Elisabeth Kucera

Vascular endothelial growth factor (VEGF) in human breast cancer : Correlation with disease-free survival

Studies have shown that microvessel density influences breast‐cancer prognosis. Since tumor angiogenesis is considered to be substantially affected by the excretion of vascular endothelial growth factor (VEGF) from tumor cells, we examined whether VEGF concentration is different in malignant and in non‐malignant breast tissue. It was also of interest to discover whether intratumoral VEGF concentration influences disease‐free survival (DFS) of breast‐cancer patients. Analysis is based on 120 tissue specimens taken from breast fibromas (n = 23), normal epithelial breast tissue adjacent to fibromas (n = 8) and invasive breast cancer (n = 89). VEGF concentration was quantified by using an immunoassay. Microvessel density was determined by immunostaining for factor‐VIII‐related antigen. Median VEGF concentration is given in pg/mg protein (25%‐quantile—75%‐quantile) and it was 0 (0–1.8) in normal breast tissue, 9.8 (0.52–43.0) in fibromas and 130.4 (50.8–362.2) in invasive carcinomas. A univariate Cox model revealed that node status, tumor size, estrogen‐receptor concentration, histological grading and microvessel density were prognostic factors for disease‐free survival in breast cancer. We found a significant correlation between VEGF concentration and microvessel count, but VEGF concentration did not significantly influence disease‐free survival. Although VEGF protein was found at a significantly higher concentration in malignant than in non‐malignant tissue, determination of intratumoral VEGF protein by an enzyme immunoassay was not prognostically relevant in our patient population. Int. J. Cancer 74:455–458, 1997.

Accuracy of Intraoperative Frozen-Section Diagnosis in Stage I Endometrial Adenocarcinoma

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.

Human papillomavirus in the cervix and placenta.

Objective To determine the prevalence and association of human papillomavirus (HPV) infection in the cervices and placentas of pregnant women. Methods Cervical samples were taken from 179 of 226 women who had placental biopsies because of abnormal ultrasound findings or were older than 35 years, to detect HPV infections with hybrid capture II tests. Polymerase chain reaction (PCR) was done on placental tissue of 147 of the 226 women to detect HPV DNA. Results We found 44 of 179 women (24.6%, 95% confidence interval 18.3, 31.0) to test positive for HPV in their cervices. Logistic regression analyses showed decreased prevalence of HPV infection with increased maternal age (P = .039). The HPV DNA E6 PCR from the villus tissue was negative in the 147 cases examined. However, a significant contingency coefficient between low-risk HPV infection and elevated risk of chromosome aberration was found (&phis; = V = 0.15, P = .050). Conclusion The infection rate of 24.6% in women without clinical symptoms of HPV infection was high, but there seemed to be no virus transmission to the placenta in women with subclinical infections. Low-risk cervical HPV infection might be associated with a slightly higher risk of abnormal fetal karyotype.

Is high-risk human papillomavirus infection associated with cervical intraepithelial neoplasia eliminated after conization by large-loop excision of the transformation zone?

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.

Treatment of endometrial carcinoma with high‐dose‐rate brachytherapy alone in medically inoperable stage I patients

PURPOSE To review the results of treatment with high-dose-rate brachytherapy alone in 228 patients with stage I endometrial carcinoma who are unfit for surgery. METHODS All patients received an exclusive radiation therapy by means of high-dose-rate Iridium 192 intracavitary brachytherapy without additional external beam radiation. RESULTS At 5 years, the overall survival rate was 59.7% and disease specific survival 85.4% at 10 years 30.2% and 75.1%. In clinical stage Ia disease specific survival was 88.6% at 5 years and 82.7% at 10 years, in stage Ib 80.2% and 63.4%, respectively (p<0.02). Disease specific survival was not affected by tumor grade or age. The rates of local control are related to the size of the uterus but not to the tumor grading. Intrauterine recurrence occurred in 17.5% but extrauterine pelvic relapse in only 0.4% of patients. The calculated probability of severe complications was 4.6% at 5 years. CONCLUSION HDR brachytherapy alone achieves excellent disease specific survival rates in patients with medically inoperable stage I endometrial carcinoma.

Prognostic Significance of Mutations in the p53 Gene, Particularly in the Zinc-binding Domains, in Lymph Node- and Steroid Receptor Positive Breast Cancer Patients

The aim of our study was to evaluate if p53 mutations, especially those in the L2/L3 domains of the p53 gene, add prognostic information for node-positive and steroid receptor positive breast cancer patients. Two hundred and five tumour samples from a randomised clinical trial of 596 lymph node- and steroid receptor positive breast cancer patients were included. All patients had been randomly allocated to receive 20 mg of adjuvant tamoxifen (TAM) daily for 2 years or TAM plus one cycle of low-dose, short-term chemotherapy. For detection of p53 mutations we used in vitro amplification by polymerase chain reaction and consecutively performed temperature gradient gel electrophoresis (PCR-TGGE) and direct sequencing. We found p53 mutations in 42/205 (20%) cases: 16/42 (38%) p53 mutations occurred within the L2/L3 domains of the p53 gene, and 26/42 (62%) outside the L2/L3 domains. p53 mutation served as a statistically significant parameter in predicting disease-free survival in univariate (P = 0.02) and multivariate (P = 0.009) analysis. For overall survival, no significant differences were observed. Patients with tumours that had p53 mutations within the L2/L3 domains of the gene showed no significant difference to those with mutations outside the L2/L3 domains for disease-free survival. For overall survival, mutations in the L2/L3 domains showed a marginally significant difference (P = 0.05) in multivariate analysis, but not in univariate analysis (P = 0.13). We conclude that mutation in the L2/L3 domains of the p53 gene is not an independent prognostic indicator of disease outcome for patients suffering from breast cancer with lymph node metastases and positive steroid receptors.

Radiotherapy alone for invasive vaginal cancer: outcome with intracavitary high dose rate brachytherapy versus conventional low dose rate brachytherapy

Purpose. Our aim was to compare the role of remote afterloaded high‐dose‐rate brachytherapy (HDRB) with traditional low‐dose‐rate brachytherapy (LDRB) for patients with invasive primary vaginal carcinoma.

Does T1, N0-1 vulvar cancer treated by vulvectomy but not lymphadenectomy need inguinofemoral radiation?

PURPOSE The objective of our study was to demonstrate differences in relapse rates, total survival times, and complication rates between inguinofemoral radiation and its absence in cases of invasive vulvar carcinoma without lymph node involvement (FIGO Stages T1, N0-1). METHODS AND MATERIALS From 1974 to 1990, 135 patients with invasive vulvar carcinoma in Stage T1 without clinical evidence of inguinal lymph node involvement underwent simple vulvectomy performed by hot-knife resection without lymphadenectomy. Although 65 patients (Group 1) received postoperative inguinofemoral radiation therapy, 70 patients (Group 2) did not, and none received local vulva irradiation. RESULTS The 5-year survival rates were 93.7% in Group 1 and 91.4% in Group 2 (p = NS). Although clitoris involvement was significantly more prevalent in the irradiation group (p = 0.04), inguinal relapse was found less frequently in Group 1 (4.6% or 3 out of 65 patients) than in group 2 (10% or 7 out of 70 patients) (p = 0.32). The complication rates were, 7.7% in Group 1 and 2.9% in Group 2, 2.7% for vaginal stenosis (two patients in each group), 1.5% for inguinal pain (one patient in Group 1), 1.5% for rectovaginal fistula (one patient in Group 1), 1.5% for vulvar infection (one patient in Group 1). CONCLUSION No statistically significant differences in the relapse rates and survival times were found. Risk factors were equally distributed in both study groups except for clitoris involvement. The 5-year survival rates in both groups were similar to those reported in the literature for radical vulvectomy and inguinal lymph-node dissection (83-96%). Morbidity in our study was low. Although our data showed similar results in both groups, we are not recommending at this time to omit groin radiation in general, but it may be justified in low-risk cases.

Loss of heterozygosity (LOH) at p53 is correlated with LOH at BRCA1 and BRCA2 in various human malignant tumors

Mutations inBRCA1 andBRCA2 predispose to breast and ovarian cancers. Several studies have demonstrated that BRCA1 and BRCA2 share similarities not only in their biochemical properties but also in the biochemical pathways by which they are regulated. The protein products of both BRCA1 and BRCA2 genes have transcription-activation function (Chapman and Verma, 1996; Milnert al.,1997). Both bind to Rad51, a protein involved in maintaining the integrity of the genome (Scullyet al., 1997; Sharanet al., 1997). Frequent LOH of both BRCA1 and BRCA2 genes was observed in breast and ovarian cancers. Furthermore, concordant loss of both genes was also reported (Kelsell t al.,1996; Silvaet al., 1999). The combined loss of BRCA1 andBRCA2 leads to the hypothesis that both genes may be induced by or function in overlapping regulatory pathways involved in tumorigenesis. Rajan et al. (1996) demonstrated that BRCA2 mRNA expression was tightly regulated during mammary epithelial proliferation and differentiation and that this regulation occurred coordinately with BRCA1. It has also been demonstrated that the mRNA levels of BRCA1 and BRCA2 were coordinately elevated by the steroid hormone estradiol in breast cancer cell lines (Spillman and Bowcock, 1996). In the ovary, BRCA1and BRCA2 exhibited a comparable hormone-independent pattern of expression in oocytes, granulosa cells and thecal cells of developing follicles (Blackshear et al.,1998). In addition, the products of both genes form a stable interaction in both mitotic and meiotic cells (Chenet al.,1998). BRCA genes are referred to as caretakers rather than gatekeepers. Inactivation of a caretaker gene does not promote tumor initiation directly; it rather leads to genetic instabilities that result in an increased mutation rate affecting all genes including gatekeepers, which directly regulate the growth of tumors by inhibiting growth or promoting death (Kinzler and Vogelstein, 1997). High frequencies ofp53 mutations were found in both breast and ovarian cancers (Hollsteinet al.,1996; Kupryjanczyket al.,1993). Somatic mutations of p53are common in association with germline mutations ofBRCA1 andBRCA2 (Rheiet al.,1998; Crook et al.,1997; Gretarsdottiret al.,1998). In breast cancer, a higher frequency of LOH at 17q and 13q was found in association with p53gene alterations. The increased LOH rate in relation to p53 abnormalities was statistically significant for the BRCA1,but not for theBRCA2, locus (Tsenget al.,1997). According to the tumor-suppressor hypothesis, LOH of 1 allele is frequently accompanied by mutational or regulatory inactivation of the remaining allele and, thus, leads to complete inactivation of a gene (Knudson, 1971). To find out whether p53 inactivation is specifically correlated with that of BRCA1 and BRCA2 in various tumor types of epithelial origin, we examined LOH of these 3 genes in 193 breast carcinomas, 66 ovarian carcinomas, 30 endometrial carcinomas and 39 headand-neck carcinomas. MATERIAL AND METHODS

Genetic alterations in endometrial hyperplasia and cancer

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.