Differences in Analgesia Methods for Open Gastrointestinal Surgery Are Not Associated With Initial Postoperative Ambulation.

Background: A characteristic of modern medical care is the reduction in the length of hospital stay, and several facilities across Japan are working towards this goal. The presence of postoperative pain is correlated with the number of days to hospital discharge. Therefore, this study investigated the relationship between the analgesic methods used in clinical practice and the initial ambulation of postoperative laparotomy patients with severe postoperative worked incisional pain to enable better analgesic management in the future. Methods: This retrospective study collected information from the medical records of 117 patients who underwent laparotomy between December 1, 2019, and October 13, 2020, at the Department of Gastroenterology of the International University of Health and Welfare Mita Hospital. Based on the failure or success of the ambulation process, the patients were divided into the delayed and successful groups, respectively. Results: In the delayed group, patient-controlled epidural analgesia (PCEA) was used in 32 patients, intravenous patient-controlled analgesia (IV-PCA) was used in two patients, continuous worked incisional infiltration anesthesia was used in one patient, and transvenous acetaminophen was used in one patient for postoperative analgesia. In the successful group, PCEA was used in 66 patients, IV-PCA was used in 11 patients, continuous worked incisional infiltration anesthesia was used in three patients, and acetaminophen administered intravenously at patient's request was used in one patient (P = 0.094). Conclusions: No significant differences were observed between different postoperative analgesia methods, suggesting that there may be no association between postoperative ambulation and the postoperative analgesia method.

Predictors of Catheter-Related Bladder Discomfort After Surgery: A Literature Review.

Background: Indwelling bladder catheters are routinely used in clinical practice. Patients may experience postoperative indwelling catheter-related bladder discomfort (CRBD). This study aimed to perform a literature review to identify predictors of postoperative CRBD. Methods: We searched PubMed for relevant articles published between 2000 and 2020 using the search items "CRBD", "catheter-related bladder discomfort", and "prediction". Additionally, we searched for articles that matched the research objectives from the references of the extracted articles. We included only prospective observational studies involving human participants and excluded interventional studies, observational studies that did not report sample sizes, or observational studies that did not research on predictors of CRBD. We narrowed our search to the keyword "prediction" and found five references. We selected five studies that met the objectives of the study as the target literature. Results: Using the keywords "CRBD" and "catheter-related bladder discomfort", we identified 69 published articles. The results were narrowed down by the keyword "prediction", and five studies that recruited 1,147 patients remained. The predictors of CRBD can be divided into four factors: 1) patient factors; 2) surgical factors; 3) anesthesia factors; and 4) device and insertion technique factors. Conclusion: Our study suggests that patients with predictors of CRBD should be closely monitored to reduce postoperative patient suffering, and their quality of life should be improved after anesthesia.

A Literature Review of Factors Related to Postoperative Sore Throat.

Postoperative sore throat can occur as a complication in patients who have undergone surgery under general anesthesia. The incidence of postoperative sore throat ranges from 12.1% to 70%, and its effects include damage to the epithelium and mucosal cells caused by airway securement, damage to the vocal cords, congestion, blood clots, and factors such as an inappropriately large tube, cuff shape, cuff pressure, and airway securement. Notably, there are individual differences in pain thresholds, and the sensation of pain is affected by mental states, such as anxiety, and varies from person to person. Therefore, we conducted a literature review using PubMed to clarify patient factors related to the development of postoperative sore throat. The extracted keywords were "postoperative sore throat," "anesthesia," and "patient factors." We found 16 articles that met our search criteria. We expanded the search period and retrieved 19 cases from 1990 to 2020. We also included references that were judged to be closely related to the list of citations of the retrieved references. The study designs included were randomized controlled trials, clinical trials, meta-analyses, reviews, and systematic reviews. The results showed that female sex, smoking, and age were the most common patient factors. However, we could not find any literature that studied the relationship between postoperative sore throat and mental states such as anxiety.

One DNA Methylation Regulates CHIP Gene Expression of Human Breast Cancer and Predicts Recurrence.

BACKGROUND/AIM: Carboxyl terminus of Hsc70-interacting protein (CHIP) is a ubiquitin ligase that induces ubiquitination and degradation of its target proteins including oncoproteins. We reported that its down-regulation is associated with tumor progression and metastasis of breast cancer. However, the mechanism through which CHIP gene affects cancer cells is unclear. MATERIALS AND METHODS: We extracted RNA from 45 primary breast cancer samples and compared CHIP mRNA expression profiles, promoter DNA methylation status, and clinicopathological information. RESULTS: CHIP mRNA expression was significantly correlated with the tumor progression status. In several samples, a pinpoint CpG methylation in the CHIP gene promoter region was significantly correlated with CHIP mRNA expression. When this specific CpG was methylated in estrogen receptor (ER)-positive cases, a significant difference in 5-year recurrence was not found compared with ER-negative cases. CONCLUSION: CpG methylation contributes to the long-term prognosis of ER-positive breast cancer.

Prostate-specific antigen nomogram to predict advanced prostate cancer using area under the receiver operating characteristic curve boosting.

BACKGROUND: For prostate cancer, accurate prediction of the pathological stage before surgery is very important. Therefore, the aim of the present study was establishing the prostate-specific antigen (PSA) threshold nomogram to predict pathologically advanced prostate cancer using the novel method of area under the receiver operating characteristic curve boosting (AUCBoost). METHODS: The medical records of patients with clinically localized prostate cancer who underwent robot-assisted radical prostatectomy were retrospectively reviewed. Multivariate logistic regression analysis was performed to identify clinical covariates significantly associated with pathological tumor stage ≥3a. The best combination of the variables was determined by validated values of the area under the curve (AUC). The optimal individualized PSA threshold values were developed using AUCBoost. RESULTS: In the multivariate logistic regression analysis, PSA, prostate volume, clinical tumor stage, Gleason Grade Group, the number of positive cores, and the percentage of positive cores were independent predictive factors for pathological tumor stage ≥3a. A combination model comprising PSA, prostate volume, clinical tumor stage, percent positive core, and Gleason Grade Group produced the highest AUC for predicting pathological tumor stage ≥3a (AUC = 0.777). The PSA threshold values for detecting pathological tumor stage ≥3a were calculated and a table of individualized PSA threshold nomogram was developed using AUCBoost. CONCLUSIONS: We developed a nomogram of the PSA threshold values for predicting adverse pathological tumor stages of prostate cancer using a novel statistical method. Further validation is necessary; however, the individualized PSA threshold nomogram may be useful in determining treatment strategies before surgery.

Predicting factors for progression to castration resistance prostate cancer after biochemical recurrence in patients with clinically localized prostate cancer who underwent radical prostatectomy.

BACKGROUND: To determine prognostic factors associated with progression to castration-resistant prostate cancer following biochemical recurrence which is lethal prostate cancer and establish a risk stratification model of progression to castration-resistant prostate cancer. METHODS: We retrospectively reviewed the data of 550 patients who experienced biochemical recurrence after radical prostatectomy. The endpoint of the present study was progression to castration-resistant prostate cancer. The actuarial probabilities of progression to castration-resistant prostate cancer-free survival were determined using Kaplan-Meier analysis. Univariate and multivariate Cox proportional hazards regression analyses were used to identify independent predictors of biochemical recurrence. RESULTS: Fifty-two patients experienced progression to castration-resistant prostate cancer during the follow-up period. The progression to castration-resistant prostate cancer-free survival rate after biochemical recurrence at 10 years was 76.8%. In multivariate analysis, pathological Gleason score ≥ 9, lymphovascular invasion, and prostate-specific antigen velocity ≥ 0.4 ng/mL/year were independent predictive factors for progression to castration-resistant prostate cancer. The patients were stratified into three groups using a risk stratification model incorporating these variables. The 10-year progression to castration-resistant prostate cancer-free survival rates were 96.7% in the low-risk group, 84.7% in the intermediate-risk group, and 24.5% in the high-risk group. CONCLUSIONS: The present results suggest that the pathological Gleason score, lymphovascular invasion, and prostate-specific antigen velocity were independent predictive factors for progression to castration-resistant prostate cancer. The risk stratification model established in the present study could be useful for patient counseling and in identifying patients with a poor prognosis.

Heregulin controls ERα and HER2 signaling in mammospheres of ERα-positive breast cancer cells and interferes with the efficacy of molecular targeted therapy.

Estrogen receptor (ER)α and the human epidermal growth factor receptor (HER) family are inversely expressed in ERα-positive cancer in association with resistance to hormonal therapy, but the mechanism underlying their relationship remains unknown. We analyzed the effect of HER family ligands on the expression of ER and the HER family in ERα-positive MCF-7 and T47D breast cancer cell lines in 3D spheroid culture. Here, we demonstrated for the first time that heregulin-1ß (HRG), a HER3 and HER4 ligand, most effectively regulated ER/HER family expression by decreasing ERα mRNA expression and increasing HER family mRNA expression. HRG treatment attenuated fulvestrant-mediated growth inhibition, and promoted the migration of MCF-7 cells. Moreover, HRG increased the CD44+/CD24- cell fraction and side population cells, both of which are recognized as prospective breast cancer stem cell markers. HRG activated both phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) and mitogen-activated protein kinase (MAPK) pathways. Inhibitors of these pathways reduced the growth of MCF-7 cells, but the addition of HRG has different effects on these pathways. HRG blocked the inhibitory effect of mTOR inhibitors, such as rapamycin and everolimus, on cell growth but not that of a PI3K inhibitor. Furthermore, HRG slightly decreased the inhibitory effect of an AKT inhibitor on cell growth. In contrast, HRG enhanced the MEK inhibitor-induced inhibition of cell growth. These findings suggest that HRG-stimulated signaling pathways allow ERα-positive breast cancer cells to escape from growth inhibition caused by everolimus, via MAPK signaling and/or other signaling pathways. Everolimus improves progression-free survival in combination with exemestane as second-line therapy for metastatic hormone receptor-positive breast cancer. Our study suggests that HRG is a novel target for ERα-positive breast cancer therapy.

Characterization of a membrane-associated estrogen receptor in breast cancer cells and its contribution to hormone therapy resistance using a novel selective ligand.

The estrogen receptor (ER) plays a role in the progression of hormone-dependent breast cancer and is a hormone therapy target. Estrogen acts as a transcription factor (genomic action) and also produces a quick non-genomic reaction through intracellular signaling pathways. The membrane associated ER (mER) may regulate both these signals and hormone therapy resistance. However, the details remain unclear because a reliable method to distinguish the signals induced by the estradiol (E2)-mER and E2-nuclear ER complex has not been established. In the present study, we prepared the novel ligand Qdot-6-E2, selective for mER, by coupling E2 with insoluble quantum dot nano-beads. We investigated the characteristics of mER signaling pathways and its contribution to hormone therapy resistance using different cell lines including estrogen depletion resistant (EDR) cells with different mechanisms. Qdot-6-E2 stimulated proliferation of nuclear ER-positive cells, but nuclear ER-negative cells showed no response. In addition, Qdot-6-E2 indirectly activated nuclear ER and increased mRNA expression of target genes. We confirmed that E2 was not dissociated from Qdot-6-E2 using a mammalian one-hybrid assay. We visually demonstrated that Qdot-6-E2 acts from the outside of cells. The gene expression profile induced by Qdot-6-E2-mER was different from that induced by E2-nuclear ER. The effect of anti-ER antibody, the GFP-ER fusion protein localization, and the effect of palmitoyl acyltransferase inhibitor also indicated the existence of mER. Regarding intracellular phosphorylation signaling pathways, the MAPK (Erk 1/2) and the PI3K/Akt pathways were both activated by Qdot-6-E2. In EDR cells, only nuclear ER-positive cells showed increased cell proliferation with increased localization of ERα to the membrane fraction. These findings suggested that Qdot-6-E2 reacts with ERα surrounding the cell membrane and that mER signals help the cells to survive under estrogen-depleted conditions by re-localizing the ER to use trace amounts of E2 more effectively. We expect that Qdot-6-E2 is a useful tool for studying the mER.

The significance of micro-lymphatic invasion and pathological Gleason score in prostate cancer patients with pathologically organ-confined disease and negative surgical margins after robot-assisted radical prostatectomy.

BACKGROUND: The development process of recurrence in prostate cancer patients with pathologically organ-confined (pT2) disease and negative surgical margins is unclear. The aim of the present study was to determine factors associated with the development of biochemical recurrence following robot-assisted radical prostatectomy among those prostate cancer patients. METHODS: We retrospectively reviewed the data of patients who underwent robot-assisted radical prostatectomy without neoadjuvant endocrine therapy. We evaluated prognostic factors in 1096 prostate cancer patients with pT2 disease and negative surgical margins. Univariate and multivariate Cox proportional hazards regression analyses were used to identify independent predictors for biochemical recurrence. RESULTS: Of the 1096 patients, 55 experienced biochemical recurrence during the follow-up period. The 5-year biochemical recurrence-free survival rate for patients with pT2 and negative surgical margins was 91.8%. On univariate analysis, clinical stage, biopsy Gleason score, percent of positive core, pathological Gleason score, and the presence of micro-lymphatic invasion were significantly associated with biochemical recurrence. On a multivariate analysis, the presence of micro-lymphatic invasion and a pathological Gleason score ≥ 4 + 3 were significant prognostic factors for biochemical recurrence. Based on these factors, we developed a risk stratification model. The biochemical recurrence-free survival rate differed significantly among the risk groups. CONCLUSIONS: The prognosis of prostate cancer patients with pT2 disease and negative surgical margins is favorable. However, patients with the presence of micro-lymphatic invasion and a pathological Gleason score ≥ 4 + 3 tend to experience biochemical recurrence more often after surgery. Therefore, careful follow-up might be necessary for those patients.

Tumor microenvironmental growth factors induce long-term estrogen deprivation resistance in breast cancer.

BACKGROUND: Hormonal therapy is an effective treatment for luminal-like breast cancer. Aromatase inhibitor (AI) is widely used for estrogen receptor-positive, postmenopausal breast cancers. However, resistance is occurred and becomes a serious clinical concern. In general, progression of cancer strongly depends on tumor microenvironment, which may, therefore, also contribute to the development of AI resistance. METHODS: We evaluated tumor microenvironment-derived factors with respect to AI resistance using typical estrogen receptor-positive breast cancer cell lines. We established tumor microenvironment-dependent AI-resistant models and elucidated the underlying mechanisms. RESULTS: T-47D cells had a higher dependence on microenvironment-derived factors, such as estrogen or growth factors, for survival than MCF-7 cells. We, therefore, evaluated tumor microenvironment growth factors with respect to AI resistance using T-47D cells. We established three resistant cell lines (V1, V2, and V3) that survived estrogen deprivation and growth factor-supplemented conditions. These cell lines were deficient in estrogen receptor α expression and estrogen-dependent growth. Among six representative growth factors, epidermal growth factor was the most influential. In these models, HER2 protein was overexpressed without gene amplification and intracellular phosphorylation pathways were activated compared to parental cell lines. Molecular targeting inhibitors revealed that V1 and V2 primarily rely on the PI3 K pathway for survival, whereas V3 relies on the MAPK pathway. CONCLUSIONS: This study demonstrates the importance of tumor microenvironment-derived factors for the development of AI resistance. These resistant models did not utilize the same resistance mechanism, suggesting that flexible strategies are essential in conquering resistance.

Utility of Ki67 labeling index, cyclin D1 expression, and ER-activity level in postmenopausal ER-positive and HER2-negative breast cancer with neoadjuvant chemo-endocrine therapy.

In this study, we investigated the relationships of pathological response after neoadjuvant chemo-endocrine therapy with alterations in the Ki67 labeling index (LI), expression of cyclin D1 (CCND1) and progesterone receptor (PgR), and estrogen receptor (ER) activity in breast cancer. A total of 43 Japanese post-menopausal ER-positive and human epidermal growth factor receptor 2-negative invasive breast cancer patients with tumors >2 cm or positive lymph nodes were enrolled. Exemestane alone was administered for 2 months. Neoadjuvant chemo-endocrine therapy included four cycles of doxorubicin plus paclitaxel followed by weekly paclitaxel. The immunohistochemical expression of Ki67 LI, CCND1, and PgR, and ER activity were evaluated using core needle biopsy samples at the pretreatment and post-exemestane alone stages. ER activity was evaluated through transfection of an adenovirus vector carrying an estrogen-response element-green fluorescent protein gene. In current study, marked pathological responses (including 4.7% with pathological complete response) were obtained in 34.9% of patients. Ki67 LI and PgR expression were significantly decreased after treatment. High Ki67 LI at pretreatment was a significant predictive factor of marked pathological response. At the stage of post-exemestane alone, Ki67 LI was not significantly associated with pathological response; however, high CCND1 expression was significantly correlated with high Ki67 LI. Moreover, low-level ER activity at the post-exemestane alone stage was significantly associated with marked pathological response. In conclusions, pretreatment Ki67 LI was a predictor of response to neoadjuvant chemo-endocrine therapy. CCND1 expression and ER activity at the post-endocrine therapy alone stage may be useful in determining further treatments.

Cancer stem-like properties of hormonal therapy-resistant breast cancer cells.

BACKGROUND: Presently, hormonal therapy targeting estrogen receptors is the most effective treatment available for luminal breast cancer. However, many patients relapse after the therapy. It has been suggested that cancer stem-like cells are involved with hormonal therapy resistance; in the present study, we evaluated this hypothesis. METHODS: In the present study, we used our previously established hormonal therapy-resistant cell lines, including aromatase inhibitor (AI)-resistant cells (Type 1 and Type 2) and fulvestrant-resistant cells (MFR). RESULTS: AI-resistant cell lines expressing ER (Type 1 V1 and V2) showed high cancer stemness in terms of their CD44/CD24 expression and side populations, which were stimulated by the addition of estrogen and inhibited by fulvestrant. However, ALDH activity was lower than in the ER-negative resistant cells, suggesting that the stemness of luminal cells is distinct from that of basal-like breast cancer cells. The migration and invasion activity of the ER-positive Type 1 V1 and V2 cells were higher than in the ER-negative cell lines, Type 2 and MFR. CONCLUSIONS: Fractionation of parental cells based on CD44/CD24 expression and colony formation assay indicated that CD44+/CD24+ cells might be the origin of hormonal therapy-resistant cells. This population reconstituted various other subpopulations under estrogen deprivation. These results indicate that hormonal therapy resistance is closely related to the cancer stem cell-like properties of luminal breast cancer.

Comparing protein and mRNA expressions of the human epidermal growth factor receptor family in estrogen receptor-positive breast cancer.

The human epidermal growth factor receptor (HER) family plays a vital role in the development of resistance to treatments in estrogen receptor (ER)-positive breast cancer. This study investigated the correlation between protein and mRNA expressions of the HER family in ER-positive breast cancer. We dissected regions of invasive cancer from the frozen tissues of 34 patients with ER-positive breast cancer using laser-capture microdissection, followed by evaluation of the mRNA levels of the ER and HER family (EGFR, HER2, HER3, and HER4) using the quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay. In addition, we assessed the protein expressions of the ER and HER family using an immunohistochemical (IHC) assay. A significant correlation was observed between the ER protein and mRNA expressions. For HER2, HER3, and HER4, protein expressions significantly correlated with mRNA levels. We established significant correlations of the mRNA level between EGFR versus HER2, as well as EGFR versus HER3. Furthermore, a significant correlation of the mRNA level between HER2 and HER3 was illustrated. In conclusion, IHC evaluation may be reliable and representable for mRNA. Hence, this study established a marked correlation between the mRNA expressions of HER family members in patients with ER-positive breast cancer.

Acquired resistance to everolimus in aromatase inhibitor-resistant breast cancer.

We previously reported the establishment of several types of long-term estrogen-depleted-resistant (EDR) cell lines from MCF-7 breast cancer cells. Type 1 EDR cells exhibited the best-studied mechanism of aromatase inhibitor (AI) resistance, in which estrogen receptor (ER) expression remained positive and PI3K signaling was upregulated. Type 2 EDR cells showed reduced ER activity and upregulated JNK-related signaling. The mTOR inhibitor everolimus reduced growth in cells similar to Type 1 EDR cells. The present study generated everolimus-resistant (EvR) cells from Types 1 and 2 EDR cells following long-term exposure to everolimus in vitro. These EvR cells modeled resistance to AI and everolimus combination therapies following first-line AI treatment failure. In Type 1 EvR cells, everolimus resistance was dependent on MAPK signaling; single agents were not effective, but hormonal therapy combined with a kinase inhibitor effectively reduced cell growth. In Type 2 EvR cells, ER expression remained negative and a JNK inhibitor was ineffective, but a Src inhibitor reduced cell growth. The mechanism of acquired everolimus resistance appears to vary depending on the mechanism of AI resistance. Strategies targeting resistant tumors should be tailored based on the resistance mechanisms, as these mechanisms impact therapeutic efficacy.

Single CpG site methylation controls estrogen receptor gene transcription and correlates with hormone therapy resistance.

Hormone therapy is the most effective treatment for patients with estrogen receptor α-positive breast cancers. However, although resistance occurs during treatment in some cases and often reflects changed estrogen receptor α status, the relationship between changes in estrogen receptor α expression and resistance to therapy are poorly understood. In this study, we identified a mechanism for altered estrogen receptor α expression during disease progression and acquired hormone therapy resistance in aromatase inhibitor-resistant breast cancer cell lines. Subsequently, we investigated promoter switching and DNA methylation status of the estrogen receptor α promoter, and found marked changes of methylation at a single CpG site (CpG4) in resistant cells. In addition, luciferase reporter assays showed reduced transcriptional activity from this methylated CpG site. This CpG region was also completely conserved among species, suggesting that it acts as a methylation-sensitive Ets-2 transcription factor binding site, as confirmed using chromatin immunoprecipitation assays. In estrogen receptor α-positive tumors, CpG4 methylation levels were inversely correlated with estrogen receptor α expression status, suggesting that single CpG site plays an important role in the regulation of estrogen receptor α transcription.

Recent trends in microRNA research into breast cancer with particular focus on the associations between microRNAs and intrinsic subtypes.

MicroRNAs (miRNAs) are short non-coding RNAs that regulate the function of target genes at the post-transcriptional phase. miRNAs are considered to have roles in the development, progression and metastasis of cancer. Recent studies have indicated that particular miRNA signatures are correlated with tumor aggressiveness, response to drug therapy and patient outcome in breast cancer. On the other hand, in routine clinical practice, the treatment regimens for breast cancer are determined based on the intrinsic subtype of the primary tumor. Previous studies have shown that miRNA expression profiles of each intrinsic subtypes of breast cancer differ. In hormone receptor-positive/human epidermal growth factor receptor 2 (HER2)-negative breast cancer, miRNA expressions are found to be correlated with endocrine therapy resistance, progesterone receptor expression and heat shock protein activity. Some miRNAs are associated with resistance to HER2-targeted therapy and HER3 expression in HER2-positive breast cancer. In triple-negative breast cancer, miRNA expressions are found to be associated with BRCA mutations, immune system, epithelial-mesenchymal transition, cancer stem cell properties and androgen receptor expression. As it has been clarified that the expression levels and functions of miRNA differ among the various subtypes of breast cancer, and it is necessary to take account of the characteristics of each breast cancer subtype during research into the roles of miRNA in breast cancer. In addition, the discovery of the roles played by miRNAs in breast cancer might provide new opportunities for the development of novel strategies for diagnosing and treating breast cancer.

Different epigenetic mechanisms of ERα implicated in the fate of fulvestrant-resistant breast cancer.

Approximately 70% of breast cancers express estrogen receptor α (ERα), which plays critical roles in breast cancer development. Fulvestrant has been effectively used to treat ERα-positive breast cancer, although resistance remains a critical problem. To elucidate the mechanism of resistance to fulvestrant, we established fulvestrant-resistant cell-lines named MFR (MCF-7 derived fulvestrant resistance) and TFR (T-47D derived fulvestrant resistance) from the ERα-positive luminal breast cancer cell lines MCF-7 and T-47D, respectively. Both fulvestrant-resistant cell lines lost sensitivity to estrogen and anti-estrogens. We observed diminished ERα expression at both the protein and mRNA levels. To address the mechanism of gene expression regulation, we examined epigenetic alteration, especially the DNA methylation level of ERα gene promoters. MFR cells displayed high methylation levels upstream of the ERα gene, whereas no change in DNA methylation was observed in TFR cells. Hence, we examined the gene expression plasticity of ERα, as there are differences in its reversibility following fulvestrant withdrawal. ERα gene expression was not restored in MFR cells, and alternative intracellular phosphorylation signals were activated. By contrast, TFR cells exhibited plasticity of ERα gene expression and ERα-dependent growth; moreover, these cells were resensitized to estrogen and anti-estrogens. The difference in epigenetic regulation among individual cells might explain the difference in the plasticity of ERα expression. We also identified an MFR cell-activating HER/Src-Akt/MAPK pathway; thus, the specific inhibitors effectively blocked MFR cell growth. This finding implies the presence of multiple fulvestrant resistance mechanisms and suggests that the optimal therapies differ among individual tumors as a result of differing epigenetic mechanisms regulating ERα gene expression.

Estrogen receptor activation by tobacco smoke condensate in hormonal therapy-resistant breast cancer cells.

The relationship between tobacco smoke and breast cancer incidence has been studied for many years, but the effect of smoking on hormonal therapy has not been previously reported. We investigated the effect of smoking on hormonal therapy by performing in vitro experiments. We first prepared tobacco smoke condensate (TSC) and examined its effect on estrogen receptor (ER) activity. The ER activity was analyzed using MCF-7-E10 cells into which the estrogen-responsive element (ERE)-green fluorescent protein (GFP) reporter gene had been stably introduced (GFP assay) and performing an ERE-luciferase assay. TSC significantly activated ERs, and upregulated its endogenous target genes. This activation was inhibited by fulvestrant but more weakly by tamoxifen. These results suggest that the activation mechanism may be different from that for estrogen. Furthermore, using E10 estrogen depletion-resistant cells (EDR cells) established as a hormonal therapy-resistant model showing estrogen-independent ER activity, ER activation and induction of ER target genes were significantly higher following TSC treatment than by estradiol (E2). These responses were much higher than those of the parental E10 cells. In addition, the phosphorylation status of signaling factors (ERK1/2, Akt) and ER in the E10-EDR cells treated with TSC increased. The gene expression profile induced by estrogenic effects of TSC was characterized by microarray analysis. The findings suggested that TSC activates ER by both ligand-dependent and -independent mechanisms. Although TSC constituents will be metabolized in vivo, breast cancer tissues might be exposed for a long period along with hormonal therapy. Tobacco smoke may have a possibility to interfere with hormonal therapy for breast cancer, which may have important implications for the management of therapy.

Prognostic value of the ubiquitin ligase carboxyl terminus of the Hsc70-interacting protein in postmenopausal breast cancer.

The carboxyl terminus of the Hsc70-interacting protein (CHIP) is considered to induce the ubiquitination and degradation of several oncogenic proteins, and play a role in the inhibition of tumor progression and invasion under experimental conditions. However, the impact of CHIP expression on the prognosis of breast cancer patients has not yet been established. In this study, using an immunohistochemical method, 272 patients with invasive breast cancer were assessed for the expression of CHIP (graded scores 0-3) and the statuses of biomarkers, such as estrogen receptor (ER), progesterone receptor (PgR), and HER2. The relationships between the statuses of CHIP and biomarkers as well as clinical features were also evaluated, and that between the expression of CHIP and patient prognosis was analyzed. We revealed that the strong expression of CHIP correlated with positive ER (P < 0.001), positive PgR (P < 0.001), and negative HER2 (P = 0.02). In postmenopausal patients, relapse-free survival (RFS) was significantly better in the high CHIP group than in the low CHIP group (P = 0.042). In addition, RFS and cancer-specific survival (CSS) were significantly better in patients with ER-positive/CHIP score 3 tumors than in those with ER-negative/CHIP score 0 tumors (RFS: P = 0.038, CSS: P = 0.0098). The methylation status of CHIP gene promoter did not always account for the down-regulation of its expression. In conclusion, the overexpression of CHIP is a potent prognostic factor of a good prognosis in ER-positive breast cancer patients in the postmenopausal phase.

Contribution of Estrone Sulfate to Cell Proliferation in Aromatase Inhibitor (AI) -Resistant, Hormone Receptor-Positive Breast Cancer.

Aromatase inhibitors (AIs) effectively treat hormone receptor-positive postmenopausal breast cancer, but some patients do not respond to treatment or experience recurrence. Mechanisms of AI resistance include ligand-independent activation of the estrogen receptor (ER) and signaling via other growth factor receptors; however, these do not account for all forms of resistance. Here we present an alternative mechanism of AI resistance. We ectopically expressed aromatase in MCF-7 cells expressing green fluorescent protein as an index of ER activity. Aromatase-overexpressing MCF-7 cells were cultured in estrogen-depleted medium supplemented with testosterone and the AI, letrozole, to establish letrozole-resistant (LR) cell lines. Compared with parental cells, LR cells had higher mRNA levels of steroid sulfatase (STS), which converts estrone sulfate (E1S) to estrone, and the organic anion transporter peptides (OATPs), which mediate the uptake of E1S into cells. LR cells proliferated more in E1S-supplemented medium than did parental cells, and LR proliferation was effectively inhibited by an STS inhibitor in combination with letrozole and by ER-targeting drugs. Analysis of ER-positive primary breast cancer tissues showed a significant correlation between the increases in the mRNA levels of STS and the OATPs in the LR cell lines, which supports the validity of this AI-resistant model. This is the first study to demonstrate the contribution of STS and OATPs in E1S metabolism to the proliferation of AI-resistant breast cancer cells. We suggest that E1S metabolism represents a new target in AI-resistant breast cancer treatment.