status. All of the primary ovarian cancers and cell lines expressed BRG-1. Estrogen stimulation of two cell lines expressing ER (SKOV3, OVCA 432) elicited low levels of ER-mediated transcription that was not enhanced by BRG-1 transfection. ICI 182,780 did not induce cell cycle arrest in these cell lines, but there was evidence of downregulation of ER, indicating a ligand-receptor interaction. However, estrogen did not elicit increased transcription of estrogen-responsive genes (PR, myc, fos, pS2). CONCLUSION: Inappropriate expression of the nuclear corepressors N-CoR and SMRT or the coactivator BRG-1 does not underlie the resistance of ovarian cancers to hormonal therapy. Further studies are needed to elucidate the mechanisms underlying the inability of ovarian cancers to undergo ER-mediated transcription if we hope to understand their resistance to hormonal therapy.

High frequency of hypermethylation at the 14-3-3 sigma locus leads to gene silencing in breast cancer.

Expression of 14-3-3 final sigma (final sigma) is induced in response to DNA damage, and causes cells to arrest in G(2). By SAGE (serial analysis of gene expression) analysis, we identified final sigma as a gene whose expression is 7-fold lower in breast carcinoma cells than in normal breast epithelium. We verified this finding by Northern blot analysis. Remarkably, final sigma mRNA was undetectable in 45 of 48 primary breast carcinomas. Genetic alterations at final sigma such as loss of heterozygosity were rare (1/20 informative cases), and no mutations were detected (0/34). On the other hand, hypermethylation of CpG islands in the final sigma gene was detected in 91% (75/82) of breast tumors and was associated with lack of gene expression. Hypermethylation of final sigma is functionally important, because treatment of final sigma-non-expressing breast cancer cell lines with the drug 5-aza-2'-deoxycytidine resulted in demethylation of the gene and synthesis of final sigma mRNA. Breast cancer cells lacking final sigma expression showed increased number of chromosomal breaks and gaps when exposed to gamma-irradiation. Therefore, it is possible that loss of final sigma expression contributes to malignant transformation by impairing the G(2) cell cycle checkpoint function, thus allowing an accumulation of genetic defects. Hypermethylation and loss of final sigma expression are the most consistent molecular alterations in breast cancer identified so far.

Inhibition of mitogen-activated protein kinase and phosphatidylinositol 3-kinase activity in MCF-7 cells prevents estrogen-induced mitogenesis.

Estrogen acts to promote DNA synthesis in the MCF-7 human breast cancer cell line via its interaction with high levels of estrogen receptor. The primary mode of estrogen action has been considered to be through transcriptional activation of genes containing estrogen response elements, including the immediate early genes c-myc and fos. Recent reports have indicated that estrogen, acting through the estrogen receptor, is capable of inducing the mitogen-activated protein kinase (MAPK) cytoplasmic signaling cascade. In this study, specific small molecule inhibitors of MAPK and phosphatidylinositol 3-kinase activity were used to determine the influence of these cascades on estrogen-mediated mitogenesis. Phosphatidylinositol 3-kinase inhibitors, LY294002 and wortmannin, as well as inhibitors of MAPK kinase-1, PD098059 and U0126, decreased the fraction of cells entering DNA synthesis after treatment with 17beta-estradiol. These compounds did not inhibit expression of myc or fos. However, the drugs did prevent the accumulation of cyclin D1 and hyperphosphorylated retinoblastoma protein, indicating that the block occurred at, or prior to, this point in the cell cycle. Although these compounds were effective in preventing estrogen-mediated mitogenesis, the downstream kinases extracellular signal-regulated kinase 1, extracellular signal-regulated kinase 2, and protein kinase B were not activated over basal levels by estrogen treatment. These studies suggest that estrogen initiates mitogenesis by inducing the transcription of immediate early genes, but cytoplasmic signaling pathways play an important role in the control of subsequent events in the cell cycle.

Phenol sulfotransferases: hormonal regulation, polymorphism, and age of onset of breast cancer.

In recent years, significant effort has been made to identify genes that influence breast cancer risk. Because the high-penetrance breast cancer susceptibility genes BRCA1 and 2 play a role only in a small fraction of breast cancer cases, understanding the genetic risk of the majority of breast cancers will require the identification and analysis of several lower penetrance genes. The estrogen-signaling pathway plays a crucial role in the pathophysiology of breast cancer; therefore, polymorphism in genes involved in this pathway is likely to influence breast cancer risk. Our detailed analysis of gene expression profiles of estrogen- and 4-OH-tamoxifen-treated ZR75-1 breast cancer cells identified members of the sulfotransferase 1A (SULT1A) phenol sulfotransferase family as downstream targets of tamoxifen. On the basis of the induction of SULT1A by 4-OH-tamoxifen and the known inherited variability in SULT1A enzymatic activity, we hypothesized that polymorphism in sulfotransferase genes might influence the risk of breast cancer. Using an RFLP that distinguishes an arginine to histidine change in exon 7 of the SULT1A1 gene, we characterized SULT1A1 genotypes in relation to breast cancer risk. An analysis of 444 breast cancer patients and 227 controls revealed no effect of SULT1A1 genotype on the risk of breast cancer (P = 0.69); however, it did appear to influence the age of onset among early-onset affected patients (P = 0.04). Moreover, individuals with the higher activity SULT1A1*1 allele were more likely to have other tumors in addition to breast cancer (P = 0.004; odds ratio, 3.02; 95% confidence interval, 1.32, 8.09). The large number of environmental mutagens and carcinogens activated by sulfotransferases and the high frequency of the SULT1A1*1 allele in human populations warrants additional studies to address the role of SULT genes in human cancer.

Estrogen-induced mitogenesis of MCF-7 cells does not require the induction of mitogen-activated protein kinase activity.

Estrogen mediates the transcription of responsive genes via its interaction with the estrogen receptor (ER). This ligand-dependent transcriptional activity has been the mechanistic basis for understanding estrogen-induced proliferation. However, recent reports suggest that estrogen stimulation results in activation of the mitogen-activated protein kinase (MAPK) cascade in an ER-dependent manner suggesting that mitogenesis may be mediated through this cytoplasmic signaling cascade. In this study, we demonstrate that estrogen stimulation of MCF-7 cells does not activate MAPK regardless of hormone concentration, serum concentration, or cell density. We also excluded the activation of MAPK through autocrine effects after estrogen treatment. Finally, concentrations required for estrogen-induced mitogenesis and estrogen-mediated transcription were shown to be the same. These results support transcriptional activation as the primary mechanism for estrogen-mediated mitogenesis.

Isolation and initial characterization of the BRCA2 promoter.

The hereditary breast cancer susceptibility gene, BRCA2, is considered to be a tumor suppressor gene that may be involved in the cellular response to DNA damage. The transcript for this gene is cell cycle regulated with mRNA levels reaching a peak just before the onset of DNA synthesis. In order to define the mechanisms by which BRCA2 is transcriptionally regulated, we have begun to study upstream regulatory sequences. In this report, we define a minimal promoter region that has strong activity in human breast epithelial cells. Deletions of this sequence narrowed the strong basal activity to a region extending from -66 to +129 with respect to the BRCA2 transcriptional start site. This sequence demonstrated cell cycle regulated activity with kinetics similar to the endogenous transcript. Examination of the sequence revealed several consensus binding sites for transcription factors including an E-box, E2F and Ets recognition motifs. Electrohoretic mobility shift assays revealed specific protein binding to two sequences upstream of the start site; the palindromic E-box and an Ets/E2F site. Site-directed mutagenesis of either of these sites reduced both the basal activity in log phase cells and the cell cycle regulated activity of the promoter. Mutational inactivation of both sites within the same construct effectively eliminated promoter activity. Antibodies to candidate transcription factors used in super shift experiments revealed specific interactions between the BRCA2 promoter and the basic region/helix - loop - helix containing USF-1 and 2 proteins and Elf-1, an Ets domain protein. Binding of these factors depended upon the presence of intact recognition sequences. The USF factors were shown to bind predominantly as a heterodimeric complex of USF-1 and 2 while Elf-1 bound the promoter when it was not occupied by USF. Co-transfection studies with USF proteins and the varicella zoster IE62 protein provide evidence for the involvement of endogenous and exogenous USF in the activation of the BRCA2 promoter. We propose that interactions between USF-1, USF-2 and Elf-1 play an important role in the transcriptional regulation of the BRCA2 gene.

Bcl10 is not a target for frequent mutation in human carcinomas.

The recently described Bcl10 gene has been suggested to be a major target gene for inactivation in a variety of human cancers. In order to further evaluate the role of this gene in human adult malignancies, we have analysed a series of carcinomas for mutations in the Bcl10 gene. We have screened a panel of 174 carcinoma samples in total, comprised of 47 breast, 36 epithelial ovarian, 36 endometrial, 12 cervical, 23 colorectal and 20 head/neck carcinomas, all unselected for grade or stage. This panel reflects, in part, tumours reported to have involvement of the 1p22 region of chromosome 1, the region harbouring the Bcl10 gene. No deleterious mutations were detected in any of the samples analysed, strongly suggesting that Bcl10 is not a common target for inactivation in adult malignancies and that BCL10 is not the gene targeted for frequent inactivation at 1p22.

Co-expression of p53 by epithelial and stromal elements in carcinosarcoma of the female genital tract: an immunohistochemical study of 19 cases.

Carcinosarcoma is an aggressive neoplasm of the female genital tract, which comprises 1-2% of malignancies of the uterine corpus. Because of the broad range of differentiation exhibited by these tumors, the precise nature of the relationship between epithelial and stromal components in this unique tumor remain unclear. Previous studies have demonstrated that mutation and consequent overexpression of the tumor suppressor gene p53 occurs frequently in carcinosarcoma and is conserved from primary to metastastic sites. We examined p53 accumulation in formalin-fixed, paraffin-embedded archival sections in 19 cases previously shown to have mutations in the p53 gene and performed semi-quantitative analysis of the intensity of staining and relative density of positive cells and stromal and glandular elements. There was a high level of concordance of immunohistochemical staining for the p53 oncoprotein between glandular and stromal elements. These results further suggest a clonal origin for the diverse elements of carcinosarcoma.

Managing hereditary ovarian cancer risk.

Ovarian cancer is the fourth leading cause of cancer deaths in American women. About 10% of cases are thought to have a hereditary basis, and family history is the strongest known risk factor. In the past, prophylactic oophorectomy has been advocated for women with two or more affected first-degree relatives. More recently, with the identification of the genes responsible for most hereditary ovarian cancers (BRCA1, BRCA2), oophorectomy can now be offered specifically to women who are mutation carriers. Conversely, noncarriers in these families can be reassured that their risk of ovarian cancer is not increased. The value of oophorectomy in mutation carriers has not yet been proven, however, and concern exists that the benefit may be less than intuitively expected. First, although the lifetime risk of ovarian cancer initially was reported to be as high as 60%, more recent studies have suggested risks in the range of 15 to 30%. A better understanding of the factors that underlie variable penetrance in mutation carriers is needed to augment our ability to counsel individual women. In addition, peritoneal papillary serous carcinoma indistinguishable from ovarian cancer occurs in some women after oophorectomy. Studies that better define the frequency with which this occurs are needed to establish the magnitude of the protective effect conferred by prophylactic oophorectomy. In view of the uncertainty regarding the efficacy of prophylactic oophorectomy, chemopreventive and early detection approaches also deserve consideration as strategies for decreasing ovarian cancer mortality in women who carry mutations in ovarian cancer susceptibility genes.

Frequency of germline and somatic BRCA1 mutations in ovarian cancer.

Germline mutations in the BRCA1 tumor suppressor gene are thought to be the most common cause of hereditary ovarian cancer. The aim of this study was to explore further the role of BRCA1 alterations in the development of ovarian cancers. We sought to determine whether somatic BRCA1 mutations are ever present in ovarian cancers and whether mutation is always accompanied by loss of the wild-type allele. The entire coding region and intronic splice sites of BRCA1 were sequenced using genomic DNA samples from 103 unselected ovarian cancers. Thirteen clearly deleterious BRCA1 mutations and two variants of uncertain significance were found. Blood DNA was available in all but two cases and demonstrated that 4 of 13 mutations and both variants of uncertain significance were germline alterations, whereas in seven cases the mutation was a somatic change present only in the cancer. Using four microsatellite markers, loss of heterozygosity at the BRCA1 locus was found in all 15 ovarian cancers with BRCA1 sequence alterations, compared with only 58% of ovarian cancers that did not have BRCA1 mutations. BRCA1-associated ovarian cancers were characterized by serous histology and moderate histological grade. These data confirm prior reports suggesting that germline mutations in BRCA1 are present in about 5% of women with ovarian cancer. In addition, somatic mutations in BRCA1 occur in the development of some sporadic cases. The finding that both germline and somatic BRCA1 mutations are accompanied by loss of heterozygosity, suggests that loss of this tumor suppressor gene is a critical event in the development of these cancers.