Marianne Waldstrøm

Molecular phenotyping of human ovarian cancer stem cells unravels the mechanisms for repair and chemoresistance

A major burden in the treatment of ovarian cancer is the high percentage of recurrence and chemoresistance. Cancer stem cells (CSCs) provide a reservoir of cells that can self-renew, can maintain the tumor by generating differentiated cells [non-stem cells (non-CSCs)] which make up the bulk of the tumor and may be the primary source of recurrence. We describe the characterization of human ovarian cancer stem cells (OCSCs). These cells have a distinctive genetic profile that confers them with the capacity to recapitulate the original tumor, proliferate with chemotherapy, and promote recurrence. CSC identified in EOC cells isolated form ascites and solid tumors are characterized by: CD44+, MyD88+, constitutive NFκB activity and cytokine and chemokine production, high capacity for repair, chemoresistance to conventional chemotherapies, resistance to TNFα-mediated apoptosis, capacity to form spheroids in suspension, and the ability to recapitulate in vivo the original tumor. Chemotherapy eliminates the bulk of the tumor but it leaves a core of cancer cells with high capacity for repair and renewal. The molecular properties identified in these cells may explain some of the unique characteristics of CSCs that control self-renewal and drive metastasis. The identification and cloning of human OCSCs can aid in the development of better therapeutic approaches for ovarian cancer patients.

The relationship of VEGF polymorphisms with serum VEGF levels and progression-free survival in patients with epithelial ovarian cancer

OBJECTIVE The vascular endothelial growth factor (VEGF) is an important regulator of angiogenesis and vascular permeability of tumors. In the present study we evaluated the relation of five single nucleotide polymorphisms (SNPs) in the VEGF gene with progression-free survival. Furthermore, we evaluated the functional significance of the SNPs as determined by the influence on serum VEGF levels in ovarian cancer. METHODS Serum from 143 consecutive ovarian cancer patients referred for first line platinum/paclitaxel treatment were analyzed for serum VEGF levels using commercially available enzyme-linked immunosorbent assay (ELISA). VEGF gene polymorphisms (-2578 C/A, -1154 G/A, -460 T/C, +405 G/C and +936C/T) were determined by real time PCR using genomic DNA extracted from whole blood samples. RESULTS VEGF serum levels were significantly higher in carriers of the 2578C, 460T and 405C, alleles compared to non-carriers (p=0.003, p=0.003 and p=0.001, respectively). There was no significant correlation between VEGF SNP genotypes and progression-free survival. In haplotype analysis, the multivariate survival analysis showed that progression-free survival (PFS) for the patients with the AGCGC haplotype was significantly improved compared to patients with other haplotypes (HR 1.9, p=0.036). CONCLUSIONS VEGF polymorphisms were found to be significantly related with serum VEGF levels. The AGCGC haplotype was found to be independently associated with improved PFS.

Risk-reducing mastectomy and salpingo-oophorectomy in unaffected BRCA mutation carriers: uptake and timing.

Skytte A‐B, Gerdes A‐M, Andersen MK, Sunde L, Brøndum‐Nielsen K, Waldstrøm M, Kølvraa S, Crüger D. Risk‐reducing mastectomy and salpingo‐oophorectomy in unaffected BRCA mutation carriers: uptake and timing.

Prevalence of Epithelial Ovarian Cancer Stem Cells Correlates with Recurrence in Early-Stage Ovarian Cancer

Epithelial ovarian cancer stem cells (EOC stem cells) have been associated with recurrence and chemoresistance. CD44 and CK18 are highly expressed in cancer stem cells and function as tools for their identification and characterization. We investigated the association between the number of CD44+ EOC stem cells in ovarian cancer tumors and progression-free survival. EOC stem cells exist as clusters located close to the stroma forming the cancer stem cell “niche”. 17.1% of the samples reveled high number of CD44+ EOC stem cells (>20% positive cells). In addition, the number of CD44+ EOC stem cells was significantly higher in patients with early-stage ovarian cancer (FIGO I/II), and it was associated with shorter progression-free survival (P = 0.026). This study suggests that quantification of the number of EOC stem cells in the tumor can be used as a predictor of disease and could be applied for treatment selection in early-stage ovarian cancer.

The relationship of platinum resistance and ERCC1 protein expression in epithelial ovarian cancer.

Objective: Although platinum-based chemotherapy remains the cornerstone for treatment of ovarian cancer, some patients are resistant to the treatment and will therefore not benefit from the standard platinum-based chemotherapy. Preclinical and clinical data have suggested a potential use of excision repair cross-complementation group 1 enzyme (ERCC1) as a molecular predictor of clinical resistance to platinum-based chemotherapy. Excision repair cross-complementation group 1 enzyme is a key enzyme in the nucleotide excision repair pathway which is involved in the DNA repair mechanisms in tumor cells damaged by treatment with platinum agents. The primary aim of the present study was to investigate if immunohistochemical expression of ERCC1 protein was associated with resistance to standard combination carboplatin and paclitaxel chemotherapy in newly diagnosed ovarian cancer patients. Methods: Formalin-fixed, paraffin-embedded tissue sections from 101 patients with newly diagnosed ovarian cancer were used for immunohistochemical staining for the ERCC1 protein. All patients received carboplatin-paclitaxel combination chemotherapy. Results: Excision repair cross-complementation group 1 enzyme protein overexpression was found in 13.9% of the tumors. Platinum resistance was found in 75% of the tumors with positive ERCC1 protein expression compared with 27% among the patients with negative tumor staining for ERCC1 (P = 0.0013). These findings translated into a significant difference in progression-free survival in both univariate (P = 0.0012) and in multivariate analysis (P = 0.006). Conclusions: The data presented suggest a positive association between positive ERCC1 protein expression and clinical resistance to platinum-based chemotherapy.

The predictive value of serum VEGF in multiresistant ovarian cancer patients treated with bevacizumab

OBJECTIVE Bevacizumab, a humanized monoclonal antibody against VEGF (vascular endothelial growth factor), has shown antitumor activity, but so far no biomarkers have been identified to predict outcome. The purpose of the present study was to investigate the efficacy of bevacizumab in patients with multiresistant ovarian cancer and, furthermore, to investigate the possible predictive value of serum VEGF, VEGFR1-2 and VEGF gene polymorphisms. METHODS Patients received single-agent bevacizumab 10 mg/kg every 3 weeks. All patients were followed with CA 125 measurements and serum VEGF/VEGFR1-2 levels prior to each cycle. Endpoints were response rate (RR), progression-free survival (PFS) and overall survival (OS). RESULTS Thirty-eight patients were included. All patients were heavily pre-treated with a median of five prior regimens. The median number of bevacizumab treatments was 4. Overall response rate was 30% according to CA 125 (GCIG criteria). Median PFS was 5.9 months (95% CI, 3.5-9.4) and median OS was 8.6 months (95% CI, 6.6-12.8). The VEGF serum level decreased during treatment in all patients. A low pre-treatment VEGF level was predictive to response. The median value was 540 pg/ml and divided the patients into two groups with a response rate of 60% and 0%, respectively (p=0.0007). The difference translated to a significant difference in PFS (p=0.047) and OS (p=0.01). VEGF gene polymorphisms -2578, -1154, -460, +405, +936 did not reveal any association with response or survival and the same applied to serum VEGFR1-2. CONCLUSIONS Single agent bevacizumab has activity in ovarian cancer patients. Pre-treatment serum VEGF seems to have predictive value.

Evaluation of p16INK4a/Ki-67 dual stain in comparison with an mRNA human papillomavirus test on liquid-based cytology samples with low-grade squamous intraepithelial lesion

The objective of the current study was to investigate the clinical performance of detecting high‐grade lesions with the CINtec PLUS p16INK4a/Ki‐67 dual stain and the APTIMA human papillomavirus (HPV) Assay in a cohort of women with low‐grade squamous intraepithelial lesion (LSIL) cytology. The authors also assessed the reproducibility of the evaluation of immunocytochemical staining.

Prognostic impact of prechemotherapy serum levels of HER2, CA125, and HE4 in ovarian cancer patients.

Objective: Human epididymis protein 4 (HE4) has attracted a lot of interest as a relatively novel biomarker for ovarian carcinoma. Research focus has been directed at HE4 as a diagnostic tool with potential for better triage of women with adnexal masses but the prognostic aspect of HE4 in ovarian cancer patients remains to be elucidated. The aim of the present study was to investigate the prognostic value of prechemotherapy serum HER2, cancer antigen 125 (CA125), and HE4 levels in ovarian cancer patients receiving standard combination chemotherapy. Methods: Serum from 139 patients with newly diagnosed ovarian cancer was analyzed for HER2, CA125, and HE4 using enzyme-linked immunosorbent assay assays. Samples were collected just before first-line chemotherapy, and all patients were treated with carboplatin-paclitaxel combination chemotherapy. Results: Increasing levels of serum HE4 (grouped into quartiles) was significantly associated with worse progression-free survival (PFS) (P < 10−5) and overall survival (P < 10−5). After adjustment in the Cox model, HE4 serum levels remained an independent prognostic parameter for PFS, with a hazard ratio of 1.77 (95% confidence interval, 1.03-3.04; P = 0.040) for patients with HE4 levels above the median compared with patients with HE4 levels below the median. The shorter PFS for patients with high levels of HE4 also translated into an independent significant difference in overall survival (hazard ratio, 3.17 [95% confidence interval, 1.41-7.10]; P = 0.005). Serum HER2 and CA125 levels did not demonstrate an independent prognostic value. Conclusions: High levels of serum HE4 is a strong and independent indicator of worse prognosis in epithelial ovarian cancer patients.

Immunohistochemical Expression of Wilms Tumor Gene Protein in Different Histologic Subtypes of Ovarian Carcinomas

CONTEXT Immunohistochemical expression of Wilms tumor gene protein (WT1) has previously been described in primary ovarian carcinomas. OBJECTIVE To evaluate differences in WT1 expression among different histologic subtypes of ovarian carcinomas and the correlation to the histologic grade. DESIGN Ninety-one primary ovarian carcinomas were reviewed, and 1 representative formalin-fixed and paraffin-embedded tissue block was selected. One slide from each case included in the study was immunostained using the WT1 clone 6F-D2. The immunoreactivity was graded according to the percentage of stained tumor cells. Only nuclear staining was considered a positive reaction. A tumor was regarded as negative if less than 1% of the tumor cells was stained. RESULTS All serous carcinomas (28/28) showed WT1 expression, whereas all mucinous (14/14) and all clear cell carcinomas (14/14) were negative. The lone malignant Brenner tumor and 3 (60%) of 5 undifferentiated carcinomas included in the study were also negative. The endometrioid carcinomas showed either no reaction for WT1 or were diffusely positive with more than 50% of the tumor cells stained. All the grade 1 tumors (10/10) were negative, whereas 5 (45%) of the 11 grade 2 tumors and 5 (63%) of the 8 grade 3 tumors showed a positive reaction. CONCLUSION The present study demonstrates differences in immunohistochemical expression of WT1 among different histologic subtypes of primary ovarian carcinomas. Regarding the endometrioid subtype, the expression seems to be correlated to the histologic grade.

Prognostic importance of vascular endothelial growth factor-A expression and vascular endothelial growth factor polymorphisms in epithelial ovarian cancer.

Introduction: Vascular endothelial growth factors (VEGFs) play a central role in angiogenesis and consequently, in various steps of ovarian carcinogenesis. Gene polymorphisms within the VEGF system have revealed a correlation with prognosis in some malignancies. The aim of the present study was to examine the possible importance of 2 VEGF polymorphisms and VEGF-A expression in ovarian cancer. Methods: We investigated 2 single nucleotide polymorphisms VEGF +405G/C and VEGF −460C/T by polymerase chain reaction and also analyzed VEGF-A expression by immunohistochemistry in 159 women with ovarian cancer. Results: Vascular endothelial growth factor-A expression revealed a significant correlation with survival in a Cox proportional hazards regression model (P = 0.012). Germline polymorphisms were not correlated with clinicopathological parameters such as stage, type, and histology. Heterozygous genotype in VEGF +405G/C predicted a better survival compared with homozygous genotypes (P = 0.034), and the heterozygous genotype in VEGF −460C/T pointed to the same direction. A multivariate analysis also indicated that heterozygosity of either of the 2 polymorphisms held independent prognostic significance (P = 0.001). Conclusions: Vascular endothelial growth factor polymorphisms +405G/C and VEGF expression seem to have independent prognostic importance.