Marek Spaczyński

Immunogenicity and tolerability of an HPV-16/18 AS04-adjuvanted prophylactic cervical cancer vaccine in women aged 15-55 years

The immunogenicity and safety of an HPV-16/18 AS04-adjuvanted vaccine were assessed in women aged 26-55 years and compared with women aged 15-25 years in a Phase III, non-randomised, open-label, age-stratified study. Overall the vaccine was well tolerated and 100% seropositivity was achieved 1 month after the third dose in all age groups. There was a high correlation between HPV-16 and HPV-18 antibody levels (IgG) in cervicovaginal secretions and sera, regardless of age. The HPV-16/18 AS04-adjuvanted vaccine induces a robust and persistent immune response in women >26 years of age and generates antibodies that transudate through the cervix epithelium.

TLR4 signaling induced by lipopolysaccharide or paclitaxel regulates tumor survival and chemoresistance in ovarian cancer.

Toll-like receptors (TLRs) expressed on immune cells trigger inflammatory responses. TLRs are also expressed on ovarian cancer (OvCa) cells, but the consequences of signaling by the TLR4/MyD88 pathway in these cells are unclear. Here, TLR4 and MyD88 expression in OvCa tissues (n=20) and cell lines (OVCAR3, SKOV3, AD10, A2780 and CP70) was evaluated by reverse transcriptase–PCR, western blots and immunohistochemistry. Cell growth, apoptosis, nuclear factor-κB (NF-κB) translocation, IRAK4 and TRIF expression and cJun phosphorylation were measured following tumor cell exposure to the TLR4 ligands, lipopolysaccharide (LPS) or paclitaxel (PTX). Culture supernatants were tested for cytokine levels. TLR4 was expressed in all tumors, tumor cell lines and normal epithelium. MyD88 was detectable in tumor tissues and in 3/5 OvCa lines but not in normal cells. In MyD88+ SCOV3 cells, LPS or PTX binding to TLR4 induced IRAK4 activation and cJun phosphorylation, activated the NF-κB pathway and promoted interleukin (IL)-8, IL-6, vascular endothelial growth factor and monocyte chemotactic protein-1 production and resistance to drug-induced apoptosis. Silencing of TLR4 in SCOV3 cells with small interference RNA resulted in phosphorylated-cJun (p-cJun) downregulation and a loss of PTX resistance. In PTX-sensitive, MyD88neg A2780 cells, TLR4 stimulation upregulated TRIF, and TLR4 silencing eliminated this effect. Thus, TLR4/MyD88 signaling supports OvCa progression and chemoresistance, promoting immune escape.

ABCC2 (MRP2, cMOAT) can be localized in the nuclear membrane of ovarian carcinomas and correlates with resistance to cisplatin and clinical outcome.

Purpose: Cisplatin resistance is a major obstacle in the treatment of ovarian carcinoma. ABCC2 is commonly localized in apical cell membranes and could confer cisplatin resistance. Here, we show that ABCC2 can be localized in the cytoplasmic membrane as well as in the nuclear membrane of various human tissues including ovarian carcinoma cells. Experimental Design: For the subcellular detection of ABCC2, immunohistochemistry was done using 41 Federation Internationale des Gynaecologistes et Obstetristes stage III ovarian carcinoma specimens prepared before treatment with cisplatin-based schemes and 35 specimens from the same group after chemotherapy. Furthermore, 11 ovarian carcinoma cell lines as well as tissue microarrays consisting of various human tissues were analyzed. Results: Nuclear membranous localization of ABCC2 was associated with response to first-line chemotherapy at primary (P = 0.0013) and secondary surgery (P = 0.0060). Cases with relapse showed higher nuclear membrane expression at primary (P = 0.0003) and secondary surgery (P = 0.0024). Kaplan-Meier analyses showed that weak nuclear membrane ABCC2 expression before treatment was associated with significantly longer overall (P = 0.04) and progression-free survival (P = 0.001); following chemotherapy, it correlated with significantly longer progression-free survival (P = 0.038). Tissue microarrays confirmed nuclear membranous localization of ABCC2, in particular, in poorly differentiated cells. In ovarian carcinoma cells, it correlated with resistance against cisplatin, whereas localization in the cytoplasmic membrane did not. Conclusions: ABCC2 confers resistance to cisplatin of ovarian carcinoma in cell culture systems and in clinics when expressed in the nuclear membrane. Thus, ABCC2 localization can predict platinum therapy outcome. Furthermore, expression of ABCC2 in nuclear membranes in human tissues is specific for poorly differentiated cells including stem cells.

Augmented expression of metallothionein and glutathione S-transferase pi as unfavourable prognostic factors in cisplatin-treated ovarian cancer patients.

Resistance to cis- or carboplatin represents the principal cause of therapeutic failures in ovarian carcinoma. The phenomenon of resistance to platinum-based drugs is partly related to expression of metallothionein (MT) and of glutathione S-transferase pi (GST-pi), but opinion on the subject is discordant. Documentation of a negative predictive effect of MT and GST-pi expression for the therapy employing platinum-based drugs would permit to select resistant cases in which other therapeutic approaches could be employed. The present study aimed at examining the relation between intensities of MT and GST-pi expressions in ovarian carcinomas and dynamics of the clinical course in the neoplastic disease in a group of cisplatin-treated patients. The analyses were performed on samples of ovarian carcinoma originating from 43 first-look laparotomies (FLLs) and, in 30 cases, from second-look laparotomies (SLL) from the same patients. Immunohistochemical reactions were performed on paraffin sections of studied tumors, using monoclonal antibodies to MT and GST-pi. The calculations showed that in cases with augmented expression of MT, mortality was higher. On the other hand, augmented expression of GST-pi predisposed to more frequent relapses, deaths and progression of the tumor. Kaplan–Meier analysis showed that a significantly shorter survival time was linked to cases of higher expression of MT at FLL and of higher expression of GST-pi at FLL, whereas a shorter progression-free time was manifested by cases with higher expression of GST-pi at FLL. The performed investigations indicate that augmented expressions of MT at FLL and GST-pi at FLL in ovarian cancer represent an unfavourable predictive factor in cisplatin-treated patients.

Nuclear metallothionein expression correlates with cisplatin resistance of ovarian cancer cells and poor clinical outcome

Elevated metallothionein (MT) expression in ovarian cancers treated with cisplatin-based schemes represents an unfavorable prognostic index. MT expression is significantly higher in tumor samples obtained after chemotherapy. The present study aimed at examining MT expression in ovarian carcinoma cells sensitive (A2780) or resistant (A2780RCIS) against platinum drug treatment as well as examining effects of exposure to cisplatin on MT expression. Subcellular expression of MT was evaluated also in samples originating from 73 ovarian tumors. Cisplatin-resistant A2780RCIS cells were exposed to increasing cisplatin concentrations, and the subcellular expression of MT was determined by immunocytochemistry. The studies demonstrated that cisplatin-resistant A2780RCIS cells exposed to cisplatin typically manifested a nuclear MT expression. The study demonstrated also that exposure to cisplatin was paralleled by growing MT expression in cell nuclei. The nuclear expression of MT was also found to be specific for ovarian cancers of poor clinical outcome. No relationship could be demonstrated between cytoplasmic expression of MT and clinical variables. Nuclear MT expression is induced by cisplatin and seems to protect DNA in the cells from toxic effects of the drug.

Estrogen Receptor Alpha Expression in Ovarian Cancer Predicts Longer Overall Survival

Estrogen as a potential factor of ovarian carcinogenesis, acts via two nuclear receptors, estrogen receptor alpha (ERα) and estrogen receptor beta (ERβ), but the cellular signal pathways involved are not completely clear so far. In this study we have described the expression of ERα, detected by immunocytochemistry in 11 ovarian carcinoma cell lines and by immunohistochemistry in 43 Federation Internationale des Gyneacologistes et Obstetristes stage III ovarian carcinoma specimens prepared before and after treatment with cisplatin-based schemes. For cisplatin resistance is a major obstacle in the treatment of ovarian carcinoma, analysis of cisplatin sensitivity in 11 ovarian carcinoma cell line was also performed. The strong nuclear ERα expression was only shown in the single A2780P cell line. Expression of ERα in tissue specimens did not reveal any correlations between histopathological parameters (histologic type and grading). We demonstrated a significant association with ERα expression in specimens from primary laparotomies (PL) and cause–specific survival. In the cases terminated by death of the patient, overall immunoreactivity score of ERα expression at PL was significantly lower than in surviving patients. In addition, Kaplan-Meier analysis revealed significantly shorter overall survival time and progression-free time in cases with lower immunoreactivity score of ERα expression at PL. Our findings support the hypothesis that aberrant hormone activity, by way of altered receptor expression, might be an important factor in the malignant transformation of ovarian cancer.

Persistence of immune response to HPV-16/18 AS04-adjuvanted cervical cancer vaccine in women aged 15-55 years

The HPV-16/18 AS04-adjuvanted vaccine (Cervarix®, GlaxoSmithKline Biologicals) has been shown to induce a robust immune response in women aged 15-55 years (103514/NCT00196937). This follow-up study is the first report of persistence of immune response and safety profile through 48 months after vaccination in women aged 15-55 years. In this open-label, age-stratified Phase III study in Germany and Poland (105882/NCT00196937), healthy women aged 15-55 years received 3 doses of HPV-16/18 AS04-adjuvanted vaccine at 0, 1, and 6 months. Anti-HPV-16/18 seropositivity rates and geometric mean antibody titers (GMTs) were assessed by enzyme-linked immunosorbent assay (ELISA) in women aged 15-25 (n=168), 26-45 (n=186) and 46-55 years (n=177) from the time of first vaccination through 48 months. At Month 48, all subjects were seropositive for anti-HPV-16 antibodies and 99.4% were seropositive for anti-HPV-18. Antibody kinetics were as previously reported, with peak response at Month 7 followed by a gradual decline tending towards a plateau in all age groups. Anti-HPV-16/18 GMTs were sustained at Month 48 in all age groups, including women aged 46-55 years in whom GMTs were respectively 11-fold and 5-fold higher than natural infection levels. The vaccine exhibited a clinically acceptable safety profile in all age groups. In summary, the HPV-16/18 AS04-adjuvanted vaccine induces high and sustained immune responses in women aged 15-55 years, with antibody levels remaining several-fold higher than natural infection levels for at least 4 years after the first vaccine dose.

Cutoff value of human chorionic gonadotropin in relation to the number of methotrexate cycles in the successful treatment of ectopic pregnancy

OBJECTIVE To assign cutoff values for human chorionic gonadotropin (beta-hCG) in pretreatment and after one methotrexate (MTX) cycle and determine its correspondence to the number of MTX cycles in successfully treated ectopic pregnancy. DESIGN Retrospective study. SETTING Polish university hospital. PATIENT(S) 68 women with ectopic pregnancies who qualified for medical treatment. INTERVENTION(S) A single-dose of MTX (50 mg/m(2)) repeated every 7 days, plus laparoscopy in cases of tubal rupture or increased (>or=50% over 1 week) beta-hCG concentration. MAIN OUTCOME MEASURE(S) Resolution of serum beta-hCG without the necessity of laparoscopy. RESULT(S) Success rate was 78% (53 of 64 women). The medians of pretreatment beta-hCG levels in the groups treated successfully and unsuccessfully (943 vs. 3085 mIU/mL) and after the first dose of MTX (564 vs. 4049 mIU/mL) were statistically significantly different. The decrease in beta-hCG level after one MTX dose differed statistically significantly only in successfully treated women. The receiver operating characteristic (ROC) curve cutoff value in the success group indicated an initial beta-hCG level of 1790 and 1218 mIU/mL after one MTX cycle. The median of beta-hCG titer was not statistically different in patients requiring one or more treatment cycles. CONCLUSION(S) When the beta-hCG level is >1790 mIU/mL, the MTX treatment of ectopic pregnancy is at risk of failure. However, the initial beta-hCG titer is not a predictor of the number of MTX cycles that can guarantee a successful outcome.

Correlation between levels of human papillomavirus (HPV)-16 and 18 antibodies in serum and cervicovaginal secretions in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine

This pooled analysis of data from four Phase III clinical trials was undertaken to assess the correlation between levels of anti-human papillomavirus (HPV)-16/18 antibodies in serum and cervicovaginal secretions (CVS) in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine. Serum and CVS samples were collected from a subset of women aged 10–65 years (N=350) at pre-specified time-points from 7 to 36 months post-vaccination. Anti-HPV-16/18 antibody levels in serum and CVS were measured by enzyme-linked immunosorbent assay. Pearson correlation coefficients between serum and CVS antibody levels, standardized for total immunoglobulin G, were calculated at each time-point in women with detectable antibodies in both serum and CVS. All subjects had seroconverted at Month 7 and remained seropositive through Month 36 for both antigens. Geometric mean titers of anti-HPV-16/18 antibodies in serum were substantially higher at all time-points than those in a control group of women who had cleared a natural HPV infection in another trial. In women with detectable antibodies in both serum and CVS, good correlation was seen between HPV-16/18 antibody levels at all time-points (Pearson correlation coefficient: 0.84–0.92 for HPV-16 and 0.90–0.91 for HPV-18). The strong correlation between levels of HPV-16/18 antibodies in serum and CVS up to 36 months post-vaccination in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine supports transudation of serum antibodies as the mechanism by which antibodies are introduced into CVS. These CVS antibodies may play a role in the protective efficacy of this vaccine.

Continuous, daily levonorgestrel/ethinyl estradiol vs. 21-day, cyclic levonorgestrel/ethinyl estradiol: efficacy, safety and bleeding in a randomized, open-label trial☆

BACKGROUND This Phase 3, randomized, open-label, multicenter study conducted at 44 sites in Europe evaluated the safety and efficacy of a continuous, daily regimen of levonorgestrel (LNG) 90 mcg/ethinyl estradiol (EE) 20 mcg compared with a 21-day, cyclic LNG 100 mcg/EE 20 mcg regimen. STUDY DESIGN Three hundred twenty-three healthy women were randomized to continuous LNG 90 mcg/EE 20 mcg and 318 subjects to cyclic LNG 100 mcg/EE 20 mcg for 1 year (13 pill packs). Pearl index, adverse event (AE) incidence and bleeding profiles were assessed. RESULTS No pregnancies occurred with the continuous oral contraceptive (OC) (Pearl index=0.00). As the study progressed, the percentage of women who achieved amenorrhea during each 28-day pill pack increased: 40% at pill pack 7, 53% at pill pack 13. The percentage of women with no bleeding [with or without spotting (defined as not requiring sanitary protection)] was 50%, 69% and 79% at pill packs 3, 7 and 13, respectively. The incidence of AEs was similar to that of the cyclic OC (except for metrorrhagia and vaginal bleeding in the first 6 months). CONCLUSIONS Continuous LNG 90 mcg/EE 20 mcg was shown to be a safe and effective OC in this direct comparison to a cyclic OC. Suppression of menses and the potential for no bleeding requiring sanitary protection may be provided by this continuous, low-dose OC.