Resumption of Trastuzumab in Patients With Disease Recurrence After (Neo-) Adjuvant Anti-HER2-therapy in Patients With HER2-positive Breast Cancer.

BACKGROUND/AIM: HER2-positive breast cancers eventually relapse in about one third of patients. Is anti-HER2-directed therapy with Herceptin® (trastuzumab) effective in re-treatment? Between 2008 and 2018, 216 patients with recurrent HER2-positive breast cancer (BC) were re-treated with Herceptin (HER) during first-line therapy. This study assessed the effectiveness and tolerability of re-treatment with HER. PATIENTS AND METHODS: After approval from Ethical committee, the NIS was conducted according to German Drug Act. Re-treatment with HER was documented at routine visits starting with a basic observational period of maximum 12 months and a follow-up period of maximum additional four years. RESULTS: HER2-positive BC relapsed after a median of 36.5 months (mos). Patients were re-treated with HER +/- chemotherapy +/- endocrine therapy. HER-containing regimens resulted in median progression-free survival (mPFS) of 12.7 (95%CI=10.5-14.8) mos and overall survival (OS-2) of 31.6 mos (95%CI=28.8-38.4) since recurrence diagnosis. Differentiation of recurrence types (local, visceral, non-visceral) unfolded worst prognosis for patients with visceral metastases. Cardiac monitoring within this non-interventional study (NIS) did not result in new safety concerns. CONCLUSION: Re-therapy with HER in the first-line setting of advanced HER2-positive breast cancer is effective and without unexpected or intensified adverse events.

Primary metastatic breast cancer in the era of targeted therapy - Prognostic impact and the role of breast tumour surgery.

BACKGROUND: Except for meeting the individual palliative need, the benefit of breast surgery in primary metastatic breast cancer (PMBC), also known as de novo metastatic breast cancer, on long-term outcomes remains controversial. Twenty-four hundred and one patients with metastatic breast cancer, enrolled between 2000 and 2011 in two prospective non-interventional studies on targeted therapy, were screened with respect to this question. METHODS: One study investigated trastuzumab therapy for human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer in addition to mainly first-line chemotherapy. The other observed bevacizumab added to chemotherapy as first-line treatment for mostly HER2-negative disease. RESULTS: Five-hundred and seventy (24%) patients presented with PMBC, and valid information on resection of the primary tumour was available for 568 women. Out of these, 426 (75%) underwent local resection. The latter group was characterised by less overall metastatic burden and a lower proportion of T4 tumours. No major differences were observed with respect to age, hormone receptor and HER2 status, visceral disease and performance status. Numerically, the surgery group showed a slightly favourable progression-free survival (PFS, medians: 13.6 versus 11.8 months; P = 0.18) and overall survival (OS, 34.1 versus 31.7; P = 0.23). However, in multivariable analysis, including all other univariably significant parameters, no trend for better outcome after surgery remained detectable, neither for PFS (hazard ratio 0.99; P = 0.92) nor for OS (0.95; P = 0.71). CONCLUSIONS: Our findings suggest no major survival benefit for local resection in the overall PMBC population treated with modern targeted therapies. However, further analyses are warranted to define specific risk groups, which may benefit from surgical removal of the primary.

Polymorphisms in the promoter region of ESR2 gene and susceptibility to ovarian cancer.

Susceptibility to ovarian cancer might be affected by genetic variations in genes involved in estrogen biosynthesis, metabolism or signal transduction. In this study we tested the hypothesis that single nucleotide polymorphisms (SNPs) in the promoter of human ESR2 gene, coding for estrogen receptor ß, may be associated with increased risk for ovarian cancer. Three SNPs in the promoter region of human ESR2 gene were genotyped by means of allele-specific tetra-primer PCR. A total of 184 ovarian cancer cases and the same numbers of controls were included in the study. With regard to homozygous analysis, the AA genotype of SNP rs3020449 was found to be significantly more frequent in ovarian cancer cases staged as FIGO III+IV than in cases staged as I+II (OR 2.717, p=0.027). With regard to allele frequency, the G allele of this SNP was less frequent in FIGO I+II cases than in cases with higher FIGO stages (OR 1.739, p=0.018). With regard to genotype frequency, allele frequency, allele positivity or haplotype frequency of SNPs rs2987983, rs3020449 and rs3020450 we did not observe a significant difference between the cancer and the control group. Our data suggest that SNPs in the promoter region of ESR2 gene do not affect susceptibility to ovarian cancer, but SNP rs3020449 might affect progression of this disease.

Icb-1 gene polymorphism rs1467465 is associated with susceptibility to ovarian cancer.

In this study, we tested the hypothesis that single nucleotide polymorphisms (SNPs) of differentiation-associated human gene icb-1 (C1orf38) may be associated with ovarian cancer susceptibility. For this purpose, we compared the genotype and allele frequencies of the SNPs rs1467465 and rs12048235 in a group of 184 ovarian cancer patients with a control group of 184 age- and gender-matched women without any malignancy. Genotype-phenotype association revealed that A allele of SNP rs1467465 was more frequent in ovarian cancer patients than in the control group (0.40 vs. 0.33, OR 1.37, 95% CI 1.013-1.853, p = 0.04). After analysis of allele positivity we observed that A-positive genotypes were more frequent in the ovarian cancer group (0.65 vs. 0.53, OR 1.63, 95% CI 1.072-2.483, p = 0.02). Furthermore, the heterozygous genotype of rs1467465 was found to be more frequent in the patients group (0.50 vs. 0.41, OR 1.63, 95% CI 1.045-2.045, p = 0.03). No significant results were obtained with regard to SNP rs1204823. Our data suggest, that SNP rs1467465 of human gene icb-1 might affect susceptibility to ovarian cancer.

Effects of estriol on growth, gene expression and estrogen response element activation in human breast cancer cell lines.

OBJECTIVE: Local application of estradiol (E2) to treat vulvovaginal atrophy in postmenopausal breast cancer patients receiving aromatase inhibitors is known to elevate serum estradiol levels and thereby might counteract breast cancer therapy. Thus, vaginal application of estriol (E3) has been recommended for these patients. However, it is unclear to what extent E3 stimulates breast cancer cell growth. In this study, we examined the effect of E3 on growth and gene expression of two human breast cancer cell lines. METHODS: We used an established in vitro cell culture assay and compared the effect of E2 and E3 on growth of the estrogen receptor alpha-positive breast cancer cell lines MCF-7 and T-47D testing a wide range of hormone concentrations of 10(-12)-10(-7)M. E3 effects on gene expression were examined by means of reporter gene assays, RT-qPCR and Western blot analysis. RESULTS: E3 acted as a potent estrogen and exerted a mitogenic effect on T-47D and MCF-7 cells at concentrations of 10(-9)M (288 pg/ml) and higher. With regard to activation of an estrogen response element (ERE) in breast cancer cells, effects of E3 were visible at 10(-10)M. The same concentrations of E3 activated expression of the estrogen-responsive gene PR and of the proliferation genes cyclin A2, cyclin B1, Ki-67, c-myc and b-myb, providing molecular mechanisms underlying the observed growth increase. CONCLUSIONS: Like E2, low levels of E3 were able to trigger a robust estrogenic response in breast cancer cells. Thus, our data suggest caution regarding use of E3 by breast cancer survivors.

Polymorphisms in the promoter region of estrogen receptor ß gene in endometrial cancer.

INTRODUCTION: The development of endometrial cancer is known to be affected by estrogens. Thus, genetic variations like single nucleotide polymorphisms (SNPs) in genes involved in estrogen biosynthesis, metabolism, and signal transduction might affect risk for endometrial cancer. In this study, we tested the hypothesis that polymorphisms in the promoter of ESR2 gene may be associated with susceptibility to this disease. METHODS: We compared the frequency of three SNPs in the promoter region of ESR2 gene (rs2987983, rs3020450, and rs3020449) in 135 women with endometrial cancer and 135 healthy women serving as controls by means of allele-specific tetra-primer PCR. RESULTS: Regarding allele frequency, allele positivity or genotype frequencies of these SNPs we did not observe any significant difference between healthy women and women with endometrial cancer. CONCLUSION: Our data clearly suggest that the tested SNPs in the promotor region of human ESR2 gene are not associated with the development of endometrial cancer.

Effects of a combined treatment with tamoxifen and estrogen receptor ß agonists on human breast cancer cell lines.

INTRODUCTION: Coexpression of estrogen receptors (ER) α and ß is present in about half of all breast cancer cases. Whereas ERα is a well-established target for endocrine therapy with the selective estrogen receptor modulator tamoxifen, the applicability of ERß as target in breast cancer therapy is unclear. In this study, we examined the effects of two synthetic ERß agonists alone and in combination with tamoxifen on ERα/ß-positive breast cancer cells. METHODS: We treated MCF-7 and T-47D breast cancer cells with the ERß agonists ERB-041 and WAY-200070 and measured the effects on cell growth. In addition, transcriptome analyses were performed by means of Affymetrix GeneChip arrays. RESULTS: When given alone, ERß agonists ERB-041 and WAY-200070 did not affect the growth of MCF-7 or T-47D cells. In contrast, addition of these drugs to tamoxifen increased its growth-inhibitory effect on both cell lines. This effect was more pronounced under serum-free conditions, but was also observed in the presence of serum in T-47D cells. Transcriptome analyses revealed a set of genes regulated after addition of ERß agonists including S100A8 and CD177. CONCLUSION: The observed enhanced growth-inhibitory effects of a combination of tamoxifen and ERß agonists in vitro encourage further studies to test its possible use in the clinical setting.

Expression of SCUBE2 gene declines in high grade endometrial cancer and associates with expression of steroid hormone receptors and tumor suppressor PTEN.

SCUBE2 (Signal peptide-CUB-epidermal growth factor-like domain-containing 2) gene codes for a cell-surface glycoprotein. In breast cancer, SCUBE2 transcript levels are part of important prognostic and predictive gene signatures and are linked to expression of estrogen receptor α (ERα). To elucidate the role of this gene in endometrial cancer, we compared SCUBE2 expression in malignant and normal endometrial tissue specimens. We then examined its correlation with steroid hormone receptors and PTEN and compared it to SCUBE2 expression in breast cancer samples. Expression of SCUBE2 was found to be decreased in G3 endometrial cancer when compared to postmenopausal endometrium or to G1 tumors (p < 0.05). In postmenopausal endometrium, SCUBE2 transcript levels were more than twice as high as in premenopausal women. In breast cancer, SCUBE2 expression was found to be notably reduced particularly in ERα-negative G3 tumors. Both in endometrial and breast cancer we observed a significant positive correlation of SCUBE2 transcript levels with expression of ERα, PR and PTEN. Our data suggest that SCUBE2, like in breast cancer, associates with ERα and might have a potential as prognostic or predictive marker in endometrial cancer.

G protein-coupled estrogen receptor (GPER) expression in endometrial adenocarcinoma and effect of agonist G-1 on growth of endometrial adenocarcinoma cell lines.

The G protein-coupled estrogen receptor (GPER, GPR30) is suggested to be involved in non-nuclear estrogen signaling and is expressed in a variety of hormone dependent cancer entities. This study was performed to further elucidate the role of this receptor in endometrial adenocarcinoma. We first analyzed GPER expression at the mRNA level in 88 endometrial cancer or normal endometrial tissue samples and compared it to those of nuclear steroid hormone receptors. GPER transcript levels were found to be about 6-fold reduced, but still present in endometrial cancer. Expression of this receptor was decreased in all grading subgroups when compared to pre- or postmenopausal endometrium. GPER mRNA expression was associated with PR mRNA levels (Spearman's rho 0.4610, p<0.001). We then tested the effect of the GPER ligand G-1 on growth of three endometrial cancer cell lines with different GPER expression. GPER protein levels were highest in RL95-2 cells, moderate in HEC-1A cells and not detectable in HEC-1B cells. The moderate expression level in HEC-1A cells was similar to average tumor tissue expression. Treatment with G-1 significantly inhibited growth of the GPER-positive cell lines RL95-2 and HEC-1A in a dose-dependent manner, whereas the GPER-negative line HEC-1B was not affected. Though GPER transcript levels were found to be reduced in endometrial cancer, our in vitro data suggest that moderate GPER expression might be sufficient to mediate growth-inhibitory effects triggered by its agonist G-1.

icb-1 Gene counteracts growth of ovarian cancer cell lines.

Human gene icb-1 has been originally identified to be involved in differentiation processes of cancer cells. To examine the function of icb-1 in ovarian cancer, we knocked down its expression in three ovarian cancer cell lines and performed microarray-based gene expression profiling with subsequent gene network modeling. Loss of icb-1 expression accelerated proliferation of SK-OV-3, OVCAR-3 and OAW-42 cells and led to upregulation of ovarian cancer biomarkers like KLK10 and CLDN16. Most of the upregulated genes were part of oncogenic pathways regulated by ERα or TNF. Our data suggest that icb-1 gene inhibits growth and progression of ovarian cancer cells.

Estrogen receptor ß agonists affect growth and gene expression of human breast cancer cell lines.

Expression of estrogen receptor ß (ERß) has been described to reduce growth of cancer cell lines derived from hormone-dependent tumors, like breast cancer. In this study we tested to what extent two ERß agonists, androgen derivative 3ß-Adiol and flavonoid Liquiritigenin, would affect growth and gene expression of different ERß-positive human breast cancer cell lines. Under standard cell culture conditions, we observed 3ß-Adiol to inhibit growth of MCF-7 cells in a dose-dependent manner, whereas growth of BT-474 and MCF-10A cells was suppressed by the maximum concentration (100 nM) only. When treated in serum-free medium, all cell lines except of MDA-MB-231 were responsive to 1 nM 3ß-Adiol, and ZR75-1 cells exhibited a dose-dependent antiproliferative response. Providing putative mechanisms underlying the observed growth-inhibitory effect, expression of Ki-67 or cyclins A2 and B1 was downregulated after 3ß-Adiol treatment in all responsive lines. In contrast, treatment with lower doses of Liquiritigenin did not affect growth. In MCF-7 cells, the highest dose of this flavonoid exerted proliferative effects accompanied by increased expression of cyclin B1, PR and PS2, indicating unspecific activation of ERα. In conclusion, the ERß agonists tested exerted distinct concentration-dependent and cell line-specific effects on growth and gene expression. The observed inhibitory effects of 3ß-Adiol on breast cancer cell growth encourage further studies on the potential of this and other ERß agonists as targeted drugs for breast cancer therapy.

Role of estrogen receptor ß in gynecological cancer.

Estrogen receptor ß is expressed in normal and neoplastic ovarian and endometrial tissues. Recent studies indicate that levels of this receptor decline in ovarian tumorigenesis, like in breast or prostate cancer. Furthermore, ERß expression has been associated with good prognosis in ovarian cancer. In contrast, previous studies on the role of this receptor in endometrial cancer suggested that ERß might play different roles in the carcinogenesis of the ovary and endometrium. Besides its possible role as a prognostic factor, ERß might be a potential target for the treatment of ovarian and endometrial cancer.

Single nucleotide polymorphisms in the regulatory region of gonadotropin-releasing hormone receptor gene and breast cancer susceptibility.

It is known that exposure to estrogens affects the pathophysiology of breast cancer. The key role of gonadotropin-releasing hormone (GnRH) in the regulation of female steroid hormone metabolism raises the question of whether polymorphisms in its receptor, GnRHR, might influence breast cancer risk. To test this hypothesis, we analyzed three single nucleotide polymorphisms (SNPs) in the 5'-regulatory region of the GnRHR gene in a total of 565 women, 254 women with breast cancer and 311 women without any malignancy by allele-specific PCR. No significant differences were observed between the breast cancer and control group in terms of genotype, allele frequency or allele positivity. In contrast, different frequencies of the SNPs rs13138607, rs12644822 and rs3756159 were observed after sub-grouping the breast cancer cases according to tumor grading. Our data suggest a potential role of GnRHR gene polymorphisms in the development of breast cancer.

A polymorphism in the nuclear receptor coactivator 7 gene and breast cancer susceptibility.

The nuclear receptor coactivator 7 (NCoA7) gene codes for an estrogen receptor-associated protein that plays a significant role in the cellular response to estrogens. Given that NCoA7 is expressed in the mammary gland, alterations in this gene may affect breast cancer risk. In this study, we compared the genotype and allele frequencies of the missense single nucleotide polymorphism (SNP) rs1567, located in the coding region of the NCoA7 gene and resulting in an amino acid exchange from asparagine to glutamine, in 305 women with sporadic breast cancer and 346 women without any malignancy. Statistical analysis of the observed frequencies did not reveal a significant difference between the cancer and control groups, nor did a comparison between histological breast cancer subgroups. In conclusion, the results of our phenotype-genotype association study indicate that NCoA7 SNP rs1567 does not affect breast cancer susceptibility.

The treatment of climacteric symptoms.

BACKGROUND: Peri- and postmenopausal women commonly suffer from climacteric symptoms. In this article, we provide information to help physicians recognize climacteric symptoms and treat them appropriately. METHODS: The information presented here is based on a selective search of the literature for pertinent articles that appeared from 2008 to early 2011, including the German S3 guideline on hormone therapy (HT) during and after menopause, which was published in 2009. RESULTS: Perimenopausal women often suffer from climacteric symptoms. Typically, women undergoing menopause complain of heat waves and vaginal dryness. According to randomized controlled trials as well as national and international guidelines, HT is the most effective treatment for vasomotor symptoms and also improves vulvovaginal atrophy; for the latter indication, HT is preferably administered locally. Vaginal estrogen therapy lowers the frequency of recurrent urinary tract infections. However, HT is associated with an increased risk for a number of diseases, including stroke, thromboembolic events, gall-bladder diseases, and breast cancer. Alternative treatments for climacteric symptoms have little or no efficacy. CONCLUSION: HT should only be used to treat climacteric symptoms after extensive patient education about its benefits and risks. Participatory decision-making is desirable. The generalized use of HT by all women with climacteric symptoms cannot be recommended.

Network analysis of icb-1 gene function in human breast cancer cells.

Icb-1 is a human gene previously described by our group to exert important functions in cancer cells of different origin. We now performed microarray-based gene expression profiling with subsequent network modeling to further elucidate the role of icb-1 in breast cancer cells. Analyzing the effect of icb-1 knockdown on the transcriptome of MCF-7 cells, we found 151 differentially expressed genes exhibiting more than twofold changes, 97 of which were up- and 54 downregulated. Most of the upregulated genes were cancer-related genes associated with poor prognosis, invasion and metastasis, building an oncogenic network of TNF target genes. On the other hand, network analysis identified the downregulated genes to be primarily involved in interferon signaling and cellular apoptosis. Confirming these network data, we observed that cells with reduced levels of icb-1 exhibited an impaired response to the apoptosis inducers tamoxifen, staurosporine, actinomycin, and camptothecin. The data of this study suggest that icb-1 might exert a tumor-suppressor function in breast cancer and that its loss might confer relative resistance of breast cancer cells to apoptotic drugs.

Effects of a combined treatment with GPR30 agonist G-1 and herceptin on growth and gene expression of human breast cancer cell lines.

Expression of G-protein-coupled receptor 30 (GPR30) is present in HER2-overexpressing breast cancer. In this study, we examined to what extent GPR30-agonist G-1 would affect the antitumoral action of trastuzumab (Herceptin). Combined treatment with both drugs exerted an additive growth-inhibitory effect on breast cancer cells which was accompanied by a significant decline of cyclin A2 expression both on the protein and the mRNA level. Combined treatment also resulted in expression changes of c-fos, cyclin D1, or p21/WAF-1. The results of our study encourage further attempts to test the relevance of these in vitro data in the clinical setting.

Estrogen receptor ß transcript variants associate with oncogene expression in endometrial cancer.

The human ESR2 gene codes for estrogen receptor ß1 and for multiple splice variants, which are suggested to exert distinct functions in the cellular estrogen response. Given that the function of ERß in endometrial cancer remains unclear, we examined the expression of ERß1, ERß2 and various further ERß transcript variants and their association with selected cancer-related genes in 74 human endometrium samples and endometrial cancer specimens by means of RT-qPCR. Additionally, we knocked down ERß expression in HEC-1A endometrial adenocarcinoma cells by means of siRNA transfection. Expression of four ERß transcript variants was significantly elevated in cancer tissue or in G3 tumors compared to postmenopausal endometrium. Expression of ERß1, ERß2, ERß5 and five further variants was associated with the oncogenes MYBL2 or HER2 in endometrial cancer. In addition, siRNA-triggered knockdown of ERß expression led to a significant decline of MYBL2 mRNA and protein levels in endometrial cancer cells. Our observation of increased ERß transcript levels in cancer tissue and particularly their correlation with the expression of oncogenes, as well as the results of our knockdown studies, suggest a role of ERß in endometrial carcinogenesis.

Expression of cysteine protease cathepsin L is increased in endometrial cancer and correlates with expression of growth regulatory genes.

Proteases contribute to tumor invasion and metastasis by degrading basement membranes and extracellular matrix (ECM). In this study, we compared gene expression levels of two proteases, cysteine protease Cathepsin L2 (CTSL2) and matrix metalloproteinase MMP11, in human endometrium and endometrial cancer. Our data demonstrate CTSL2 transcript levels to be strongly elevated in endometrial cancer, particularly in G3 tumors. Furthermore, we observed a highly significant positive correlation of CTSL2 with expression of growth regulatory genes Ki-67, cyclin B1, MYBL2, p21/WAF, and HER2 receptor tyrosine kinase. Our data suggest that CTSL2 might be involved in progression of endometrial cancer.

Silencing of the icb-1 gene inhibits the induction of differentiation-associated genes by vitamin D3 and all-trans retinoic acid in gynecological cancer cells.

Icb-1 (C1orf38) is a human gene initially described by our group to be involved in differentiation processes of cancer cells. To further elucidate the function of the icb-1 gene in differentiation of breast and endometrial cancer cells, we knocked down its expression by means of shRNA transfection. Knockdown of icb-1 inhibited the vitamin D3-induced up-regulation of E-cadherin expression in both MCF-7 and HEC-1B cells. Induction of E-cadherin expression by all-trans retinoic acid (ATRA) was also blocked in both cell lines expressing icb-1 siRNA. Examination of icb-1 and E-cadherin expression in 66 breast cancer tissue samples revealed a significant positive correlation between the two genes. In MCF-7 cells, silencing of the icb-1 gene inhibited the ATRA- and the vitamin D3-induced up-regulation of lactoferrin and estrogen receptor ß expression. The data of our knockdown study suggest that icb-1 may act as a mediator of differentiation signals in breast cancer cells induced by ATRA or vitamin D3. These findings together with the observed co-expression of icb-1 with E-cadherin in breast cancer samples support an important role of the icb-1 gene in cancer cell differentiation.