AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens

Kanaya, Noriko; Nguyen, Duc T.; Lu, Hannah; Wang, Yuanzhong; Hsin, Li-Yu; Petreas, Myrto; Nelson, David; Guo, Weihong; Reynolds, Peggy; Synold, Timothy W.; Chen, Shiuan

doi:10.1007/s10549-015-3398-z

Cited by 9 publications

(9 citation statements)

References 31 publications

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“…In order to elucidate the impact cell cycle-driven ER can have on palbociclib response, we utilized the C4-12 cell line which is a variant of the MCF-7 cell line but lacks an endogenous ERα [ 27 ]. We generated a stably transfected ERα (C4-12/ERα) cell line which has a transcriptionally functional ER [ 28 ] but does not need estrogen for cell growth ( Supplementary Figure 1 ). CDK4/6 inhibitors did not affect the proliferation of C4-12/ERα (Figure 1 ), indicating that proliferation of these cells does not require ER even though they have biologically active.…”

Section: Resultsmentioning

confidence: 99%

“…We have established that C4-12/ERα cells do not require estrogen for cell growth ( Supplementary Figure 1 ) but ER is activated by estrogen as seen with the up-regulation of ER regulated genes such as GREB1, TFF1 and PGR, which is also observed in the estrogen dependent MCF-7 cell line ( Supplementary Table 2 ). Analysis of C4-12/ERα cells shows no change in the G2/M-phase or checkpoint regulation genes ( Supplementary Figure 3B ); thus, the transcriptionally functional ER in this cell line [ 28 ] is not involved in cell cycle progression. Moreover, even though this cell line expresses RB, which alone is not a predictive factor of palbociclib response [ 18 , 20 , 40 – 42 ], a transcriptionally functional ER involved in cell cycle progression is required for the inhibitor response.…”

Section: Discussionmentioning

confidence: 99%

“…C4-12 was cultured in phenol red-free high glucose DMEM with 10% charcoal/dextran treated FBS [ 27 ]. C4-12/ERα cell line was generated as previously reported [ 28 ]. Culture media were supplemented with 1 mM sodium pyruvate and 100 U/mL penicillin-streptomycin.…”

Section: Methodsmentioning

confidence: 99%

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ERα-mediated cell cycle progression is an important requisite for CDK4/6 inhibitor response in HR+ breast cancer

Petrossian

Kanaya

et al. 2018

Oncotarget

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While ER has multiple biological effects, ER-cyclin D1-CDK4/6-RB is a critical pathway for the action of estrogen on the cell cycle, especially for breast cancers that rely on estrogen for growth. The latest and most efficient CDK4/6 inhibitors target the phosphorylation of retinoblastoma (RB) tumor suppressor gene; thus, altering levels of many cell cycle molecules. Estrogen receptor (ER)+/HER2- breast cancers have shown great progression free survival when CDK4/6 inhibitors are combined with endocrine therapies. Here we report the mechanism of antiestrogen (fulvestrant) combination with CDK4/6 inhibitors is due to synergism in the suppression of ER-mediated cell cycle progression. Furthermore, we performed single cell analysis of cells from an estrogen dependent/hormone receptor-positive patient derived xenograft (PDX) tumor model treated with palbociclib. These single cells expressed various levels of ER and RB which are involved in cell cycle regulation; and the response to palbociclib treatment relies not only on the ER-cyclin D1-CDK4/6-RB pathway but it is also dependent on elevated levels of ER and/or RB. Our preclinical studies show that palbociclib response is dependent on cells with ER, which is directly involved in cell cycle progression in hormone receptor positive (HR+) breast cancer.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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ERα-mediated cell cycle progression is an important requisite for CDK4/6 inhibitor response in HR+ breast cancer

Petrossian

Kanaya

et al. 2018

Oncotarget

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“…Using a luciferase reporter system, each PBDE was tested in a C4-12ERa ERE assay for estrogenic and/or anti-estrogenic properties. The C4-12ERa ERE cells were established in our laboratory from C4-12 cells, derived from MCF-7 breast cancer cells, that are deficient in endogenous ER expression; we previously confirmed that the reporter signal is highly specific for ERa (not ERb) (Kanaya et al, 2015). In the agonist mode of this assay, cells were treated only with the individual PBDE (no cotreatment).…”

Section: Effects Of Bde-47 Bde-100 and Bde-153 On Era Through Cell-mentioning

confidence: 94%

“…MCF7aro cells (Zhou et al, 1990) were maintained in MEM supplemented with 10% FBS, 2 mM L-glutamine, 1 mM sodium pyruvate, 100 U/ml penicillin-streptomycin, and 100 lg/ml of G418. C4-12 ERaERE cells (Kanaya et al, 2015) were maintained in phenol red-free high-glucose DMEM supplemented with 10% charcoal-dextran stripped FBS (Omega Scientific, Tarzana, California), 2 mM L-glutamine, 1 mM sodium pyruvate, 100 U/ml penicillin-streptomycin, and 100 lg/ml of hygromycin. T47D cells were maintained in RPMI-1640 supplemented with 10% FBS, 2 mM L-glutamine, 1 mM sodium pyruvate, and 100 U/ml penicillin-streptomycin.…”

Section: Methodsmentioning

confidence: 99%

Molecular Mechanisms of Polybrominated Diphenyl Ethers (BDE-47, BDE-100, and BDE-153) in Human Breast Cancer Cells and Patient-Derived Xenografts

Kanaya

Bernal

Chang

et al. 2019

Toxicological Sciences

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Polybrominated diphenyl ethers (PBDEs) have been used as flame retardants in household materials. Their environmental persistence has led to continuous human exposure and significant tissue levels. Three PBDE congeners (BDE-47, BDE-100, and BDE-153) have been frequently detected in human serum. Although these compounds appear to possess endocrine disrupting activity, studies are largely missing to determine the biological mechanisms of PBDEs in breast cancer cells. Here, we assessed PBDE bioactivities with three complementary strategies: receptor binding/activity assays; nonbiased RNA-sequencing analysis using an estrogen-dependent breast cancer cell line MCF-7aroERE; and in vivo assessments using patient-derived xenograft (PDX) models of human breast cancer. According to the results from in vitro experiments, the PBDE congeners regulate distinct nuclear receptor signaling pathways. BDE-47 acts as a weak agonist of both estrogen receptor a (ERa) and estrogen-related receptor a (ERRa); it could stimulate proliferation of MCF-7aroERE and induced expression of ER-regulated genes (including cell cycle genes). BDE-153 was found to act as a weak antagonist of ERa. BDE-100 could act as (1) an agonist of aryl hydrocarbon receptor (AhR), inducing expression of CYP1A1 and CYP1B1 and (2) as a very weak agonist/antagonist of ERa. In vivo, a mixture of the three congeners with ratios detected in human serum was tested in an ERþ PDX model. The mixture exhibited estrogenic activity through apoptosis/cell cycle regulation and increased the expression of a proliferation marker, Ki-67. These results advance our understanding of the mechanisms of PBDE exposure in breast cancer cells.

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Superior suppression of serum estrogens during neoadjuvant breast cancer treatment with letrozole compared to exemestane

Bertelsen,

Almås,

Fjermeros

et al. 2024

Breast Cancer Res Treat

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Purpose The aromatase inhibitor letrozole and the aromatase inactivator exemestane are two of the most pivotal cancer drugs used for endocrine treatment of ER-positive breast cancer in all phases of the disease. Although both drugs inhibit CYP19 (aromatase) and have been used for decades, a direct head-to-head, intra-patient-cross-over comparison of their ability to decrease estrogen synthesis in vivo is still lacking. Methods Postmenopausal breast cancer patients suitable for neoadjuvant endocrine therapy were randomized to receive either letrozole (2.5 mg o.d.) or exemestane (25 mg o.d.) for an initial treatment period, followed by a second treatment period on the alternative drug (intra-patient cross-over study design). Serum levels of estrone (E1), estradiol (E2), letrozole, exemestane, and 17-hydroxyexemestane were quantified simultaneously using a novel, ultrasensitive LC–MS/MS method established in our laboratory. Results Complete sets of serum samples (baseline and during treatment with letrozole or exemestane) were available from 79 patients, including 40 patients starting with letrozole (cohort 1) and 39 with exemestane (cohort 2). Mean serum estrone and estradiol levels in cohort 1 were 174 pmol/L and 46.4 pmol/L at baseline, respectively. Treatment with letrozole suppressed serum E1 and E2 to a mean value of 0.2 pmol/L and 0.4 pmol/L (P < 0.001). After the cross-over to exemestane, mean serum levels of E1 and E2 increased to 1.4 pmol/L and 0.7 pmol/L, respectively. In cohort 2, baseline mean serum levels of E1 and E2 were 159 and 32.5 pmol/L, respectively. Treatment with exemestane decreased these values to 1.8 pmol/L for E1 and 0.6 pmol/L for E2 (P < 0.001). Following cross-over to letrozole, mean serum levels of E1 and E2 were significantly further reduced to 0.1 pmol/L and 0.4 pmol/L, respectively. Serum drug levels were monitored in all patients throughout the entire treatment and confirmed adherence to the protocol and drug concentrations within the therapeutic range for all patients. Additionally, Ki-67 values decreased significantly during treatment with both aromatase inhibitors, showing a trend toward a stronger suppression in obese women. Conclusion To the best of our knowledge, we present here for the first time a comprehensive and direct head-to-head, intra-patient-cross-over comparison of the aromatase inhibitor letrozole and the aromatase inactivator exemestane concerning their ability to suppress serum estrogen levels in vivo. All in all, our results clearly demonstrate that letrozole therapy results in a more profound suppression of serum E1 and E2 levels compared to exemestane.

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AroER tri-screen™ is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens

Cited by 9 publications

References 31 publications

ERα-mediated cell cycle progression is an important requisite for CDK4/6 inhibitor response in HR+ breast cancer

ERα-mediated cell cycle progression is an important requisite for CDK4/6 inhibitor response in HR+ breast cancer

Molecular Mechanisms of Polybrominated Diphenyl Ethers (BDE-47, BDE-100, and BDE-153) in Human Breast Cancer Cells and Patient-Derived Xenografts

Superior suppression of serum estrogens during neoadjuvant breast cancer treatment with letrozole compared to exemestane

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