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Liquid biopsy is a promising tool for therapy monitoring of cancer patients, but a need for further research in this field exists in order to improve sensitivity, specificity, standardization and minimize costs. In our present study, we evaluated two panels of transcripts related with the presence of circulating tumor cells (CTCs) (Panel 1: CK19, EpCAM, SCGB2A2 and Panel 2: EMP2, SLC6A8, HJURP, MAL2, PPIC and CCNE2) in two cohorts of breast cancer patients (metastatic and early). A blood cell fraction possibly containing CTCs was isolated with density gradient centrifugation, followed by RNA isolation and qPCR using TaqMan® or RT-qPCR using hybridization probes. The positivity rates of the investigated panels were similar, albeit higher in metastatic (69.4% Panel 1, 75.0% Panel 2; total 86.1%) compared to early (18.9% Panel 1, 23.3% Panel 2; total 31.1%) breast cancer patients. CK19, SCGB2A2, EMP2, HJURP, MAL2, and CCNE2 individually correlated with shorter overall survival in the metastatic patient cohort. The findings highlight the additional value of Panel 2 markers, which are in contrast to CK19 and EpCAM not solely linked to an epithelial phenotype.

Background: While there is a proven association of upper genital tract Ureaplasma infection during pregnancy with adverse pregnancy outcome, the effect of vaginal Ureaplasma colonization on preterm delivery has been controversially debated. Objectives: We hypothesized that women with isolation of vaginal U. parvum but not U. urealyticum are at increased risk for spontaneous preterm birth (SPB) compared to women with negative results. Methods: A vaginal swab taken between 12 and 14 weeks of gestation was analyzed for the presence of Ureaplasma biovars by PCR in 4,330 pregnant women. Results: Of the study cohort, 37% were positive for U. parvum, 5.9% for U. urealyticum, and 3.1% for both. The rates of SPB were 10.4% (OR 1.7, 95% CI 1.3, 2.2, p < 0.001) and 8.9% (OR 1.4, 95% CI 0.9, 2.3, p = 0.193) in the groups with isolation of U. parvum and U. urealyticum, respectively, compared to 6.4% in the group with negative PCR results. Multiple logistic regression and interaction analyses showed that vaginal colonization with U. parvum but not U. urealyticum was a statistically significant risk factor for SPB (adjusted OR 1.6, 95% CI 1.2, 2.1, p < 0.001), independent of other risk factors such as bacterial vaginosis and history of SPB. Conclusion: Our study demonstrates a statistically significant and independent association between first-trimester vaginal colonization with U. parvum and subsequent SPB.

While there is a proven association of upper genital tract Ureaplasma infection during pregnancy with adverse pregnancy outcome, the effect of vaginal Ureaplasma colonization on preterm delivery has been controversially debated.We hypothesized that women with isolation of vaginal U. parvum but not U. urealyticum are at increased risk for spontaneous preterm birth (SPB) compared to women with negative results.A vaginal swab taken between 12 and 14 weeks of gestation was analyzed for the presence of Ureaplasma biovars by PCR in 4,330 pregnant women.Of the study cohort, 37% were positive for U. parvum, 5.9% for U. urealyticum, and 3.1% for both. The rates of SPB were 10.4% (OR 1.7, 95% CI 1.3, 2.2, p0.001) and 8.9% (OR 1.4, 95% CI 0.9, 2.3, p = 0.193) in the groups with isolation of U. parvum and U. urealyticum, respectively, compared to 6.4% in the group with negative PCR results. Multiple logistic regression and interaction analyses showed that vaginal colonization with U. parvum but not U. urealyticum was a statistically significant risk factor for SPB (adjusted OR 1.6, 95% CI 1.2, 2.1, p0.001), independent of other risk factors such as bacterial vaginosis and history of SPB.Our study demonstrates a statistically significant and independent association between first-trimester vaginal colonization with U. parvum and subsequent SPB.

Objective Prohibitin is an important antiproliferative protein inhibiting cell proliferation by blocking the G1/S transition of the cell cycle. Recent findings indicate that the presence of at least one mutant allele within a certain prohibitin gene polymorphism causes inactivation of bioactive RNA resulting in the loss of its pro-apoptotic function and a subsequent risk for malignant growth. Based on these findings we studied whether the presence of this prohibitin polymorphism increases the risk and worsens the prognosis of ovarian cancer in Caucasian women. Study design A polymorphism within the 3′ untranslated region (UTR) of the prohibitin gene was evaluated by pyrosequencing in 136 Caucasian patients with epithelial ovarian cancer and 129 healthy Caucasian controls. Results The wild-type C/C, heterozygous C/T, and the mutant T/T prohibitin genotype was found in 88 (64.7%), 46 (33.8%), and 2 (1.5%) patients with ovarian cancer and in 84 (65.1%), 39 (30.2%), and 6 (4.7%) healthy controls. Presence of at least one mutant allele of the prohibitin 3′ UTR polymorphism was not associated with an increased risk of ovarian cancer as compared to healthy controls ( P = 0.9). No association was found between presence of the prohibitin 3′ UTR polymorphism and the clinico-pathological parameters tumor stage, tumor grade, and patients’ age at diagnosis. Presence of at least one mutant allele of the prohibitin 3′ UTR polymorphism was not associated with disease-free and overall survival. Conclusion The prohibitin 3′ UTR polymorphism was not associated with risk and prognosis of ovarian cancer in Caucasian women.

Objective To determine whether the cyclooxygenase (COX)-2 inhibitor rofecoxib increases the regression rates of cervical intraepithelial neoplasia (CIN) grade II and III. Study design A prospective, randomized, placebo-controlled, double-blind study with rofecoxib 25 mg daily for six months as active treatment for patients with CIN II and III was started in May 2005 and was halted after rofecoxib withdrawal in October 2005. Results A total of 16 patients with CIN II (n = 9) and CIN III (n = 7) were included in our study. Eight and eight patients received rofecoxib and placebo, respectively. Regression rates in the rofecoxib and placebo arm were statistically not significant (25% versus 12.5%) after a mean of 87 (46.3) days of treatment. No severe side effects were noted during the therapy; no dropouts were recorded. Conclusion A conservative treatment of CIN II and III is feasible. Inhibitors of COX, especially COX-2, were seen as candidates for cancer chemoprevention. Our study was halted after rofecoxib was withdrawn. The results obtained should be added to those already published for planning future studies on medical therapies for high grade CIN.

Objective. The enzyme matrix metalloproteinase (MMP)-1 is involved in ovarian carcinogenesis. A common guanine insertion–deletion promoter polymorphism within the gene encoding MMP-1 ( MMP1 ) has been suggested to be a candidate gene for ovarian cancer. We investigated whether this common polymorphism can also serve as independent prognostic parameter in a large series of affected women. Methods. The MMP1 promoter polymorphism was examined in 151 Caucasian patients with epithelial ovarian cancer using polymerase chain reaction. Results were correlated with clinical data. Results. No associations were ascertained between the MMP1 polymorphism and tumor stage ( P = 1.0, odds ratio [OR] 1.08), lymph node involvement ( P = 1.0, OR 0.8), tumor grading ( P = 0.2, OR 0.5), and patient's age at diagnosis ( P = 1.0, OR 1.04). Besides the clinically established prognosticators, tumor stage and histological grade, presence of the MMP1 polymorphism was associated with a shortened disease-free and overall survival in a univariate Kaplan–Meier analysis ( P = 0.01) and a multivariate Cox regression model ( P = 0.04). Conclusion. Presence of the MMP1 gene promoter polymorphisms was found to be a negative prognostic parameter in patients with ovarian cancer.

Objective. To determine whether SCC-Ag serum levels can be used as a prognostic parameter in surgically treated early stage vulvar cancer. Methods. SCC-Ag serum levels were measured preoperatively in 61 surgically staged patients with squamous cell vulvar cancer (UICC pT1 and pT2). Results were correlated to clinical data. Results. Mean (standard deviation) SCC-Ag serum levels in patients with vulvar cancer were 1.5 (1.99) ng/mL. SCC-Ag serum levels were significantly higher in patients with pT2 vulvar cancer (2.2 [2.6] ng/mL) compared with patients with pT1 vulvar cancer (1.0 [1.2] ng/mL, P = 0.034). SCC-Ag serum levels were not associated with lymph node involvement ( P = 0.1), tumor grade ( P = 0.6), and patients' age ( P = 0.5). Multivariate Cox regression models considering tumor stage, lymph node involvement, patients' age, and SCC-Ag serum levels as covariates showed that lymph node involvement ( P = 0.04 and P = 0.01) and tumor stage ( P = 0.006 and P = 0.009), but not SCC-Ag serum levels ( P = 0.8 and P = 0.6), and patients' age ( P = 0.08 and P = 0.22) are prognostic factors for disease-free and overall survival, respectively. Conclusion. SCC-Ag serum levels cannot be used as an additional prognostic parameter in patients with surgically treated early stage vulvar cancer.

Objective. The aim of the study was to determine the diagnostic accuracy and feasibility of sentinel lymph node (SLN) detection using a gamma probe in patients suffering from vulvar cancer. Methods. From May 1998 to November 2000, 26 patients with early vulvar cancer, planned for local wide excision or vulvectomy including groin dissection, were eligible for the study. Two to 3 h before the planned procedure we injected technetium 99 m-labeled microcolloid intradermally at four locations around the tumor. Dynamic and static images were recorded using a gamma camera. SLN locations were marked on the overlying skin. In the operating theater SLNs were identified at the beginning of the procedure using a handheld gamma-detection probe. After resection of suspected SLNs a standard unilateral or bilateral groin dissection was performed, subsequently followed by local wide excision or, if indicated, radical vulvectomy. Sentinel node detection using technetium 99 m-labeled microcolloid was compared with final histopathological and immunohistochemical results. Results. Scintigraphy showed focal uptake in all 26 patients. Intraoperatively we detected all sentinel nodes by handheld gamma probe. In 20 patients, one sentinel node was identified unilaterally, while in 6 patients two or more nodes were identified bilaterally. Histologically positive SLNs were found in 9 patients. In our preliminary series we did not find any false-negative SLN. Conclusion. Identification of sentinel nodes in vulvar cancer is feasible with preoperatively administered technetium 99 m-labeled microcolloid. We confirm the results of previous studies and improve the evidence that the SLN procedure could be implemented in future therapy concepts.

Putative precursors of endometrial cancer such as complex endometrial hyperplasia with atypia have been described to be monoclonal and considered to be genetically related. In order to identify a genetic marker that could serve as a putative predictor of endometrial cancer we analyzed 14 endometrial hyperplasia and 29 endometrial cancer samples for instabilities and loss of heterozygosity (LOH) in microsatellite sequences. Deletions on the short arm of chromosome 8 were frequently detected in both endometrial hyperplasia and cancer samples, suggesting that these deletions are early events in the development of endometrial cancer.

Objective : To investigate whether high-risk HPV infection associated with cervical intraepithelial neoplasia (CIN) was successfully eliminated after electrosurgical conization by large-loop excision of the transformation zone (LLETZ). Study Design : 142 women, who were admitted for conization of CIN 1–3 were recruited into a prospective follow-up study of HPV infection, including cervical sampling for HPV DNA before, and then 3, 6 and 12 months after surgery. We examined whether there were any differences in the rate of HPV DNA positivity after LLETZ between specific risk groups, such as patients with primary (i.e. before surgical treatment) high-risk HPV infection, CIN of different grades, and positive margins. Results : We did not detect statistically significant differences between specific risk groups. According to the assay used (hybrid capture II) at the last follow-up visit 94% of primarily infected patients were completely free from infection with high-risk HPV types, while 6% had persisting HPV infection. Conclusions : With a detection limit of 5000 genomes/ml HPV DNA the hybrid capture II results revealed, that after electrosurgical removal of CIN in 94% of patients testing positive for high-risk HPV DNA prior to surgery were negative 12 months post-surgery.

The aim of this study was to determine the diagnostic accuracy and feasibility of sentinel lymph node (SLN) detection using a gamma probe in patients with Figo IB cervical cancer. Between January 1999 and September 2000, 14 patients with cervical cancer, planned for radical hysterectomy were eligible for the study. The day before radical hysterectomy we injected technetium99m-labelled nanocolloid in each quadrant of the cervix. Dynamic and static images were recorded using a gamma camera. SLNs were identified intraoperatively using a handheld gamma-detection probe. After resection of SLNs a standard radical hysterectomy with pelvic lymph node dissection was performed. Patients and tumour characteristics were compared with sentinel node detection and with final histopathological and immunohistochemical results. Scintigraphy showed focal uptake in 13 of the 14 patients. Intraoperatively we detected 26 sentinel nodes by gamma probe. In 8 of 13 patients, one or more sentinel nodes were identified unilaterally, in 5 women bilaterally. Histologically positive SLNs were found in only 1 patient. We did not find any false-negative SLN in our series. In conclusion identification of sentinel nodes in cervical cancer is feasible with preoperatively administered technetium99m-labelled nanocolloid. A larger series will be required to establish sentinel node detection in cervical cancer for further therapy concepts and planning. © 2001 Cancer Research Campaignhttp://www.bjcancer.com

The main reason for the restricted use of methotrexate in the treatment of ectopic pregnancy (EP) obviously is the fear of tubal rupture in patients with lower abdominal pain after the administration of methotrexate. Therefore, we wanted to find out if patient characteristics at first presentation, such as age, pretreatment β-hCG level, adnexal mass as visualized by transvaginal ultrasonography, or history of prior EP, would identify patients at risk for tubal rupture if they were hemodynamically stable and showed no signs of peritoneal irritation. We examined whether more patients could have been treated medically with methotrexate, because tubal rupture was unforeseeable at first presentation and inclusion criteria for methotrexate treatment were fulfilled. From January 1996 to August 1998, 122 patients diagnosed as having EP were treated at the Gynecologic Department of the University Hospital of Vienna. Inclusion criteria for medical treatment with intramuscular methotrexate (50 mg/ m2 body surface area) were (1) hemodynamic stability, (2) an unruptured ectopic mass ≤5 cm at the greatest dimension demonstrated at transvaginal ultrasonography; (3) β-hCG level ≤5,000 mIU/ml; (4) no cardiac activity of the extrauterine embryo; (5) wish of future fertility, and (6) informed consent. Patients with hemodynamic instability, severe abdominal pain, an ectopic mass ≥5 cm at the greatest dimension, β-hCG levels ≥5,000 mIU/ml, cardiac activity of the extrauterine embryo, and no wish of future fertility, or disagreement with methotrexate treatment, primarily underwent surgery. Despite the fact that none of the above patient characteristics at first presentation identified patients at risk for tubal rupture, only 60/122 patients (49%) actually underwent medical treatment whereas our inclusion criteria would have granted medical treatment in 101/122 patients (83%). We determined the actual and maximal possible percentages of patients with unruptured EP eligible for medical treatment of EP with intramuscular single-dose methotrexate 50 mg/m2 body surface area. Our data show that tubal rupture in hemodynamically stable patients is not foreseeable and should not lead to a restricted use of medical treatment in patients preferring methotrexate.

Our aim was to evaluate the clinical usefulness of serum concentrations of squamous-cell carcinoma antigen (SCC-Ag) and tissue polypeptide antigen (TPA) in the follow-up of patients with vulvar cancer. We measured SCC-Ag and TPA in 480 serum samples of 82 patients with squamous-cell vulvar cancer. Results were correlated with clinical data. SCC-Ag, TPA and the combination of SCC-Ag and TPA reached a sensitivity and specificity of 27%/97%, 28%/75% and 40%/73%, respectively. The sensitivity and specificity of the marker combination SCC-Ag and TPA was not significantly higher compared with SCC-Ag alone (McNemar's test, p = 0.6 and p = 0.09, respectively). Of the 35 patients with recurrent disease during follow-up, 19, 6 and 10 developed local, regional and distant recurrent disease, respectively. SCC-Ag showed lead-time effects in 26%, 75% and 50% and TPA in 25%, 0% and 33% of patients with local, regional and distant recurrent disease, respectively. The combination of SCC-Ag and TPA showed lead-time effects in 50%, 75% and 50% of patients with local, regional and distant recurrent disease, respectively. The difference between median lead-times of the combination of SCC-Ag and TPA and SCC-Ag alone was not statistically significant (Mann-Whitney U-test, p = 0.4). Our data show that serum SCC-Ag displays good sensitivity/specificity characteristics in the follow-up of vulvar cancer patients, with lead-time effects seen in 75% and 50% of patients with regional and distant recurrent disease, respectively. Furthermore, our data indicate that combining SCC-Ag with TPA is not a successful strategy to improve the sensitivity or the duration of lead-time effects in the follow-up of patients with vulvar cancer. Int. J. Cancer 83:167–170, 1999. © 1999 Wiley-Liss, Inc.

Expression of alternatively spliced CD44 isoforms has been reported to correlate with poor prognosis in human squamous cell cancers, i.e. squamous cell cancer of the lung and cervix. The aim of this study was to evaluate whether CD44 isoform expression is a prognostic factor in early-stage squamous cell cancer of the vulva. Seventy cases of squamous cell carcinoma of the vulva International Federation of Gynaecology and Obstetrics (FIGO) stage I were examined immunohistochemically for expression of CD44 isoforms. We used four different variant exon sequence-specific murine monoclonal antibodies to epitopes encoded by exons v3, v5, v6 and v7-8 of human variant CD44. The correlation of CD44 expression with histological grade and disease-free and overall survival was investigated. CD44 isoforms CD44v3, CD44v5, CD44v6 and CD44v7-8 were detected in 28% (20/70), 47% (33/70), 33% (23/70) and 17% (12/70) of the tumour samples respectively. Patients suffering from tumours expressing CD44v6 had a poorer relapse-free (log-rank test, P = 0.02) and overall survival (log-rank test, P = 0.03). Likewise, patients suffering from tumours expressing CD44v3 had a poorer relapse-free (log-rank test, P = 0.04) and overall survival (log-rank test, P = 0.01). Expression of CD44v5 and CD44v7-8 did not compromise the patients' outcome. Histological grade did not correlate with CD44 isoform expression. Immunohistochemically detected expression of CD44 isoforms containing variant exon v6 or v3 is correlated with a poor relapse-free and overall survival in FIGO stage I vulvar cancer patients.

The genotoxic properties of 2',2'-difluorodeoxycytidine (dFdC) were characterised using diploid, mortal low-passage fibroblasts (LPF cells) and the spontaneously transformed fibroblast cell line V79. In both cell types, incorporation of dFdC into the DNA led to an increase of DNA single-strand breaks evaluated by an in situ nick translation assay and to an accumulation of cells in the S-phase of the cell cycle. At concentrations below those leading to cell cycle arrest, dFdC neither induced sister chromatid exchange (SCE) nor structural chromosome aberrations in LPF cells, whereas V79 cells accumulated SCEs as well as chromosome breaks over a broad dose range. In LPF cells treated with dFdC, chromosomal alterations were detected by the micronucleus assay within a narrow concentration range, whereas in V79 cells, a dose-dependent increase in the appearance of micronuclei was seen up to cytotoxic concentrations. In addition, V79 cells went into apoptosis, as evaluated by nuclear fragmentation and condensation, whereas this phenomenon was not detectable in LPF cells.

Background. Cytokines are intercellular hormones, believed to play a functional role in the natural history of various malignant diseases. In vitro and in vivo studies have indicated that interleukin 6 (IL-6) may provide autocrine and paracrine growth stimulation in ovarian cancer cells. Methods. In the present study we measured IL-6 in the serum of 73 patients with FIGO stage I to IV ovarian cancer. Enzyme-linked immunosorbent assay (ELISA) was used to determine IL-6 serum levels. Results were correlated to clinical data. Serum levels of IL-6 were additionally evaluated in a panel of 50 normal controls. Results. Median serum levels of IL-6 in patients with ovarian cancer and normal controls were 55.6 (minimum 0, maximum 2869.0) pg/ml and 0.5 (minimum 0, maximum 2.14) pg/ml, respectively (Mann–Whitney U test, P = 0.0001). When serum levels of IL-6, taken prior to therapy, were grouped by FIGO stage, lymph node involvement, and grading of tumor cells, we found a statistically significant correlation with FIGO stage (Mann–Whitney U test, P = 0.04). Lymph node involvement and grading of tumor cells were not correlated with IL-6 levels. Elevated IL-6 serum levels prior to therapy were significantly correlated with poorer disease-free (log-rank test, P = 0.003) and overall survival (log-rank test, P = 0.01). Conclusion. Elevated IL-6 serum levels prior to therapy are correlated with a poor relapse-free and overall survival in ovarian cancer patients.

Clinical and experimental evidence is consistent with a key role of point mutations of the p53 tumor suppressor gene in the etiology of squamous cell carcinoma. To determine the relation of tumor behavior and patient survival in vulvar cancer in regard to p53 status, we retrospectively analyzed 38 paraffin-embedded specimens of primary vulvar cancer for genetic alterations of exons 5-8 of the p53 gene. For detection of p53 point mutations we used in vitro amplification by polymerase chain reaction (PCR) and temperature gradient gel electrophoresis (TGGE) and, as a detection method, direct sequencing for mutation verification. p53 point mutations were detected in 12/38 tumor specimens. Patients bearing p53 point mutations showed a significantly shorter relapse-free (log-rank test, P = 0.002) and overall survival time (log-rank test, P = 0.0006). We conclude that PCR-TGGE is an appropriate method for detection of p53 point mutations in paraffin-embedded material. We show that loss of wild-type p53 is an adverse prognostic factor in patients suffering from vulvar cancer.

The extent of conization seems to influence the risk of preterm birth. The aim of this study was to compare the cone volume after surgical resection with large loop excision of the transformation zone (LLETZ) and cold knife conization (CKC). The present retrospective multi-center study comprises 804 consecutive women, who underwent LLETZ (n = 412) or CKC (n = 392) between 2004 and 2009. Univariate and multivariable analyses were performed to compare cone volumes removed by LLETZ and CKC and identify independent risk factors for large cone volume. The median resected cone volume after LLETZ was significantly smaller [1.6 cm3 (0.8–2.9)] than after CKC [2.1 cm3 (1.4–3.5)] (

Microvessel density (MVD) and expression of vascular permeability factor/vascular endothelial growth factor (VPF/VEGF), acting as a highly specific inducer of angiogenesis, were evaluated in tissue specimens of 25 patients with squamous cell cancer of the vulva. MVD was quantified by immunostaining for factor VIII-related antigen at one field of 0.25 mm2. VPF/VEGF expression was evaluated immunohistochemically using a monoclonal anti-VEGF antibody. FIGO stages I, II, and III were diagnosed in 12, 7, and 6 patients, respectively. MVD >20/field was found in 10 of 25 tumors and moderate or strong expression of VPF/VEGF in 10 of 25 tumors. High MVD was significantly more frequent in tumors with moderate or strong VPF/VEGF expression compared to tumors with no or weak VPF/VEGF expression (P = 0.01). Overall survival rates of patients with tumors of high MVD (P = 0.01) and strong VPF/VEGF expression (P < 0.01) were significantly poorer compared to those patients with low MVD or poor VPF/VEGF expression. Strong VPF/VEGF expression and high MVD are considered important parameters of tumor angiogenesis and therefore are related to poor survival probability in vulvar cancer patients.

The conversion of plasminogen to active plasmin is thought to be a crucial step in the process of extracellular matrix degradation associated with metastatic spread. Activation of plasminogen is initiated by urokinase plasminogen activator (uPA). The binding of uPA to the uPA cell surface receptor (uPA-R) accelerates plasmin generation from plasminogen and localizes uPA activity to the cell surface. We investigated the mRNA-expression of uPA-R in 19 different human breast cancer cell lines. In a competitive reverse transcription polymerase chain reaction (cRT-PCR) we simultaneously co-amplified two different RNA templates bearing the same primer recognition sequences, the cell line RNA and a known amount of an in vitro synthesized uPA-R-RNA internal standard. We analyzed the two PCR products differing 50 bp in size by agarose gel electrophoresis and calculated the initial uPA-R-RNA template concentration from the relative intensities of the bands quantified by video densitometry. We grouped the investigated cell lines according to their in vitro invasiveness according to literature. Cell lines with a high potential of invasiveness showed a higher expression of uPA-R compared to those with a low potential of invasiveness (Student's t-test, p 0.04). In addition to that we compared the uPA-R mRNA levels with uPA-R, uPA, and PAI-1 protein levels in culture supernatants and cell lysates. The obtained results in breast cancer cell lines with different invasiveness and in benign epithelial cell lines revealed the complex cooperation of the urokinase type proteolytic pathway. uPA, uPA-R, and PAI-1 are to be considered as a diagnostic tool rather than assaying a particular molecule alone. Our findings support the hypothesis that the urokinase proteolytic pathway plays a central role in the acquisition of an invasive phenotype and favors its potential use as a prognostic marker in patients with breast cancer.

Heat shock proteins have been reported to confer resistance to certain antineoplastic drugs. We investigated the impact of hsp70 overexpression on the efficacy of two new anti-cancer drugs, topotecan and gemcitabine. We used the fibrosarcoma WEHI-S cells stably transfected to overexpress the hsp70 cDNA from the constitutive SV40 promoter and appropriate control cells. After topotecan and gemcitabine treatment hsp70-overexpressing cells showed a marked elevation in cell survival, suggesting that hsp70 overexpression was sufficient to confer resistance to the drugs. In addition, hsp70-overexpressing cells were capable of starting cell proliferation after treatment with drug dosages that were lethal to control cells. Our results demonstrate that hsp70 overexpression represents a possible cause of drug resistance. In order to transfer these data to tumour cells constitutively expressing stress hsp70 due to the constitutive activity of the original hsp70 promoter we sought to supress the heat shock response pathway by the natural flavonoid quercetin, known to inactivate the heat shock transcription factor (HSF). Using a suitable cell line, we demonstrated the sensitising activity of quercetin. We found that antineoplastic drug concentrations exerting cytotoxic activity were markedly lower when cells were pretreated with quercetin. Concomitantly, hsp70 expression was strongly down-regulated under quercetin treatment. Our data indicate that quercetin may be useful as a sensitiser in chemotherapeutically treated patients suffering from hsp70-overexpressing tumours. Images Figure 4

Microvessel density in the area of the most intense neovascularization in invasive breast carcinoma is reported to be an independent prognostic factor. The established method of enumeration of microvessel density is to count the vessels using an ocular raster (counted microvessel density [CMVD]). The vessels were detected by staining endothelial cells using Factor VIII-related antigen. The aim of the study was to compare the CMVD results with the percentage of factor VIII-related antigen-stained area using computer-assisted image analysis. A true color red-green-blue (RGB) image analyzer based on a morphologically reduced instruction set computer processor was used to evaluate the area of stained endothelial cells. Sixty invasive breast carcinomas were included in the analysis. There was no significant correlation between the CMVD and the percentage of factor VIII-related antigen-stained area (Spearman correlation coefficient = 0.24, confidence interval = 0.02-0.46). Although high CMVD was significantly correlated with poorer recurrence free survival (P = .024), percentage of factor VIII-related antigen-stained area showed no prognostic value. Counted microvessel density and percentage of factor VIII-related antigen-stained area showed a highly significant correlation with vessel invasion (P = .0001 and P = .02, respectively). There was no correlation between CMVD and percentage of factor VIII-related antigen-stained area with other prognostic factors. In contrast to the CMVD within malignant tissue, the percentage of factor VIII-related antigen-stained area is not suitable as an indicator of prognosis in breast cancer patients.

The stress inducible heat shock protein 70 (hsp70) confers resistance to a variety of adverse environmental conditions including certain anticancer drugs. In the present study we explored cellular consequences of hsp70 overexpression with respect to genotoxic stress. Employing an isogenic set of stable transfectant cells, one overexpressing hsp 70 and the other serving as control, we found that a high hsp70 expression level is sufficient to provide protection against the cytotoxicity of doxorubicin. In addition, hsp70-protected cells showed the capability to restart cell proliferation at concentrations where control cells arrested. However, the DNA lesion density was comparable in the lines used. Recording the cell cycle control revealed a dramatic shortening of the doxorubicin-mediated cell cycle arrest in the G2 phase upon hsp70 overexpression. Our data suggest an involvement of hsp70 in the regulation of the cell cycle.

The expression of specific cell adhesion molecule CD44 isoforms (splice variants) has been shown to be associated with poor prognosis in human malignancies, such as breast cancer. We used three different variant exon sequence-specific murine monoclonal antibodies to epitopes encoded by exons v5, v6 or v7-v8 of human variant CD44, to study the expression of CD44 splice variants by immunohistochemistry in human stage III cervical cancer. We investigated 40 pretreatment punch biopsies of cervical cancer FIGO stage III. CD44 splice variants CD44v5, CD44v6 and CD44v7-8 were detected by means of immunohistochemistry in 90%, 55% and 25%, respectively. CD44 epitopes encoded by exon v5 were not correlated with prognosis. Expression of CD44 splice variants containing epitopes encoded by exon v6 were correlated with significantly poorer prognosis (Mantel test, P = 0.008). Five-year survival rates with or without CD44v6 expression were 20% versus 71%, respectively. Expression of CD44v7-8 was also correlated with significantly poorer overall survival (Mantel test, P = 0.02). Expression of CD44 splice variants containing epitopes encoded by exons v7-v8 and especially exon v6 is associated with significantly poorer prognosis in stage III cervical cancer patients.

We present the data of 105 serum samples from 20 patients suffering from cervical cancer. Mean serum levels of basic fibroblast growth factor (bFGF) in patients with or without tumor present were 31.3 +/- 32.1 (minimum 0, maximum 156.7) pg/ml and 4.8 +/- 6.8 (minimum 0, maximum 29.6) pg/ml, respectively (P = 0.0001). bFGF reached a sensitivity of 65.7% at a specificity of 91.5% when applying a cut-off level of 15 pg/ml. Four patients relapsed after complete remission. A continuous increase of bFGF serum levels before the clinical detection of relapse (lead time) was seen in two cases with a mean lead time of 4 months. Preoperative serum levels were not of prognostic value and showed no correlation with pelvic lymph node metastasis. These preliminary results indicate that in cervical cancer patients soluble bFGF may be useful in early detection of primary tumors, recurrences and monitoring of therapy.

Cervical cytology screening programmes have significantly reduced cervical cancer mortality. In 1988 the National Cancer Institute developed a new classification for reporting cervical cytology--"The Bethesda System". The aim of our study was to compare the clinical usefulness of this new classification with the classification used in German-speaking countries (which discriminates cytologically between moderate and severe dysplasia) including a verbal classification predicting the histological grade of cervical intraepithelial neoplasia (CIN I, II, III). 671 patients with abnormal cervical cytology results and subsequent conisation or hysterectomy were included. We correlated cytological and histological diagnosis (gold standard). In cases of cytologically suggested moderate dysplasia we found histologically CIN I, CIN II and CIN III in 11%, 41% and 42%, respectively. Our results show that discrimination of CIN II and CIN III by means of cytology is limited. Using our therapy schedules we found that adequate therapy is possible by each of the cytological classification systems. The cervical smear is used as a screening test to determine which women require further examination using colposcopy. Under these circumstances an adequate therapy is possible with both classifications. The results underline the importance of colposcopy in the management of cervical dysplasia. As long as conisation is commonly used in German-speaking countries as the only diagnostic tool after an abnormal cervical smear the Bethesda System increases the risk of overtreatment.

The purpose of this study was to evaluate the serum tumor markers CYFRA 21-1 and squamous-cell carcinoma antigen (SCC) in the follow-up of squamous-cell cervical cancer patients. One hundred-ninety-three serum samples, which were collected pretherapeutically and during follow-up of 30 patients suffering from squamous-cell cervical cancer FIGO stage III, were analyzed for SCC and CYFRA 21-1. Cutoff values for SCC and CYFRA 21-1 were 3 and 3.3 μg/liter, respectively. Fifteen cases were keratinizing and 15 nonkeratinizing squamous-cell carcinomas. Serum tumor marker results were correlated to the results of the clinical and radiologic examinations. We calculated sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of serum SCC and serum CYFRA 21-1 for the whole study group and separately for the keratinizing and nonkeratinizing squamous-cell carcinoma group. Sensitivity, specificity, PPV, and NPV of serum SCC were 66, 91, 93, and 60%, respectively. Serum CYFRA 211 showed a sensitivity of 63%, specificity of 96%, PPV of 96%, and NPV of 59%. The combination of SCC and CYFRA 21-1 increased the sensitivity to 78%, with a specificity, PPV, and NPV of 87, 91, and 69%, respectively. In keratinizing squamous-cell carcinoma serum SCC, CYFRA 21-1 and the combination of both had a sensitivity of 76, 64, and 85%, respectively. In nonkeratinizing squamous-cell carcinoma sensitivity was 58, 61, and 72%, respectively. The detection of cervical cancer recurrences with SCC is improved by the combination with CYFRA 21-1. Especially in nonkeratinizing squamous-cell carcinoma CYFRA 21-1 showed promising results.

In this study, correlations between the epidermal growth factor receptor (EGF-R), steroid receptors, and other prognostic parameters (grading, pTNM-status, menopausal status) were analysed in 326 primary breast carcinomas. 19% of the tumour samples were EGF-R positive, 63% were estrogen receptor (ER) and 54% progesteron receptor (PR) positive. Both steroid receptors were positive in 46% of all samples. We found a highly significant inverse correlation between EGF-R and steroid receptors. 88% of the ER positive tumours were EGF-R negative (p less than 5 x 10(-5)), 90% of the PR positive tumours were EGF-R negative (p less than 5 x 10(-5)) and 91% of the ER plus PR positive tumours were ERF-R negative (p less than 1 x 10(-6)). Grading was available in 170 cases. Six (4%) of the carcinomas were highly differentiated (G1), 82 (48%) were classified as G2, and another 82 (48%) were poorly differentiated (G3). A combination of negative ER and positive EGF-R was found more often in the population of G3 tumours. EGF-R was also positively correlated to tumour size. With regard to receptor status, we did not find a correlation with lymph node involvement. The ER correlated negatively (p less than 1.3 x 10(-5) and the EGF-R positively (p less than 0.042) with menopausal status. Thus, EGF-R overexpression seems to be a marker of morphological and functional dedifferentiation which is associated with a loss of steroid dependency and an increase of an autostimulatory-paracrine growth control. These changes seem to be related to poor prognosis.

The clinical value of colposcopy and biopsy prior to large loop excision of the cervical transformation zone (LLETZ) with respect to negative histology and positive specimen margin rates, was evaluated.A total of 2,050 consecutive patients who had undergone LLETZ from 1997 to 2005, were included. As diagnostic workup prior to LLETZ, 290 patients underwent repeat PAP test only and 1,760 patients had undergone colposcopy/biopsy.The diagnostic approach using colposcopy/biopsy reduced the rates of negative histology and positive margins from 6.0% to 1.9% (p0.001) and from 28.2% to 21.7% (p = 0.002), respectively. The rates of invasive carcinomas were not different between the groups (6.6% vs. 6.5%; p = 0.9).Performing colposcopy and biopsy prior to LLETZ reduces unnecessary surgical procedures and decreases positive margin rates.

DNA was extracted from tumour samples of 77 patients with primary breast carcinoma and HER-2 proto-oncogene amplification was assessed. Prognostic indices such as number of positive lymph nodes, tumour size and histological grading were strongly associated with overall survival. No statistically significant correlation between amplification of HER-2 and overall survival was observed. In addition, prognostic indices, HER-2 amplification and disease-free interval was not correlated. Analysis of HER-2 amplification alone is not a useful guide in the management of patients with mammary carcinoma.

Laparoscopic surgery in patients with endometrial cancer has been more widely used in the last 5-10 years, and numerous reports concerning recurrence rate and the surgical technique, laparoscopic vs. open, have been published. However, no data are available concerning the surgical technique of an open or closed vaginal cuff and the vaginal recurrence rate in patients with endometrial cancer.Medical records of 273 patients with endometrial cancer first diagnosed between January 2000 and December 2005 have been reviewed. All patients underwent total abdominal hysterectomy (TAH) or laparoscopically assisted vaginal hysterectomy (LAVH), bilateral salpingo-oophorectomy, peritoneal washings at the Department of Gynecology and Obstetrics of the Medical University of Vienna, Austria. Pelvic lymphadenectomy was performed in high risk patients with grade 3, grade 22 cm diameter, adeno-squamous, clear cell or papillary serous histology, tumor invading the outer half of the myometrium and cervical extension.Thirty-two out of 273 patients with endometrial cancer had recurrent disease. Sixteen patients out of 126 with primary closure of the vaginal cuff had recurrent disease, 8 of them locally; 12 patients out of 130 with open vaginal cuff had recurrent disease, 6 of them locally. Four patients out of 17 with no available information had recurrent disease, 2 of them locally. No significant correlation between the local recurrence rate and surgical technique was found (Spearman's correlation coefficient 0.001, p = 0.994).The surgical technique of an open or closed vaginal cuff during open or laparoscopic surgery for endometrial cancer showed no significant impact on the local recurrence rate.

Objective. To evaluate whether C-reactive protein (CRP) serum levels can be used as a prognostic parameter in patients with cervical cancer. Methods. In the present study, CRP serum levels were measured in 215 patients with cervical cancer. CRP serum levels were measured prior to therapy for cervical cancer and were correlated to clinical data. Results. The median CRP serum level in patients with cervical cancer was 0.5 mg/dl (interquartile range 0.5–0.9 mg/dl). CRP serum levels were significantly associated with advanced tumor stage ( p p =0.01) and patients' age ( p =0.01), but not with histological grade ( p =0.1) and histological type ( p =0.9). In a univariate and multivariate analysis CRP serum levels, tumor stage, and lymph node involvement, but not histological grade, histological type and patients' age were associated with overall and disease-free survival. Conclusion. CRP serum levels can be used as additional prognostic parameter in patients with cervical cancer.

Interleukin-1 (IL-1) is a multifunctional cytokine playing a central role in the immune response and displaying direct cytotoxic activity in vitro. Serum IL-1 alpha and beta levels were measured by enzyme linked immunosorbent assay (ELISA) in 75 ovarian cancer patients, 30 patients with benign ovarian cysts and 50 healthy controls. Both serum IL-1 alpha and IL-1 beta levels were more often elevated in ovarian cancer patients compared with healthy controls (chi-square test, P < 0.001 and P < 0.001, respectively). Mean serum IL-1 alpha and beta levels decreased significantly after surgical intervention (paired t-test, P = 0.0001 and P = 0.0002, respectively). No correlation with histopathological parameters and overall and disease-free survival was found. These preliminary results indicate that serum levels of IL-1 alpha and beta represent a host defence reaction rather than an autonomous tumour cell production.

Cytokeratins are polypeptides which constitute a subclass of intermediate filaments in epithelial cells. The serum tumour marker M 3 M 21 is based on monoclonal antibodies against the epitopes M3 and M21 of cytokeratin 18. In the present study, we measured M 3 M 21 serum levels in 50 patients with FIGO stage IB–IIB cervical cancer and in 50 control subjects using a two-site radiometric immunoassay directed against soluble fragments of cytokeratin 18. Median serum levels of M 3 M 21 in patients with cervical cancer and in normal controls were 70.6 U/ml (range 0–397.7) and 6.5 U/ml (range 0–205.2), respectively (Mann-Whitney U-test, P = 0.0001). Median serum levels of M 3 M 21 prior to therapy and 4 weeks after therapy were 104.2 U/ml (range 24.6–397.7) and 39.3 U/ml (range 0–234.7), respectively (Mann-Whitney U-test, P = 0.004). We found a significant correlation between elevated M 3 M 21 serum levels and metastatic disease in pelvic lymph nodes (Mann-Whitney U-test, P = 0.002). 24 patients relapsed after complete remission. In these patients, elevated M 3 M 21 serum levels before the detection of relapse by computed tomography was observed in 13 cases. Considering these preliminary results, further studies with an increased number of patients are justified to clarify the prognostic value and the monitoring abilities of M 3 M 21 in cervical cancer patients.

To determine whether the cyclooxygenase (COX)-2 inhibitor rofecoxib increases the regression rates of cervical intraepithelial neoplasia (CIN) grade II and III.A prospective, randomized, placebo-controlled, double-blind study with rofecoxib 25 mg daily for 6 [corrected] months as active treatment for patients with CIN II and III was started in May 2004 [corrected] and was halted after rofecoxib withdrawal in October 2004 [corrected]A total of 16 patients with CIN II (n=9) and CIN III (n=7) were included in our study. Eight and eight patients received rofecoxib and placebo, respectively. Regression rates in the rofecoxib and placebo arm were statistically not significant (25% versus 12.5%) after a mean of 87 (46.3) days of treatment. No severe side effects were noted during the therapy; no dropouts were recorded.A conservative treatment of CIN II and III is feasible. Inhibitors of COX, especially COX-2, were seen as candidates for cancer chemoprevention. Our study was halted after rofecoxib was withdrawn. The results obtained should be added to those already published for planning future studies on medical therapies for high grade CIN.

Besides its important role in immune response and inflammatory processes the cytokine interleukin-6 (IL-6) is crucially involved in carcinogenesis. A common polymorphism within the gene encoding IL-6 (IL6) is known to alter IL-6 protein expression and has been associated with patients’ prognosis in various malignancies. No data are available with respect to vulvar cancer. Therefore, we determined the prognostic potential of the common — 174(G→C) single nucleotide polymorphism in the promoter region of IL6 in a series of patients with this disease. The IL6 promoter polymorphism was investigated in 81 Caucasian patients with surgically treated squamous cell vulvar cancer using pyrosequencing. Results were correlated with clinical data. association was ascertained between the IL6 promoter polymorphism and the investigated clinicopathologic parameters, ie, tumor stage, lymph node involvement, tumor grade, and patient’s age at diagnosis. In an univariate analysis, lymph node involvement and patients’ age at diagnosis were associated with patient prognosis. In a multivariate analysis, including tumor stage and lymph node involvement as established prognostic factors and the IL6 promoter polymorphism, lymph node involvement, and the presence of at least one mutant allele, but not tumor stage, were associated with increased disease-free and overall survival. Our data suggest that the IL6 — 174(G→C) promoter polymorphism might serve as an additional prognostic parameter in patients with vulvar cancer.

Objectives. Thrombospondin-1 (TSP-1) is a multifunctional matricellular glycoprotein involved in several mechanisms critical to the formation and progression of solid tumors including cell adhesion, proliferation, migration, invasion, and angiogenesis. The present study was designed to investigate the expression of TSP-1 in invasive vulvar squamous cell carcinoma. Methods. A total of 75 invasive vulvar squamous cell carcinomas were evaluated for TSP-1 expression by immunohistochemistry. Results were correlated with the clinicopathologic parameters including tumor stage, groin lymph node status, tumor grade, patient's age, patients' disease-free, and overall survival. Results. TSP-1 expression was detected in 35/75 (46.7%) specimens of invasive vulvar squamous cell carcinomas. The expression of TSP-1 was generally localized to the cytoplasm and occasionally seen in the nucleus. An increased TSP-1 expression was detected in patients with an advanced tumor stage ( P = 0.01) and a positive groin lymph nodes status ( P = 0.01). Tumor stage and groin lymph node status were associated with patients' disease-free and overall survival. All other parameters failed to be of prognostic significance. Conclusions. We are the first to report on the immunohistochemical expression of TSP-1 in invasive vulvar squamous cell carcinoma. Increased TSP-1 expression was associated with an advanced tumor stage and a positive groin lymph node status, suggesting its pro-angiogenic potential in vulvar carcinogenesis.

5561 Background: Neopterin is produced by activated macrophages upon stimulation with interferon-gamma (IFN-g) and thus elevated concentrations in patients indicate cellular immune response. Most s...

Programmed cell death by apoptosis occurs in fetal and maternal tissues during early pregnancy and plays an important role during implantation, decidualization, and in fetal development. In the regulation of apoptosis, bcl-2 is one of the central controlling genes, and acts by protecting the cell against apoptosis. It is postulated that invasiveness of ectopic trophoblast towards and through the muscularis zone of the tubal wall consequently leading to tubal rupture might be due to disturbed regulation of apoptosis. By means of immunohistochemistry and a computerized image analysis, bcl-2 immunostaining was localized and quantified in 36 randomly selected paraffin-embedded ectopic trophoblast tissue specimens collected from women undergoing surgery for ruptured (n = 18) and non-ruptured (n = 18) tubal ectopic pregnancies. Immunostaining was found in the villi syncytiotrophoblast in all patients, while the percentage of positive bcl-2 immunostained area (%PA) (P = 0.0009) and staining intensity (P = 0.0042) were consistently greater in the group of ruptured ectopic pregnancies. Including the variables %PA and saturation into a logistic regression model for a probability threshold of 0.5 (0.5 = non-ruptured ectopic pregnancy,0.5 = ruptured ectopic pregnancy) to identify tubal rupture, a sensitivity and specificity of 94.4% were found. It is suggested that elevated bcl-2 immunostaining in the syncytiotrophoblast layer reflects unlimited cell survival of ectopic trophoblast and could lead to the establishment of a circulating marker for tubal rupture.

The purpose of our study was to determine if frozen-section diagnosis accurately identified patients suffering from endometrial adenocarcinoma FIGO stage I for surgical staging consisting of total abdominal hysterectomy, bilateral salpingo-oophorectomy, peritoneal cytology, and complete bilateral pelvic lymphadenectomy in moderately differentiated tumors with myometrial invasion. In all poorly differentiated tumors, and in all tumors with deep myometrial invasion (more than 50%) surgical staging included additional para-aortic lymphadenectomy. We performed a retrospective study including 70 patients. Frozen-section diagnosis of myometrial invasion and tumor grade was compared with permanent-section diagnosis. The accuracy rates were determined, and compared with accuracy rates of frozen-section diagnosis in the literature, and a total accuracy rate for 624 patients suffering from stage I endometrial adenocarcinoma was evaluated. In our patient collective, the overall accuracy rate of frozen-section diagnosis for myometrial invasion and tumor grade was 80 and 84%, respectively. In the five comparable studies, the mean accuracy rate for myometrial invasion and tumor grade was 89 and 84%, respectively. In combination with the five comparable studies our recent study produced an accuracy rate of frozen-section diagnosis for myometrial invasion and tumor grade of 88 and 84% in 624 patients, respectively. Despite an accuracy level of frozen-section diagnosis for myometrial invasion of 80 and 84% for tumor grade in our patient collective, all patients who required surgical staging were accurately identified.

Uterine rupture during the first trimester of pregnancy is an extremely rare, but life-threatening cause of intraperitoneal hemorrhage. Up to the knowledge of the authors all reports of first trimester uterine ruptures are related to scar dehiscences following previous cesarean sections or occurred in unscarred uteri of multiparous women. In cases of multiparity silent ruptures cannot be precluded, so that the uterus might be scarred during the following pregnancy. In early pregnancy of approximately 4-5 weeks, vaginal ultrasonography may clearly verify a scar pregnancy, but sonographical diagnostic findings may change with the pregnancy progress. In all cases of reported first trimester ruptures in pregnancies with previous cesarean sections or in pregnancies of multiparous women reported in literature, dating scans were performed too late for to preclude pregnancies in the scar. We postulate our hypotheses, that all first trimester uterine ruptures are caused by scar implantation of the trophoblast.

Cytokeratins are polypeptides that constitute a subclass of intermediate filaments in epithelial cells. The aim of the present study was to evaluate the clinical usefulness of the serum evaluation of M3/M21 in patients with ovarian cancer. This retrospective study comprises 75 patients suffering from ovarian cancer FIGO stages Ia-III. M3/M21 reached a sensitivity of 78%, a specificity of 85%, a PPV of 89% and a NPV of 83% using a cut-off level of 45 U 1(-1). Forty-four women developed recurrent disease after complete remission during the observation period. M3/M21 showed lead time effects in 19 patients, ranging from 2 to 8 months (median 3.2 months). Elevated M3/M21 serum levels before therapy were associated with a poor overall survival (log-rank test, P = 0.02). Considering these preliminary results, the value of M3/M21 as a serum tumour marker, i.e. to evaluate the tumour burden, seems promising.

For various human tumor cell lines (neuroblastoma, cervix carcinoma) the presence of constitutive nitric oxide synthase (cNOS) has been documented. Here, for the first time, we report about cNOS expression in 10 of 16 human breast cancer cell lines. cNOS expression correlates strongly with expression of estrogen receptor (ER). Competitive reverse transcription-polymerase chain reaction (cRT-PCR) was used to detect cNOS and ER mRNA expression. Our findings suggest that estradiol could stimulate constitutive NO release in breast tissue, where it acts as a free radical.

The surface glycoprotein CD44 is widely distributed in different tissues. In contrast to healthy tissue, tumour samples show a more complex pattern of CD44 expression, indicating a loss of splice control. Beside cell-surface expression, the measurement of soluble CD44 in serum of cancer patients could be useful in early diagnosis and assessment of disease status. We evaluated the surface expression of CD44 isoforms in 22 ovarian cancer patients by means of immunohistochemistry. Additionally, we investigated 134 serological samples of these patients for the occurrence of CD44 isoform expression. For CD44 standard, CD44v5 and CF44v6 mean serum levels in patients with clinically detectable or non-detectable ovarian cancer were 422.4 +/- 143.8 ng ml-1 and 547.4 +/- 148.2 ng ml-1, 12.3 +/- 7.9 ng ml-1 and 21.9 +/- 12.2 ng ml-1 and 105.5 +/- 37.9 ng ml-1 and 144.9 +/- 50.9 ng ml-1 respectively (P-values not significant). CD44 surface proteins containing epitopes encoded by splice variants CD44v5, CD44v6 and CD44v7-8 were immunohistochemically detected in 9% (n = 2), 13% (n = 3) and 4% (n = 1) of the 22 tumour samples respectively. In the present study we showed that in ovarian cancer CD44 isoforms CD44v5 and CD44v6 are expressed in very low amounts by the tumours. In accordance with this, we found that the presence of tumour is not associated with higher serum levels of CD44standard, CD44v5 and CD44v6 in preoperative serum samples in ovarian cancer patients.