Seija Grénman

Amplification and Deletion of Topoisomerase IIα Associate with ErbB-2 Amplification and Affect Sensitivity to Topoisomerase II Inhibitor Doxorubicin in Breast Cancer

Topoisomerase IIalpha (topoIIalpha) is a key enzyme in DNA replication and a molecular target for many anti-cancer drugs called topoII inhibitors. The topoIIalpha gene is located at chromosome band 17q12-q21, close to the ErbB-2 oncogene (HER-2/neu), which is the most commonly amplified oncogene in breast cancer. Because of the physical proximity to ErbB-2, copy number aberrations may also occur in the topoIIalpha gene. These topoIIalpha gene copy number aberrations may be related to the altered chemosensitivity to topoII inhibitors that breast cancers with ErbB-2 amplification are known to have. We used fluorescence in situ hybridization to study copy number aberrations of both topoIIalpha and ErbB-2 in nine breast cancer cell lines and in 97 clinical breast tumors, which were selected for the study according to their ErbB-2 status by Southern blotting. TopoIIalpha-protein expression was studied with Western blot and sensitivity to doxorubicin (a topoII inhibitor) with a 96-well clonogenic in vitro assay. Two of the five cell lines with ErbB-2 gene amplification (SK-BR-3 and UACC-812) showed amplification of topoIIalpha. In MDA-361 cells, ErbB-2 amplification (14 copies/cell) was associated with a physical deletion of topoIIalpha (four copies of chromosome 17 centromere and two copies of topoIIalpha). The topoIIalpha amplification in UACC-812 cells was associated with 5.9-fold-increased topoIIalpha protein expression and 2.5-fold-increased sensitivity to the topoII inhibitor, doxorubicin, whereas the deletion in MDA-361 leads to decreased protein expression (45% of control) and a 2.4-fold-increased chemoresistance in vitro. Of 57 ErbB-2-amplified primary breast carcinomas, 25 (44%) showed ErbB-2-topoIIalpha coamplification and 24 (42%) showed a physical deletion of the topoIIalpha gene. No topoIIalpha copy number aberrations were found in 40 primary tumors without ErbB-2 amplification. TopoIIalpha gene amplification and deletion are common in ErbB-2-amplified breast cancer and are associated with increased or decreased sensitivity to topoII inhibitors in vitro, respectively. These findings may explain the altered chemosensitivity to topoII inhibitors reported in ErbB-2-amplified breast cancers.

Quality of life and cost-effectiveness of levonorgestrel-releasing intrauterine system versus hysterectomy for treatment of menorrhagia: a randomised trial

BACKGROUND Heavy menstrual blood loss is a common reason for women to seek medical care. The levonorgestrel-releasing intrauterine system (IUS) is an effective medical treatment for menorrhagia. We report a randomised comparison of this approach with hysterectomy in terms of the quality of life of women with menorrhagia and cost-effectiveness. METHODS Of 598 women referred with menorrhagia to five university hospitals in Finland, 236 were eligible and agreed to take part. They were randomly assigned treatment with the levonorgestrel-releasing IUS (n=119) or hysterectomy (n=117). The amount of menstrual blood loss was objectively measured. The primary outcome measure was health-related quality of life at 12-month follow-up. Analyses were by intention to treat. FINDINGS In the group assigned the levonorgestrel-releasing IUS, 24 (20%) women had had hysterectomy and 81 (68%) continued to use the system at 12 months. Of the women assigned to the hysterectomy group, 107 underwent the operation. Health-related quality of life improved significantly in both the IUS and hysterectomy groups (change 0.10 [95% CI 0.06-0.14] in both groups) as did other indices of psychological wellbeing. There were no significant differences between the treatment groups except that women with hysterectomy suffered less pain. Overall costs were about three times higher for the hysterectomy group than for the IUS group. INTERPRETATION The significant improvement in health-related quality of life highlights the importance of treating menorrhagia. During the first year the levonorgestrel-releasing IUS was a cost-effective alternative to hysterectomy in treatment of this disorder.

Sequential adjuvant chemotherapy and radiotherapy in endometrial cancer - results from two randomised studies

INTRODUCTION Endometrial cancer patients with high grade tumours, deep myometrial invasion or advanced stage disease have a poor prognosis. Randomised studies have demonstrated the prevention of loco-regional relapses with radiotherapy (RT) with no effect on overall survival (OS). The possible additive effect of chemotherapy (CT) remains unclear. Two randomised clinical trials (NSGO-EC-9501/EORTC-55991 and MaNGO ILIADE-III) were undertaken to clarify if sequential combination of chemotherapy and radiotherapy improves progression-free survival (PFS) in high-risk endometrial cancer. The two studies were pooled. METHODS Patients (n=540; 534 evaluable) with operated endometrial cancer International Federation of Obstetrics and Gynaecology (FIGO) stage I-III with no residual tumour and prognostic factors implying high-risk were randomly allocated to adjuvant radiotherapy with or without sequential chemotherapy. RESULTS In the NSGO/EORTC study, the combined modality treatment was associated with 36% reduction in the risk for relapse or death (hazard ratio (HR) 0.64, 95%confidence interval (CI) 0.41-0.99; P=0.04); two-sided tests were used. The result from the Gynaecologic Oncology group at the Mario Negri Institute (MaNGO)-study pointed in the same direction (HR 0.61), but was not significant. In the combined analysis, the estimate of risk for relapse or death was similar but with narrower confidence limits (HR 0.63, CI 0.44-0.89; P=0.009). Neither study showed significant differences in the overall survival. In the combined analysis, overall survival approached statistical significance (HR 0.69, CI 0.46-1.03; P=0.07) and cancer-specific survival (CSS) was significant (HR 0.55, CI 0.35-0.88; P=0.01). CONCLUSION Addition of adjuvant chemotherapy to radiation improves progression-free survival in operated endometrial cancer patients with no residual tumour and a high-risk profile. A remaining question for future studies is if addition of radiotherapy to chemotherapy improves the results.

Transmission of High-Risk Human Papillomavirus (HPV) between Parents and Infant: a Prospective Study of HPV in Families in Finland

ABSTRACT The Finnish HPV Family Study is a prospective cohort study assessing the dynamics of human papillomavirus (HPV) transmission between parents and infant. Serial genital and oral scrapings from 76 families, including mother, father, and infant, and semen samples were collected over 2 years of follow-up, analyzed by nested PCR, and confirmed by hybridization with 12 high-risk (HR) HPV types. The most common HPV profile was HR HPV in all family members (29%), followed by HPV-positive mother-infant pairs (26%). HPV-positive father-infant pairs were less frequent (11%), and in six (8%) families, only the infant was HR HPV positive. The prevalence of genital HR HPV in the parents ranged from 13 to 25%, and that of oral HPV ranged from 8 to 34%. In the infants, HPV DNA was detected in 15% of the genital and 10% of the oral samples at birth, reaching peaks of 18 and 21%, respectively, at 6 months, and declining to 10% at 24 months. Persistent HPV in the mother was a risk factor for oral HPV in the infant (odds ratio [OR], 5.69; 95% confidence interval [95% CI], 1.5 to 21.3), while oral HPV in the mother at 6 months was a risk factor for genital HR HPV (OR, 6.38; 95% CI, 1.15 to 35.32). No such independent risk could be attributed to subclinical HPV in the father. Persistent maternal cervical HPV and subclinical oral HPV affect the risk of infant HPV. The age of 6 months is a critical point for the infant to acquire or be free of HR HPV DNA.

Collagenase-3 (MMP-13) is Expressed by Tumor Cells in Invasive Vulvar Squamous Cell Carcinomas

Collagenase-3 (MMP-13) is a human matrix metalloproteinase specifically expressed by invading tumor cells in squamous cell carcinomas (SCCs) of the head and neck. Here, we have further elucidated the role of MMP-13 in tumor invasion by examining its expression in invasive malignant tumors of the female genital tract. Using in situ hybridization, expression of MMP-13 mRNA was detected in 9 of 12 vulvar SCCs, primarily in tumor cells, but not in intact vulvar epithelium, in cervical SCCs (n = 12), or in endometrial (n = 11) or ovarian adenocarcinomas (n = 8). MMP-13 expression was especially abundant in vulvar carcinomas showing metastasis to lymph nodes and was associated with expression of membrane type 1 MMP by tumor cells and gelatinase-A (MMP-2) by stromal cells, as detected by immunohistochemistry. MMP-13 mRNAs were detected in 9 of 11 cell lines established from vulvar carcinomas and in 4 of 6 cell lines from cervical carcinomas, whereas endometrial (n = 10) and ovarian (n = 9) carcinoma cell lines were negative for MMP-13 mRNA. No correlation was detected between MMP-13 expression and p53 gene mutations in vulvar SCC cell lines. However, MMP-13 expression was detected in 5 of 6 vulvar and cervical SCC cell lines harboring HPV 16 or 68 DNA. These results show that MMP-13 is specifically expressed by malignantly transformed squamous epithelial cells, including vulvar SCC cells, and appears to serve as a marker for their invasive capacity.

Frequency of anal sphincter rupture at delivery in Sweden and Finland ‐ result of difference in manual help to the baby's head

BACKGROUND Anal sphincter rupture is a serious complication of vaginal delivery and almost half the affected women have persistent defecatory symptoms despite adequate primary repair. During the past decade, the incidence of anal sphincter ruptures has been increasing in Sweden and is currently estimated to occur in 2.5% of vaginal deliveries. The aim of the study was to report the frequency of anal sphincter ruptures in two university hospitals in two Scandinavian countries, Malmö in Sweden and Turku in Finland, and analyze the potential determinants. METHODS Retrospective analysis of a population of 30,933 deliveries (26,541 vaginal) during the years 1990 to 1994. RESULTS The incidence of anal sphincter ruptures in Malmö, Sweden was 2.69%, and in Turku, Finland 0.36%. There were no significant population differences for the known risk factors (fetal weight, nulliparity or fetal head circumference). However, there is a difference in manual support given to the perineum and to the babys head when crowning through the vaginal introitus between Malmö and Turku. The proportion of operative vaginal deliveries and abnormal presentations was significantly higher in Turku reflected in the lower Apgar score at 5 minutes and longer duration of second phase of labor. When high risk deliveries (operative vaginal delivery, abnormal presentation and newborns over 4,000 g) were excluded, the risk for anal sphincter ruptures was estimated to be 13 times higher in Malmö than in Turku. CONCLUSIONS The difference in the incidence of anal sphincter rupture between Malmö, Sweden and Turku, Finland may be due to the difference in manual control of the babys head when crowning.

High-Risk Types of Human Papillomavirus (HPV) DNA in Oral and Genital Mucosa of Infants during Their First 3 Years of Life: Experience from the Finnish HPV Family Study

BACKGROUND This study is aimed to clarify data on the acquisition, persistence, and clearance of high-risk types of human papillomavirus (HPV) DNA from the mucosa and the determinants of persistent mucosal HPV infection in infants. METHODS Oral and genital scrapings from 324 infants were collected at birth, 3 days after delivery, and 1, 2, 6, 12, 24, and 36 months after delivery and tested for the presence of HPV DNA by nested polymerase chain reaction and hybridization with 12 high-risk HPV oligoprobes. HPV status and demographic data for parents were analyzed. RESULTS During the follow-up period (median duration, 26.2 months), HPV DNA was found to be present in 12%-21% of oral scrape samples and in 4%-15% of genital scrape samples obtained from the infants. Oral HPV infection was acquired by 42% of children, cleared by 11%, and persisted in 10% of the infants, whereas 37% were never infected. The corresponding figures for genital HPV infection were 36%, 14%, 1.5%, and 47%. Kaplan-Meier analysis revealed that both the cumulative incidence of infection and clearance of HPV were parallel in oral and genital sites. Persistent oral HPV infection in the child was significantly associated with persistent oral HPV infection in the mother at month 36 of follow-up, hand warts in the mother, young age at onset of sexual activity for the mother, and the mothers use of oral contraception, as well as with the fathers oral HPV status at 24 months. Persistent genital HPV infection in the infant was predicted by if the mother had started smoking at 18-21 years of age and by a history of genital warts. CONCLUSIONS Persistent carriage of high-risk HPV types was detected in oral and genital mucosa specimens obtained from 10% and 1.5% of the infants during their first 26 months of life. The rates of acquisition and clearance of HPV were similar in oral and genital mucosa.

Metalloelastase (MMP-12) expression by tumour cells in squamous cell carcinoma of the vulva correlates with invasiveness, while that by macrophages predicts better outcome

Human metalloelastase (MMP‐12) has been implicated in elastin degradation and macrophage migration in many pathological conditions. It also generates angiostatin, thus having a potential to prevent tumour angiogenesis. It has previously been shown that transformed epithelial cells express MMP‐12 in skin cancer. The aim of this study was further to elucidate the role of metalloelastase in squamous cell cancer (SCC) progression. By in situ hybridization, expression of MMP‐12 mRNA was detected in 28/33 vulvar SCC samples in CD‐68‐positive macrophages, while 10 samples had positive cancer cells. By immunohistochemistry, MMP‐12 protein was seen in the same area as the mRNA. MMP‐12 mRNA expression in tumour cells correlated with more aggressive histology (p = 0.0099). In contrast, macrophage‐derived MMP‐12 mRNA was more abundant in well‐differentiated grade I than grade III tumours (p = 0.01). However, the level of MMP‐12 mRNA, regardless of its origin, did not correlate with metastasis or patient survival. No significant correlation was found between macrophage‐derived MMP‐12 mRNA and a low amount of blood vessels, as quantitated after von Willebrand staining. In agreement with vulvar SCCs in vivo, MMP‐12 was expressed in cultured SCC cells by northern and western blot analysis. In HaCaTs and epithelial MCF‐10f cells, MMP‐12 mRNA was induced by transforming growth factor‐β1 (TGF‐β1) and tumour necrosis factor‐α (TNF‐α) as measured by quantitative RT‐PCR (TaqMan). Two MMPs capable of generating angiostatin in vivo, matrilysin (MMP‐7) and gelatinase B (MMP‐9), were also examined in these tumours. MMP‐7 mRNA was mainly expressed by epithelial tumour cells, particularly in less differentiated tumours. MMP‐9 was usually expressed by neutrophils and macrophages; epithelial protein was predominantly found in grade II/III tumours. These results suggest a dual role for MMP‐12 in tumour progression. Copyright

Surgically staged high-risk endometrial cancer: Randomized study of adjuvant radiotherapy alone vs. sequential chemo-radiotherapy

OBJECTIVE Our purpose was to establish whether platinum-based chemotherapy combined with standard surgery and radiotherapy will improve overall and disease-free survival and lower the recurrence rate in patients with high-risk endometrial cancer. STUDY DESIGN A total of 156 patients with Stage IA-B Grade 3 (n=28), or Stage IC-IIIA Grade 1-3 (n=128) were postoperatively randomized to receive radiotherapy (56 Gy) only (Group A, n=72) or radiotherapy combined with three courses of cisplatin (50 mg/m(2)), epirubicin (60 mg/m(2)) and cyclophosphamide (500 mg/m(2)) (Group B, n=84). RESULTS The disease-specific overall five-year survival was in Group A 84.7% vs. 82.1% in Group B (p=0.148). The median disease-free survival in A was 18 (range 9-36) months and 25 (range 12-49) months in B (p=0.134), respectively. During a five-year follow-up 32 patients relapsed. Of the recurrences 5 were local and 20 distant, while 7 were combined. As calculated from the operation, the median time to recurrence was 15 (range 6-37) months in Group A, and 20 (range 8-60) months in Group B, respectively (p=0.170). Twenty-six patients died of the disease during the five-year follow-up, 11 in A and 15 in B. The patients succumbing in A lived a median 23 (range 15-44) months as compared to 37 (range 13-50) months in B (p=0.148). Chemotherapy was associated with an acceptable rate of acute toxicity. Less than 8% of the patients complained of Grade 3/4 nausea. The rate of Grade 3/4 leucopenia was at the highest at 16.6% during the third cycle but only 6.2% of the patients had Grade 3 infection. A total of 10 patients developed intestinal complications demanding surgery, 2 in Group A (2.7%) and 8 (9.5%) in Group B, respectively. CONCLUSION Adjuvant chemotherapy with cisplatin, epirubicin and cyclophosphamide failed to improve overall survival or lower the recurrence rate in patients operated on and radiated for high-risk endometrial carcinoma. Chemotherapy was associated with a low rate of acute toxicity but appeared to increase the risk of bowel complications.

Human papillomavirus in the placenta and umbilical cord blood

Objective. To analyze human papillomavirus (HPV) DNA in umbilical cord blood and in placenta, including its cellular localization. Design. Longitudinal prospective study. Setting. Maternity Unit of Turku University Hospital, and MediCity, University of Turku. Samples. Placental and cord blood samples obtained at delivery from 315 mothers and 311 neonates included in the Finnish HPV Family Study. Methods. HPV testing by nested PCR and sequencing. Tyramide amplified in situ hybridization (ISH) for viral DNA localization in placenta. Correlation to mothers and neonates oral and genital HPV status and maternal demographic data. Main outcome measures. Detection and cellular localization of HPV DNA. Results. HPV DNA was detected in 4.2 and 3.5% of placenta and cord blood samples, respectively, including HPV types 16, 6, 83 and 39. In placenta, HPV6 and 16 DNA was localized in syncytiotrophoblasts. Abnormal cytology increased the risk of HPV+ placenta and cord blood. History of genital warts was the only independent predictor of cord blood HPV in multivariate analysis (adjusted OR=4.0, 95% CI: 1.09–14.54, p = 0.036). HPV DNA in cord blood increased the risk of genital (OR=4.0, 95% CI: 1.08–14.83, p = 0.048) and oral (OR=4.4, 95% CI: 1.17–16.14, p = 0.039) HPV DNA carriage of the neonate. HPV+ placenta increased the risk of oral HPV of the neonate (OR=8.6, 95% CI: 2.73–27.13, p = 0.0001). Delivery mode did not predict HPV status of the neonate. Conclusions. HPV DNA is detected in placental trophoblasts and umbilical cord blood. The presence of HPV DNA at these sites increases the risk of a neonate testing HPV‐positive at birth.