Targeting Src in endometriosis-associated ovarian cancer.

The SRC proto-oncogene is commonly overexpressed or activated during cancer development. Src family kinase inhibitors are approved for the treatment of certain leukemias, and are in clinical trials for the treatment of solid tumors. Src signaling is activated in endometriosis, a precursor of clear cell and endometrioid subtypes of epithelial ovarian cancers (OCs). We examined the expression of phosphorylated Src (Src-pY416) in 381 primary OC tissues. Thirty-six percent of OCs expressed Src-pY416. Src-pY416 expression was most common in endometriosis-associated OCs (EAOCs) (P=0.011), particularly in clear cell OCs where 58.5% of cases expressed Src-pY416. Src-pY416 expression was associated with shorter overall survival (log rank P=0.002). In vitro inhibition of Src signaling using 4-amino-5-(4-chlorophenyl)-7-(dimethylethyl)pyrazolo[3,4-d]pyrimidine (PP2) resulted in reduced anchorage-independent and -dependent growth, and in three-dimensional cell culture models PP2 disrupted aggregate formation in Src-pY416-positive but not in Src-pY416-negative cell lines. These data suggest that targeting active Src signaling could be a novel therapeutic opportunity for EAOCs, and support the further pre-clinical investigation of Src family kinase inhibitors for treating OCs expressing Src-pY416.

The frequency of human papilloma virus types 16, 18 in upper genital tract of women at high risk of developing ovarian cancer.

AIM: To investigate the incidence of human papilloma virus (HPV) types 16, 18 in upper genital tract of women considered at a high risk (HR) of developing epithelial ovarian cancer (EOC). METHODS: HPV 16 and 18 E6 ORF specific semiquantitative PCR was used to screen the incidence of HPV in 20 women at HR of developing EOC and 10 women with no ovarian disease (control). RESULTS: The HR subset of fallopian tubes and ovarian tissues showed greater positivity for HPV E6 ORF (40%) as compared to control (10%) tissues. Of all the samples, two (10%) were positive for HPV 16, two (10%) were positive for HPV 18, and four (20%) showed positivity for mixed HPV 16/18 infection. The presence of HPV E6 ORF was found both in the fallopian tubes and ovarian DNA from 6 (30%) patients. In two cases (10%) we detected HPV ORF only in the fallopian tube derived genomic DNA. CONCLUSION: It has been shown the presence of HPV in the upper genital tract in women at HR of developing EOC in close proximity of HPV susceptible tissue cervix.

Pair-Box (PAX8) protein-positive expression is associated with poor disease outcome in women with endometrial cancer.

BACKGROUND: The Pax8 transcription factor genes have a role in cell differentiation and cell growth, and silencing of Pax8 in cell cultures results in cell death. The aims of this study were to determine the expression and correlation of Pax8 protein with several clinicopathological variables in patients with endometrial cancer. METHODS: The following clinical parameters from 229 patients were used for correlation with Pax8 expression; age, histological subtype, myometrial depth of invasion, lymphovascular invasion (LVI), the International Federation of Gynecology and Obstetrics grade, lymph nodes status, and disease status. RESULTS: A positive association of Pax8(+) expression was found with high tumour grade (P=0.002), LVI+(P=0.0186), and type II tumour subtype (P<0.0001) in univariate analysis. Survival analysis showed an association of Pax8 and 5-year overall survival probability (P=0.01486), 80.04% for patients with Pax8(-) and 55.59% for patients with Pax8(+). There was also an association of Pax8 and 5-year disease-free survival probability (P=0.02028), 72.12% for patients with Pax8(-) vs 49.88% for patients with Pax8(+). Finally, an association of Pax8(+) and shorter recurrence-free survival was also found (P=0.00203), with 74.36% for Pax8(-) and 52.11% for Pax8(+). CONCLUSION: Overexpression of Pax8 protein by endometrial cancer is associated with poor disease outcomes. Inhibition of Pax8 may be a very attractive targeted therapy for selective patients.

Laparoscopic management of adnexal masses the opportunities and the risks.

Suspected ovarian neoplasm is a common clinical problem affecting women of all ages. Although the majority of adnexal masses are benign, the primary goal of diagnostic evaluation is the exclusion of malignancy. It has been estimated that approximately 5-10% of women in the United States will undergo a surgical procedure for a suspected ovarian neoplasm during their lifetime. Despite the magnitude of the problem, there is still considerable disagreement regarding the optimal surgical management of these lesions. Traditional management has relied on laparotomy to avoid undertreatment of a potentially malignant process. Advances in detection, diagnosis, and minimally invasive surgical techniques make it necessary now to review this practice in an effort to avoid unnecessary morbidity among patients. Here, we review the literature on the laparosopic approach to the treatment of the adnexal mass without sacrificing the principles of oncologic surgery. We highlight potentials of minimally invasive surgery and address the risks associated with the laparoscopic approach.

[Residual choledocholithiasis in elective cholecystectomy and choledocholithotomy]. / Rezidualna holedoholitijaza kod elektivne holecistektomije i holedoholitotomije.

Cholecystectomy is one of the most frequent abdominal operation. Common bile duct stone may be found in 6-15% of elective cholecystectomics. One or more stones may be left behind within common bile duct. In a series of 106 consecutive cholecystectomics performed in the First surgical clinic in Belgrade over six months period (from 07.08.1997.-31.12.1997.) operative cholangiography was performed routinely in every single case. Unexpected common bile duct stone was found in 13 patients (12.26%). After removal of stones choledochoscopy was performed in 7 out of 13 patients with common bile duct stone (53.8%). An operative T-tube cholangiography was performed in all 13 cases before closure of the abdomen. Postoperatively, usually from 8th to 10th day a secondary T-tube cholangiography in the X-ray department was performed in all 13 cases. Retained stone was found in 2 patients, both in group of patients in whom choledochoscopy had not been carried out. The retained stone was successfully removed endoscopically in one case but the second patient had to be reoperated as endoscopy was unsuccessful. We conclude that choledochoscopy reduces the risk of retained common bile duct stone and it should be practiced whenever possible.

No BRCA1 germline mutation in a family with uterine papillary serous carcinoma: a case report.

The purpose of the study was to examine BRCA1 germline mutation and its relationship to BRCA1 expression in two patients, a mother and a daughter, both diagnosed with uterine papillary serous carcinoma (UPSC). DNA was screened for BRCA1 and BRCA2 germline mutations common in the Jewish population (185delAG, 5382insC, and 6174delT) by PCR-based assay and with a protein truncation test (PTT) to detect mutation in exon 11 of BRCA1 and exons 10 and 11 of BRCA2. BRCA1 expression in fixed tumor tissues was assessed by immunocytochemistry (IHC). No germline mutation in either BRCAI or BRCA2 gene was found in the two patients. Both samples showed reduced levels of BRCAI expression. Taken together, these results suggest that undetected or unscreened for germline mutation may be associated with occurrence of this rare tumor type in two members of the same family. Alternatively, an epigenetic mechanism such as BRCA1 promoter hypermethylation may be responsible for reduced expression of BRCA1 in the absence of DNA mutations.

Comparative genomic hybridization of fine needle aspirates from breast carcinomas.

Detailed knowledge of chromosomal aberrations in a specific tumor may facilitate the development of individually tailored chemotherapy, hormone or gene therapy. Unfortunately, karyotype analysis requires living cells and is complicated by the low number of good metaphase spreads obtained. Comparative genomic hybridization (CGH), however, is capable of detecting and mapping genome-wide amplifications and deletions using an equimolar mixture of normal and tumor cell DNA. We show here that even the few cells from a fine needle aspirate of a tumor are sufficient for a direct CGH assay, independent of DNA amplification. Ten primary breast cancers were analyzed by CGH. A fresh frozen fine needle aspirate and a formalin-fixed and paraffin-embedded section were used for each tumor. Metaphases from each CGH reaction were imaged, and a sum ratio profile was determined for every chromosome. The ratio profiles of DNA isolated from the 2 material sources were then compared. Fine needle aspirates and the paraffin-embedded material of a single tumor yielded the same fluorescence ratio profiles, albeit with slightly different confidence intervals. Different tumors showed a variety of aberrations. The most frequently observed changes were 1q+, 8q+, 14q-, 16p+, 16q-, 17p-, 17q+, 19q+, 20q+, 21q- and 22q-. The ability of CGH to assess chromosomal changes in breast cancer from fine needle aspirates could facilitate genetic evaluation of tumors prior to surgery.

Frequent amplification of chromosomal region 20q12-q13 in ovarian cancer.

DNA amplification at chromosomal region 20q12-q13, which is common in breast cancer, has recently been described also in ovarian tumors. We studied the amplification of the recently identified candidate oncogenes in this region in 24 sporadic, 3 familial and 4 hereditary ovarian carcinomas, and in 8 ovarian cancer cell lines. High-level amplification of at least one of the five nonsyntenic regions at 20q12-q13.2 was found in 13 sporadic (54%) and in all four hereditary tumors. Typically, two or more distinct amplicons (separated by nonamplified DNA) were found coamplified in various combinations. The regions defined by the AIB1 and PTPN1 genes (at 20q12 and 20q13.1, respectively) were amplified in 25% and 29% of the sporadic tumors, also without simultaneous coamplification of other regions. Amplification of AIB1 (a steroid receptor coactivator gene) was associated with estrogen receptor positivity in sporadic ovarian carcinomas (P = 0.01) and showed a tendency to correlate with poor survival of patients. Of the genes amplified in breast cancer, the BTAK gene was amplified in 21%, the MYBL2 gene in 17%, and the ZNF217 gene in 12.5% of the sporadic tumors. The high frequency of gene amplification at 20q12-q13.2 suggests that the genes amplified therein may play a central role in the pathogenesis of sporadic and hereditary ovarian carcinoma.

Cytogenetical diagnosis in paraffin-embedded fetoplacental tissue using comparative genomic hybridization.

Comparative genomic hybridization (CGH) is a FISH-related technique used to assess global chromosomal aberrations in a variety of human tumours. Recently CGH has been applied to cytogenetic analysis of fresh frozen fetoplacental tissues. Here we report the application of CGH to paraffin-embedded placental samples. Ten samples from paraffin-embedded blocks of 6 control placentas and fetoplacental tissue from 10 aneuploidies, and 2 unbalanced aberrations were evaluated. Balanced karyotype profiles were obtained from samples of healthy placentas and all samples from the same placenta appeared to have similar confidence intervals. CGH analysis of four cases of trisomy 21, three cases of trisomy 18, one case of trisomy 13, one case of trisomy 15 and one case of trisomy 7 all showed overrepresentation of the respective trisomic chromosome. The CGH profile was also in accordance with the karyotyping of a case with isochromosome 21. The CGH profile of a case with der (2)t(2;6)(q37.3;q22.2) revealed partial trisomy for chromosome 6 between q21 and q27. CGH may be a useful adjunct in prenatal genetic diagnosis when retrospective diagnosis is needed from archival samples.

BRCA1 and BRCA2 mutations in ovarian cancer: Covariation with specific cytogenetic features.

We analyzed 37 primary invasive carcinomas for BRCA1 and BRCA2 mutations by screening the entire coding regions of both genes. Seven predicted truncating mutations (four in BRCA1 and three in BRCA2) and one novel BRCA1 missense variant (S1542C) were identified (8/37, 22%). Two of the BRCA1 mutations were somatic changes, whereas the remaining three BRCA1 changes and all mutations of BRCA2 were found to be of germline origin. All eight BRCA-positive tumors were serous or seropapillary carcinomas (8/27 serous tumors, 30%), and all but one were poorly differentiated. The correlation between tumor karyotype and BRCA status showed that clonal chromosomal aberrations were present in all BRCA-positive tumors (8/8) compared with 20 of 29 BRCA-negative ones. The most consistently affected region in BRCA-positive tumors was the long arm of chromosome 6; alterations within this arm with a breakpoint in band 6q21 were seen in four of five BRCA1-positive and in two of three BRCA2-positive tumors, but only in four of 20 karyotypically abnormal tumors without BRCA mutations, suggesting that the genetic pathways of tumor progression differ in the two groups. The high frequency of germline BRCA mutations detected in this pilot study (16% of 37 invasive carcinomas) points to the need for more extended analyses of population-based series of patients to determine the true contribution of these predisposing genes to the overall incidence of ovarian cancer in this population.

Principles of evidence based screening for cancers.

Cancer is a major public health concern. Current estimates suggest that approximately 3 out of every 10 people will be diagnosed with cancer at some point during their lifetime. The American Cancer Society (ACS) estimates that about 1.25 million people in the US will be diagnosed with cancer each year, with nearly 550,000 subsequent deaths. The financial burden of cancer and cost in human lives is staggering. In 1990,it was estimated that the economic burden from cancer in the US was $104 billion: $35 billion for direct costs of prevention, diagnosis and treatment; $12 billion for morbidity costs (economic losses caused by days lost from productive activity because of illness-related disability); and $57 billion in mortality costs (lost economic output measured by loss in earnings because of premature death of productive individuals).

Well-differentiated mucinous carcinoma of the ovary and a coexisting Brenner tumor both exhibit amplification of 12q14-21 by comparative genomic hybridization.

Although the coexistence of mucinous ovarian neoplasms and Brenner tumors is well established, the histogenesis and developmental relationship between the two remain unknown. We used comparative genomic hybridization to analyze two such tumors occurring simultaneously, one in each ovary, in a patient. Amplification of 12q14-21 sequences was found in both tumors; in addition, both tumors also had other, different changes, four identified in the Brenner tumor and six in the mucinous carcinoma. The occurrence of the same genetic alteration in both tumors in this woman suggests that the mucinous carcinoma and Brenner tumor may be clonally related, i.e., one arose from the other by means of metastatic spreading of transformed cells from one ovary to the other. An alternative explanation is that some unknown, putative tumorigenic agent induced similar and synchronous pathogenetic changes in the epithelium of both ovaries. The phenotypic differences between the tumors are presumably attributable to the other unique genetic abnormalities identified in both tumor types.

Cytogenetics of uterine sarcomas: presentation of eight new cases and review of the literature.

Tissue from 14 uterine tumor samples from eight patients-four with endometrial stromal sarcoma (ESS), two with leiomyosarcoma (ULMS), and two with malignant mixed mesodermal tumor (MMMT)-were investigated cytogenetically after short-term culturing. Clonal chromosome aberrations were found in 12 tumors. One ESS showed a recombination between 7p14 and 17q12, a rearrangement characterizing a subset of ESSs. In our series, chromosomes 1, 6, 7, and 16 were involved in structural aberrations most frequently (four cases each). Net loss of 6q material was found in four cases and bands 11q13, 16q13, and 22q13 were each rearranged in four cases. Among 43 uterine sarcomas, including 12 MMTs, now available for evaluation, some differences in breakpoint distribution among different tumor types were found. Rearrangements of bands 1p32, 3p24, and 10q22 were found exclusively in ULMS, whereas aberrations of bands 6p21, 7p21, and 17q12 were found predominantly in ESS.

Genetic aberrations in hypodiploid breast cancer: frequent loss of chromosome 4 and amplification of cyclin D1 oncogene.

The evolution of somatic genetic aberrations in breast cancer has remained poorly understood. The most common chromosomal abnormality is hyperdiploidy, which is thought to arise via a transient hypodiploid state. However, hypodiploidy persists in 1 to 2% of breast tumors, which are characterized by a poor prognosis. We studied the genetic aberrations in 15 flow cytometrically hypodiploid breast cancers by comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH). Surprisingly, numerous copy number gains were detected in addition to the copy number losses. The number of gains per tumor was 4.3 +/- 3.2 and that of losses was 4.5 +/- 3.3 (mean +/- SD), which is similar to that previously observed in hyperdiploid breast cancers. Gains at chromosomes or chromosomal regions at 11q13, 1q, 19, and 16p and losses of 2q, 4, 6q, 9p, 13, and 18 were most commonly observed. Compared with unselected breast carcinomas, hypodiploid tumors showed certain differences. Loss of chromosome 4 (53%) and gain of 11q13 (60%) were significantly more common in hypodiploid tumors. The gain at 11q13 was found by FISH to harbor amplification of the Cyclin D1 oncogene, which is therefore three to four times more common in hypodiploid than in unselected breast cancers (15 to 20%). Structural chromosomal aberrations (such as Cyclin D1 amplification) were present both in diploid and hypodiploid tumor cell populations, as assessed by FISH and CGH after flow cytometric sorting. Together these results indicate that hypodiploid tumors form a distinct genetic entity of invasive breast cancer, although they probably share a common genetic evolution pathway where structural chromosomal aberrations precede gross DNA ploidy changes.

Identification of TP53 gene mutations in uterine corpus cancer with short follow-up.

The involvement of the TP53 tumor suppressor gene in uterine corpus cancer was investigated by single-stranded conformation polymorphism and sequence analysis of its exons 4 to 10. Mutations were found in 12 (18.5%) of 65 cases. Ten of these 12 were single-base substitutions (8 missense and 2 nonsense mutations), whereas 2 were frame-shifting mutations. TP53 gene mutations correlated significantly with advanced surgical stage of disease (P = 0.006) and unfavorable tumor histology types (P = 0.003), whereas the association to myometrial wall invasion did not reach statistical significance (P = 0.054). TP53 gene mutations also correlated significantly with allelic loss at TP53 locus (P = 0.024), absence of estrogen (P = 0.045) and progesterone receptors (P = 0.001), DNA nondiploidy (P = 0.002), and high S-phase fraction values (P = 0.002). Our results suggest that inactivation of the TP53 checkpoint function is associated with disease transition into a stage of rapid progression and spread.

Karyotypic characteristics of borderline malignant tumors of the ovary: trisomy 12, trisomy 7, and r(1) as nonrandom features.

Clonal karyotypic abnormalities were detected in five of 14 cytogenetically analyzed borderline malignant ovarian tumors of clinical stages I-II. One mucinous and one seropapillary tumor had trisomy 7 and r(1)(p36q42) as the sole chromosome abnormality, respectively. Trisomy 12 was found in the remaining three cases. It was the only change in one mucinous and one serous tumor, whereas the third, a seropapillary borderline tumor, had the karyotype 49,XX,+5,+8, +12. These findings, especially when collated with those of previous reports on ovarian borderline tumor cytogenetics, indicate that +12 is the most consistent chromosomal aberration in this group of neoplasms and that also +7 and r(1) are nonrandom features. From the karyotypic point of view, benign ovarian tumors and well-differentiated carcinomas are similar to borderline ovarian tumors, with the possible exception that the former have no tendency to form r(1). Highly malignant carcinomas, on the other hand, are typically much more complex. Chromosome-level changes therefore cannot account for the putative phenotypic passage through the most innocuous tumor stages as epithelial ovarian neoplasms go from benign to fully malignant.

Characterisation of seven human ovarian tumour cell lines.

Characteristics of a panel of seven human ovarian tumour cell lines are presented. Positive staining with HMFG2 and ultrastructural identification of desmosomes confirmed the epithelial nature of the cell lines. The lines showed wide variations in ploidy, doubling times and clonogenicity in soft agar. Both vimentin and keratin were equally expressed in five lines, one line showed strong preferential expression of keratin and one line showed preferential expression of vimentin. Karyotypic changes associated with ovarian cancer were identified in all the lines. Four of the seven cell lines showed loss of chromosome material distal to 11p13-15. These cell lines offer considerable potential for research into the biology and genetics of ovarian cancer.