Purpose: Several lines of evidence support targeting the androgen signaling pathway in breast cancer. Enzalutamide is a potent inhibitor of androgen receptor signaling. Preclinical data in estrogen-expressing breast cancer models demonstrated activity of enzalutamide monotherapy and enhanced activity when combined with various endocrine therapies (ET). Enzalutamide is a strong cytochrome P450 3A4 (CYP3A4) inducer, and ETs are commonly metabolized by CYP3A4. The pharmacokinetic (PK) interactions, safety, and tolerability of enzalutamide monotherapy and in combination with ETs were assessed in this phase I/Ib study.

Experimental Design: Enzalutamide monotherapy was assessed in dose-escalation and dose-expansion cohorts of patients with advanced breast cancer. Additional cohorts examined effects of enzalutamide on anastrozole, exemestane, and fulvestrant PK in patients with estrogen receptor–positive/progesterone receptor–positive (ER+/PgR+) breast cancer.

Results: Enzalutamide monotherapy (n = 29) or in combination with ETs (n = 70) was generally well tolerated. Enzalutamide PK in women was similar to prior data on PK in men with prostate cancer. Enzalutamide decreased plasma exposure to anastrozole by approximately 90% and exemestane by approximately 50%. Enzalutamide did not significantly affect fulvestrant PK. Exposure of exemestane 50 mg/day given with enzalutamide was similar to exemestane 25 mg/day alone.

Conclusions: These results support a 160 mg/day enzalutamide dose in women with breast cancer. Enzalutamide can be given in combination with fulvestrant without dose modifications. Exemestane should be doubled from 25 mg/day to 50 mg/day when given in combination with enzalutamide; this combination is being investigated in a randomized phase II study in patients with ER+/PgR+ breast cancer. Clin Cancer Res; 23(15); 4046–54. ©2017 AACR.

Androgen signaling may play a role in the biology of breast cancers. Enzalutamide is a potent androgen receptor signaling inhibitor approved for the treatment of men with castration-resistant prostate cancer. A favorable risk–benefit profile is well established for 160 mg/day enzalutamide in prostate cancer. Preclinical data suggest complementary effects between enzalutamide and endocrine therapies (ET) in estrogen receptor–positive breast cancer xenografts. To facilitate future trials of safety and effectiveness, and determine the suitability of 160 mg in women, this study investigated the pharmacokinetics, safety, and tolerability of enzalutamide alone or in combination with ETs in women with breast cancer. As enzalutamide strongly induces cytochrome P450 3A4 (CYP3A4), drug–drug interactions were assessed for enzalutamide in combination with the potential CYP3A4 substrates anastrozole, exemestane, and fulvestrant. Results of this phase I/Ib study supported an ongoing phase II study of enzalutamide in combination with exemestane.

Several lines of evidence support the role that androgen receptor (AR) signaling may have in the biology of breast cancer (1). In a study evaluating more than 3,000 breast tumor samples, AR expression was observed in 77% of invasive tumors and across all molecular phenotypes [91% luminal A, 68% luminal B, 59% erb-b2 receptor tyrosine kinase 2 (HER2)–amplified, 32% basal-like, and 46% unclassified breast cancer; ref. 2].

Enzalutamide is a potent AR signaling inhibitor that binds the AR to competitively inhibit (i) androgen-induced receptor activation, (ii) nuclear translocation of the activated AR, and (iii) association of the AR with chromatin (3, 4). These complementary actions result in reduced expression of AR-dependent genes, decreased growth of cancer cells, induction of cancer cell death, and tumor regression in preclinical models of prostate cancer (5). Enzalutamide 160 mg/day is approved for the treatment of metastatic castration-resistant prostate cancer in more than 50 countries for prechemotherapy and more than 75 countries for postchemotherapy treatment (6).

AR expression has been shown to induce resistance to both tamoxifen and aromatase inhibitors (AI; refs. 7, 8) in estrogen receptor (ER)–expressing cell lines. In mouse xenograft models, enzalutamide monotherapy was as effective as tamoxifen at inhibiting the estrogen-stimulated growth of MCF7, a human breast carcinoma cell line that expresses both AR and ER (9). In addition, enzalutamide blocked ERα phosphorylation, signaling activity, and cell proliferation in tamoxifen-resistant breast cancer cells (10).

AIs are widely used endocrine therapies (ET) for ER-positive/progesterone receptor–positive (ER+/PgR+) breast cancers. They block the conversion of androgens to estrone and estradiol, thereby diverting estrogen precursors to androgen synthesis, resulting in increases in androgen concentrations in the systemic circulation (11, 12). This increase in circulating androgens could stimulate the growth of androgen-expressing and -sensitive breast cancer. It follows that an AI combined with enzalutamide could have clinical utility in ER+/PgR+ breast cancer (13).

Fulvestrant is a selective ER antagonist that competitively binds to the ER with a binding affinity that is approximately 100 times greater than that of tamoxifen (14). In a preclinical ER-positive/AR-positive breast cancer model using BCK4 cells, enzalutamide and fulvestrant synergistically inhibited tumor cell growth (13). These preclinical data support the clinical evaluation of enzalutamide combined with fulvestrant.

ETs are commonly metabolized by cytochrome P450 3A4 (CYP3A4) enzymes. Because enzalutamide is a strong CYP3A4 inducer, an evaluation of potential pharmacokinetic (PK) and pharmacodynamic (PD) interactions was performed to assess the clinical feasibility of combining enzalutamide with commonly used ETs for breast cancer. Here, we report the results of a phase I/Ib study to evaluate the PK, safety, and tolerability of enzalutamide monotherapy and in combination with ETs in women with advanced breast cancer.

Study design and treatments

This two-stage study consisted of a dose-escalation stage (stage 1) followed by a dose-expansion stage (stage 2; Fig. 1). Stage 1 used a standard 3 + 3 design to evaluate two dosage levels of enzalutamide monotherapy (80 mg/day and 160 mg/day orally). The enzalutamide dose defined in stage 1 was evaluated in stage 2.

Figure 1.

Study schematic of (A) dose escalation (stage 1) and (B) dose expansion (stage 2). DDI, drug–drug interaction.

Figure 1.

Study schematic of (A) dose escalation (stage 1) and (B) dose expansion (stage 2). DDI, drug–drug interaction.

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In stage 1, a minimum of 3 patients received a single dose of enzalutamide 80 mg. PK blood samples were collected through day 8, and once-daily dosing began on day 8, with a dose-limiting toxicity (DLT) window of 35 days from day 1 (Fig. 1). If none of the 3 or 1 or fewer of the 6 patients experienced a DLT, then enrollment into the 160 mg/day dose at the same schedule could occur. The DLTs were defined as any of the following events determined to be related to enzalutamide: seizure; grade 3 or greater fatigue, diarrhea, nausea, or vomiting that did not improve to grade 1 within 14 days of initiating standard of care therapy; and grade 3 or greater hematologic or nonhematologic toxicity other than laboratory abnormalities thought to be attributable to disease progression or of no clinical significance.

The stage 2 portion of the study evaluated enzalutamide monotherapy and in combination with either anastrozole (1 mg/day orally), exemestane (25 mg/day or 50 mg/day orally), or fulvestrant [500 mg intramuscularly once every 28 (±2) days]. These ETs were initially administered as monotherapy to ensure attainment of each agent's steady-state concentration (15–17). Once PK data were available indicating decreased exposure to both anastrozole and exemestane, an additional cohort to evaluate enzalutamide combined with exemestane 50 mg was added, and the protocol was amended to allow patients receiving those treatments to switch to exemestane 50 mg. Treatment in all cohorts was continued until disease progression, unacceptable toxicity, initiation of another cytotoxic or investigational agent, or discontinuation at the investigator's discretion.

Study population

The study enrolled patients with locally advanced or metastatic breast cancer that was not amenable to curative therapy. Eligible patients were at least 18 years of age and were required to have adequate organ and bone marrow function and an Eastern Cooperative Oncology Performance Status (ECOG PS) of 2 or less. Patients who received enzalutamide monotherapy were required to have received at least two prior lines of systemic therapy for advanced breast cancer. Patients enrolled in all of the combination cohorts were required to have ER+/PgR+ disease and be postmenopausal. Treatment with luteinizing hormone-releasing hormone agonists to induce menopause was permitted.

Patients enrolled in the stage 2 enzalutamide monotherapy and the fulvestrant cohorts were required to submit a tumor specimen for determination of nuclear AR expression level by immunohistochemistry (IHC). Tissue submission from other cohorts for AR IHC determination was optional. Local pathology assessment of AR expression was allowed. Only patients whose tumor specimens had at least 10% nuclear AR IHC staining were eligible for enrollment in those cohorts where AR assessment was required (enzalutamide monotherapy and fulvestrant cohorts). Previous studies of bicalutamide and abiraterone acetate in triple-negative breast cancer have used a cutoff of 10% nuclear AR by IHC to define patients with AR-positive disease (18, 19).

Patients were ineligible for enrollment if they had a history of seizure, were using medications that could reduce seizure threshold, or had known or suspected brain metastases or leptomeningeal disease. Initially, patients who had HER2-positive breast cancer were allowed, but the protocol was amended to restrict to HER2 normal after the first 64 patients were enrolled. Concomitant treatment with potent CYP3A4 inducers (including tamoxifen) or inhibitors was prohibited.

The study was conducted at six sites in the United States. The study protocol and its amendments were approved by the respective Institutional Review Boards, and all patients provided written informed consent prior to participating in the study. The study was conducted under the principles of the World Medical Association, Declaration of Helsinki, and Good Clinical Practice guidelines of the International Conference on Harmonisation.

PK assessments

In stage 1, enzalutamide PK samples for single- and multiple-dose characterization were collected at day 1 and around day 50 at 0.5, 1, 2, 4, 6, 24, and 48 hours, with two additional samples collected any time between days 4 and 7. In addition, enzalutamide predose [minimum plasma concentration (Cmin)] PK samples were collected on days 29 and 57.

The combination cohorts followed a single-sequence crossover design with within-subject comparisons of PK and PD endpoints for ET monotherapy versus ET in combination with enzalutamide. The start of the combination treatment (ET plus enzalutamide) was designated as day 1. PK determinations for AI monotherapy and AI combined with enzalutamide occurred on day −1 and day 29, respectively. Anastrozole and exemestane PK samples were collected predose and at 0.5, 1, 1.5, 2, 3, 4, 6, 8, and 24 hours after dosing. Fulvestrant predose (Cmin) PK samples were collected on day 1 (prior to the enzalutamide dose) and day 29. Enzalutamide predose Cmin PK samples were collected on days 1, 15, 29, 43, and 57.

Bioanalytical methods have been previously published for anastrozole (20), exemestane (21), enzalutamide (22), and fulvestrant (23). Validated liquid chromatography with tandem mass spectrometry methods were used to measure plasma concentrations of anastrozole, exemestane, fulvestrant, enzalutamide, and N-desmethyl enzalutamide (active metabolite). The lower limit of quantification (LLOQ) was 0.100 ng/mL for anastrozole and exemestane, 1.00 ng/mL for fulvestrant, and 0.020 μg/mL for enzalutamide and N-desmethyl enzalutamide.

PK parameters were calculated from plasma time-concentration data using standard noncompartmental methods in Phoenix WinNonlin version 5.2 (Pharsight Corporation) and SAS version 9.1.3 (SAS Institute) software. Enzalutamide PK parameters included Cmin, maximum plasma concentration (Cmax), apparent oral clearance (CL/F), and area under the plasma concentration-time curve during one dosing interval at steady state (AUCtau). Anastrozole and exemestane PK parameters included Cmax and AUCtau. Plasma concentration data for fulvestrant and enzalutamide were summarized by descriptive statistics.

In the AI cohorts, the PK evaluable population consisted of all patients who received their assigned dose of anastrozole or exemestane from days −14 to 29 and 160 mg/day enzalutamide without missing more than two doses from days 1 to 29. These patients also had sufficient PK samples for calculation of at least one PK parameter for anastrozole or exemestane (AUCtau or Cmax) on days −1 and 29. In the fulvestrant cohort, the PK evaluable population consisted of all patients who received at least three consecutive 500 mg doses of fulvestrant prior to day 1, one 500 mg dose of fulvestrant on day 1, and 160 mg/day enzalutamide without significant interruption from days 1 to 29. These patients also had suitable PK samples for calculation of Cmin on days 1 and 29.

PD assessments

PD assessments consisted of an evaluation of plasma concentrations of estradiol and estrone in patients treated with anastrozole or exemestane. Residual plasma PK samples were pooled to create day −1 and day 29 samples for each patient. Validated methods based on gas chromatography with tandem mass spectrometry (20) were used to measure plasma concentrations of estradiol and estrone with an LLOQ of 0.625 pg/mL and 1.56 pg/mL, respectively.

Safety and antitumor assessments

The safety population included all patients who received at least one dose of enzalutamide. Safety was assessed weekly in stage 1 and biweekly in stage 2 for the first 8 weeks, then monthly thereafter for both stage 1 and stage 2, and for 30 days after the last dose of enzalutamide or prior to the initiation of a new treatment, whichever occurred first. Safety and tolerability were determined by assessment of adverse events (AE), physical examinations, ECOG PS, vital signs, and laboratory tests. The severity of abnormal laboratory values and AEs were classified using the National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE), version 4.0 (24).

For patients in both stages 1 and 2, radiographic assessments of disease status were performed at baseline, at week 7, and approximately every 3 months or earlier if clinically indicated. Tumor responses were defined using RECIST version 1.1 criteria.

Statistical analysis

Sample sizes per cohort were based on clinical rather than formal statistical considerations. Evaluable patients were those who were able to receive sufficient therapy to enable adequate PK analysis. Approximately 14 evaluable patients in each AI cohort and 10 evaluable patients in the fulvestrant cohort were required. Dosing, AEs, and baseline patient characteristics were summarized using descriptive statistics.

Estradiol and estrone concentrations that were less than the LLOQ were imputed to the LLOQ values prior to statistical calculations. To assess any potential effect of enzalutamide on PK or PD of the ETs, a linear mixed-effects model, with fixed effects for treatment period and measurements within patient as repeated measures, was performed on natural logarithmic-transformed values of AUCtau, Cmax, Cmin, estradiol concentration, and estrone concentration. Point estimates and 90% confidence intervals (CI) for the treatment differences (test minus reference) on the natural logarithmic scale were exponentiated to obtain estimates for ratios of geometric means (test/reference) on the original scale. Treatment differences were considered significant if the 90% CI did not include 1.00.

Patients

A total of 101 patients [n = 15 in stage 1; n = 84 in stage 2 (monotherapy n = 14, ET cohorts n = 70)] were enrolled between April 30, 2012, and February 3, 2015 (Fig. 1). Two patients were enrolled but not treated with enzalutamide (one admitted to hospice and one with clinical disease progression). There were 14 of 20 enrolled patients evaluable for PK in the anastrozole cohort, 12 of 16 in the exemestane 25 mg/day cohort, 16 of 23 in the exemestane 50 mg/day cohort, and 8 of 11 patients in the fulvestrant cohort. Of the 97 patients who discontinued, 89 (92%) discontinued due to disease progression, 5 (5%) due to AEs, 2 (2%) due to withdrawal of consent, and 1 (1%) was admitted to hospice. Data are presented as of January 8, 2016.

Demographics and baseline disease characteristics

Patient baseline demographic and disease characteristics are shown in Table 1. All but 1 patient treated with enzalutamide monotherapy was postmenopausal including 2 patients treated with ovarian suppression. The median age was 57 (range, 37–78) years in patients treated with enzalutamide monotherapy (stage 1 and stage 2) and 62 (range, 30–84) years in the ET combination cohorts. Of the 70 patients in the ET combination cohorts, 63 (90%) had received prior therapy for metastatic disease and 26 (37%) received prior treatment with the same hormonal agent as they received in the current study. Visceral disease and bone disease were present in the majority of all patients; 41 (59%) of all patients had at least three metastatic sites of disease. Although not required in all cohorts, tissue was submitted by 76 patients for AR IHC testing. The rate of AR+ disease in those tested by IHC was 54/76 (71%; range, 56%–91%) across all cohorts (Table 1). Five patients had HER2-positive breast cancer.

Table 1.

Demographics and baseline disease characteristics

Enza monotherapy (80/160 mg/d)Anastrozole (1 mg/d) + enzaaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enza
(n = 29)(n = 20)(n = 16)(n = 23)(n = 11)
Age, median (range), y 57 (37–78) 62 (30–84) 56 (42–79) 65 (34–75) 59 (53–78) 
ECOG PS, n (%) 1b 22 (76) 5 (25) 8 (50) 9 (39) 6 (55) 
Number of prior agents for aBC, median (range) 6 (2–15) 5 (0–13) 4 (1–6) 6 (0–16) 1 (0–11) 
 Hormonal 2 (0–5) 3 (0–5) 2 (0–3) 3 (0–5) 1 (0–4) 
 Nonhormonal 5 (0–12) 2 (0–10) 2 (0–5) 4 (0–13) 0 (0–7) 
Prior neoadjuvant/adjuvant, n (%) 21 (72) 13 (65) 9 (56) 17 (74) 6 (55) 
Months from aBC to study entry, median (range) 45 (11–133) 49 (2–125) 33 (9–97) 39 (1–333) 11 (3–182) 
Number of metastatic sites ≥3, n (%) 12 (41) 8 (40) 5 (31) 12 (52) 4 (36) 
Bone metastasis, n (%) 23 (79) 16 (80) 11 (69) 19 (83) 9 (82) 
 Bone only, n (%) 3 (10) 5 (25) 4 (25) 3 (13) 2 (18) 
Visceral disease, n (%) 16 (55) 13 (65) 8 (50) 16 (70) 4 (36) 
Measurable disease, n (%) 19 (66) 11 (55) 9 (56) 13 (57) 4 (36) 
Breast cancer subtype      
 ER+/PgR+ 22 (76) 20 (100) 16 (100) 23 (100) 10 (91) 
 HER2 amplifiedc 3 (10) 0 (0) 1 (6) 1 (4) 0 (0) 
 Triple negative 6 (21) 0 (0) 0 (0) 0 (0) 1 (9)d 
AR ≥ 10%, n/Ne 18/25 9/13 7/9 10/18 10/11 
Enza monotherapy (80/160 mg/d)Anastrozole (1 mg/d) + enzaaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enza
(n = 29)(n = 20)(n = 16)(n = 23)(n = 11)
Age, median (range), y 57 (37–78) 62 (30–84) 56 (42–79) 65 (34–75) 59 (53–78) 
ECOG PS, n (%) 1b 22 (76) 5 (25) 8 (50) 9 (39) 6 (55) 
Number of prior agents for aBC, median (range) 6 (2–15) 5 (0–13) 4 (1–6) 6 (0–16) 1 (0–11) 
 Hormonal 2 (0–5) 3 (0–5) 2 (0–3) 3 (0–5) 1 (0–4) 
 Nonhormonal 5 (0–12) 2 (0–10) 2 (0–5) 4 (0–13) 0 (0–7) 
Prior neoadjuvant/adjuvant, n (%) 21 (72) 13 (65) 9 (56) 17 (74) 6 (55) 
Months from aBC to study entry, median (range) 45 (11–133) 49 (2–125) 33 (9–97) 39 (1–333) 11 (3–182) 
Number of metastatic sites ≥3, n (%) 12 (41) 8 (40) 5 (31) 12 (52) 4 (36) 
Bone metastasis, n (%) 23 (79) 16 (80) 11 (69) 19 (83) 9 (82) 
 Bone only, n (%) 3 (10) 5 (25) 4 (25) 3 (13) 2 (18) 
Visceral disease, n (%) 16 (55) 13 (65) 8 (50) 16 (70) 4 (36) 
Measurable disease, n (%) 19 (66) 11 (55) 9 (56) 13 (57) 4 (36) 
Breast cancer subtype      
 ER+/PgR+ 22 (76) 20 (100) 16 (100) 23 (100) 10 (91) 
 HER2 amplifiedc 3 (10) 0 (0) 1 (6) 1 (4) 0 (0) 
 Triple negative 6 (21) 0 (0) 0 (0) 0 (0) 1 (9)d 
AR ≥ 10%, n/Ne 18/25 9/13 7/9 10/18 10/11 

Abbreviations: aBC, advanced breast cancer; enza, enzalutamide.

aEnza dose in stage 2 was 160 mg/day.

bTwo patients had ECOG PS of 2.

cProtocol amended to restrict to HER2 normal after first 64 patients enrolled.

dPatient with initial diagnosis of triple-negative breast cancer, but was ER+/PgR+ in metastatic setting.

eN = number of patients who underwent AR testing.

AR IHC staining results were available for 24 patients who received enzalutamide monotherapy (n = 13) or enzalutamide with fulvestrant (n = 11) in stage 2. AR IHC of at least 10% was reported in all but 2 patient tumor samples (range, 5–70) with AR IHC of at least 50% reported in 4 patients who received enzalutamide monotherapy and 6 patients who received enzalutamide with fulvestrant. Given the small sample size of these patient cohorts, an evaluation between the extent of AR expression and clinical outcomes was not performed.

Dose escalation

In stage 1, a CTCAE grade 3 DLT of adrenal insufficiency in a patient with a preexisting history of adrenal insufficiency was observed in 1 of the first 3 patients treated with enzalutamide 80 mg/day, and therefore the cohort was expanded to 7 patients. As no additional DLTs were observed with enzalutamide 80 mg/day, the dose was increased to the target dose of enzalutamide 160 mg/day. No further DLTs were observed, and enzalutamide 160 mg/day was selected for further testing in stage 2.

Drug treatment and compliance

The median duration of treatment for enzalutamide monotherapy was 8 (range, 3–65) weeks. The median duration of treatment for enzalutamide in combination with AIs was 8 (range, 1–152) weeks and with fulvestrant was 19 (range, 4–77) weeks.

PK and PD parameters

With multiple-dose administration at enzalutamide 160 mg/day, the mean SD CL/F was 0.51 (0.09) L/hour in women versus 0.52 (0.09) L/hour in men, indicating similar CL/F in women with breast cancer and men with prostate cancer (Supplementary Tables S1 and S2). Furthermore, the mean Cmin and Cmax values for enzalutamide, N-desmethyl enzalutamide, and the sum of enzalutamide plus N-desmethyl enzalutamide were similar in the two patient populations (Supplementary Tables S1 and S2).

In the ET cohorts, concomitant use of enzalutamide resulted in a mean AUCtau decrease by 89% for anastrozole and 43% to 57% for exemestane. The mean Cmin for fulvestrant decreased by 8%, which was well below the threshold of 20% for a clinically relevant drug interaction (ref. 25; Table 2 and Supplementary Fig. S1). Comparisons based on Cmax produced similar results to those based on mean AUCtau for all agents (Table 2).

Table 2.

PK and PD in ETs combined with enzalutamide versus ETs alone

Geometric mean
Hormone therapy and PK or PDa parameterET alone (reference)ET + enza (test)Geometric mean ratio (test/reference) (90% CI)
Anastrozole 1 mg/d, n = 14 
 AUCtau, ng·h/mL 755 84.0 0.111 (0.067–0.185) 
 Cmax, ng/mL 41.2 8.17 0.198 (0.124–0.316) 
 Estradiol, pg/mL 0.80 1.13 1.40 (1.04–1.90) 
 Estrone, pg/mL 4.16 7.00 1.68 (1.07–2.63) 
Exemestane 25 mg/d, n = 13 
 AUCtau, ng·h/mLb 106 60.9 0.575 (0.500–0.661) 
 Cmax, ng/mL 19.3 9.17 0.475 (0.354–0.636) 
 Estradiol, pg/mL 0.63 0.65 1.04 (0.98–1.11) 
 Estrone, pg/mL 1.56 1.76 1.13 (0.99–1.28) 
Exemestane 50 mg/d, n = 16 
 AUCtau, ng·h/mL 232 101 0.434 (0.364–0.519) 
 Cmax, ng/mL 45.0 17.2 0.382 (0.311–0.467) 
 Estradiol, pg/mL 0.63 0.71 1.13 (0.99–1.28) 
 Estrone, pg/mL 2.06 2.13 1.03 (0.84–1.27) 
Fulvestrant 500 mg/mo, n = 8 
 Cmin, ng/mL 13.5 12.5 0.920 (0.800–1.080) 
Geometric mean
Hormone therapy and PK or PDa parameterET alone (reference)ET + enza (test)Geometric mean ratio (test/reference) (90% CI)
Anastrozole 1 mg/d, n = 14 
 AUCtau, ng·h/mL 755 84.0 0.111 (0.067–0.185) 
 Cmax, ng/mL 41.2 8.17 0.198 (0.124–0.316) 
 Estradiol, pg/mL 0.80 1.13 1.40 (1.04–1.90) 
 Estrone, pg/mL 4.16 7.00 1.68 (1.07–2.63) 
Exemestane 25 mg/d, n = 13 
 AUCtau, ng·h/mLb 106 60.9 0.575 (0.500–0.661) 
 Cmax, ng/mL 19.3 9.17 0.475 (0.354–0.636) 
 Estradiol, pg/mL 0.63 0.65 1.04 (0.98–1.11) 
 Estrone, pg/mL 1.56 1.76 1.13 (0.99–1.28) 
Exemestane 50 mg/d, n = 16 
 AUCtau, ng·h/mL 232 101 0.434 (0.364–0.519) 
 Cmax, ng/mL 45.0 17.2 0.382 (0.311–0.467) 
 Estradiol, pg/mL 0.63 0.71 1.13 (0.99–1.28) 
 Estrone, pg/mL 2.06 2.13 1.03 (0.84–1.27) 
Fulvestrant 500 mg/mo, n = 8 
 Cmin, ng/mL 13.5 12.5 0.920 (0.800–1.080) 

Abbreviations: Cmin, minimum (predose) plasma concentration; enza, enzalutamide.

aEstradiol and estrone concentrations that were less than LLOQ were imputed to the LLOQ values (0.625 and 1.56 pg/mL, respectively) prior to calculating statistics.

bn = 12.

Of note, the exemestane PK profiles were virtually identical for exemestane 50 mg/day combined with enzalutamide versus exemestane 25 mg/day alone (Fig. 2), and a comparison of PK parameters for these two groups produced geometric mean ratios for AUCtau and Cmax of 0.949 (90% CI, 0.650–1.39) and 0.888 (90% CI, 0.602–1.31), respectively; thus, exposure to exemestane was essentially the same in these two groups.

Figure 2.

Mean plasma concentration–time profiles for exemestane and anastrozole. A, Exemestane 25 mg/day alone (exemestane 25 mg alone; n = 13) and exemestane 50 mg/day in combination with enzalutamide (enzalutamide + exemestane 50 mg; n = 16). B, Anastrozole 1 mg/day alone (anastrozole 1 mg alone; n = 15) and in combination with enzalutamide (enzalutamide + anastrozole 1 mg; n = 15.

Figure 2.

Mean plasma concentration–time profiles for exemestane and anastrozole. A, Exemestane 25 mg/day alone (exemestane 25 mg alone; n = 13) and exemestane 50 mg/day in combination with enzalutamide (enzalutamide + exemestane 50 mg; n = 16). B, Anastrozole 1 mg/day alone (anastrozole 1 mg alone; n = 15) and in combination with enzalutamide (enzalutamide + anastrozole 1 mg; n = 15.

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Assessments of drug–drug interactions where enzalutamide is the target typically focus on the sum of enzalutamide plus the active metabolite, which corresponds to total exposure to enzalutamide-active moieties (26). Steady-state Cmin values for enzalutamide plus its active metabolite were slightly lower in the exemestane cohorts (Supplementary Table S2), but the sample sizes were small and Cmin values were not significantly different.

An assessment of relationships between plasma exposures to the AIs both alone and in combination with enzalutamide and plasma concentrations of estrogens (estradiol and estrone) is summarized in Supplementary Fig. S2. For anastrozole, graphical trends suggested that an AUCtau value lower than approximately 200 ng·h/mL was associated with a higher likelihood of elevated plasma concentrations of estradiol but not estrone. For exemestane, graphical trends suggested a possible association between an AUCtau value lower than approximately 150 ng·h/mL and a higher likelihood of elevation in estradiol and estrone. Similar to the impact of enzalutamide on PK parameters, estradiol and estrone concentrations varied more significantly in the anastrozole combination cohort than in the exemestane combination cohorts (Table 2 and Supplementary Fig. S2).

Safety

Treatment-related AEs reported in at least 10% of patients are shown in Table 3. As previously stated, 1 patient treated with enzalutamide 80 mg/day monotherapy experienced a grade 3 DLT of adrenal insufficiency. The most common enzalutamide-related AEs with monotherapy were nausea (34%), fatigue (28%), vomiting (17%), increased aspartate aminotransferase levels (14%), diarrhea (14%), dizziness (10%), and hot flush (10%). The most common treatment-related AEs as assessed by the treating investigator in the combination cohorts were fatigue (53%), nausea (46%), decreased appetite (29%), vomiting (17%), hot flush (17%), diarrhea (13%), weight decrease (11%), cognitive disorder (11%), and insomnia (10%). The most common grade 3 or higher AEs, regardless of causality, were anemia (7%) with enzalutamide monotherapy and hypertension (7%), fatigue (6%), and neutropenia (4%) in the combination cohorts (Supplementary Table S3). Serious AEs considered to possibly be related to enzalutamide occurred in 5 patients (17%) treated with enzalutamide monotherapy and 10 patients (14%) treated with any ET combination therapy. AEs resulting in treatment discontinuation included obstructive uropathy (n = 1, enzalutamide monotherapy cohort), fatigue (n = 2, anastrozole combination cohort), fatigue plus nausea (n = 1, exemestane 25 mg/day combination cohort), fatigue plus arthritis (n = 1, exemestane 50 mg/day combination cohort), and anorexia (n = 1, fulvestrant combination cohort).

Table 3.

Treatment-emergent–related adverse events (reported in ≥10% of patients treated with enzalutamide monotherapy or in combination with any ET)

Enza monotherapyAnastrozole (1 mg/d) + enzaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enzaTotal (enza + ET)
Adverse event, n (%)(n = 29)(n = 20)(n = 16)(n = 23)(n = 11)(n = 70)
Total 24 (83) 20 (100) 14 (88) 18 (78) 10 (91) 62 (89) 
 Fatigue 8 (28) 12 (60) 8 (50) 9 (39) 8 (73) 37 (53) 
 Nausea 10 (34) 9 (45) 7 (44) 9 (39) 7 (64) 32 (46) 
 Decreased appetite 2 (7) 10 (50) 4 (25) 4 (17) 2 (18) 20 (29) 
 Vomiting 5 (17) 3 (15) 2 (13) 5 (22) 2 (18) 12 (17) 
 Hot flush 3 (10) 4 (20) 2 (13) 4 (17) 2 (18) 12 (17) 
 Diarrhea 4 (14) 4 (20) 1 (6) 1 (4) 3 (27) 9 (13) 
 Cognitive disorder 2 (7) 1 (5) 3 (19) 0 (0) 4 (36) 8 (11) 
 Decreased weight 2 (7) 2 (10) 3 (19) 1 (4) 2 (18) 8 (11) 
 Constipation 1 (3) 2 (10) 1 (6) 2 (9) 2 (18) 7 (10) 
 Insomnia 0 (0) 3 (15) 2 (13) 1 (4) 1 (9) 7 (10) 
 Headache 1 (3) 2 (10) 1 (6) 1 (4) 2 (18) 6 (9) 
 Dizziness 3 (10) 2 (10) 0 (0) 2 (9) 1 (9) 5 (7) 
 Increased AST 4 (14) 1 (5) 0 (0) 0 (0) 0 (0) 1 (1) 
Enza monotherapyAnastrozole (1 mg/d) + enzaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enzaTotal (enza + ET)
Adverse event, n (%)(n = 29)(n = 20)(n = 16)(n = 23)(n = 11)(n = 70)
Total 24 (83) 20 (100) 14 (88) 18 (78) 10 (91) 62 (89) 
 Fatigue 8 (28) 12 (60) 8 (50) 9 (39) 8 (73) 37 (53) 
 Nausea 10 (34) 9 (45) 7 (44) 9 (39) 7 (64) 32 (46) 
 Decreased appetite 2 (7) 10 (50) 4 (25) 4 (17) 2 (18) 20 (29) 
 Vomiting 5 (17) 3 (15) 2 (13) 5 (22) 2 (18) 12 (17) 
 Hot flush 3 (10) 4 (20) 2 (13) 4 (17) 2 (18) 12 (17) 
 Diarrhea 4 (14) 4 (20) 1 (6) 1 (4) 3 (27) 9 (13) 
 Cognitive disorder 2 (7) 1 (5) 3 (19) 0 (0) 4 (36) 8 (11) 
 Decreased weight 2 (7) 2 (10) 3 (19) 1 (4) 2 (18) 8 (11) 
 Constipation 1 (3) 2 (10) 1 (6) 2 (9) 2 (18) 7 (10) 
 Insomnia 0 (0) 3 (15) 2 (13) 1 (4) 1 (9) 7 (10) 
 Headache 1 (3) 2 (10) 1 (6) 1 (4) 2 (18) 6 (9) 
 Dizziness 3 (10) 2 (10) 0 (0) 2 (9) 1 (9) 5 (7) 
 Increased AST 4 (14) 1 (5) 0 (0) 0 (0) 0 (0) 1 (1) 

Abbreviations: AST, aspartate aminotransferase; enza, enzalutamide.

Antitumor activity

Antitumor activity was assessed in patients enrolled in stage 2 of the study. Two patients (14%) in the enzalutamide monotherapy cohort and 14 patients (20%) in the combination ET cohort achieved clinical benefit, defined as confirmed complete or partial response or stable disease for at least 16 weeks' duration. The clinical benefit rate at 24 weeks was 7% (1/14) and 9% (6/70) in the enzalutamide monotherapy and combination ET cohorts, respectively (Table 4). One patient who received enzalutamide combined with exemestane 25 mg followed by exemestane 50 mg/day experienced stable disease for more than 3 years.

Table 4.

Clinical benefit as assessed by investigators per RECIST in stage 2

Enza monotherapyAnastrozole (1 mg/d) + enzaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enza
(n = 14)(n = 20)(n = 16)(n = 23)(n = 11)
Patients with ≥1 postbaseline tumor assessment, n (%) 13 (93) 16 (80) 15 (94) 20 (87) 10 (91) 
CBR 16, n 
 % (95% CI) 14 (3–42) 10 (2–32) 13 (2–37) 22 (9–43) 46 (20–74) 
CBR 24, n 
 % (95% CI) 7 (0.3–31) 5 (0.3–24) 6 (0.3–30) 9 (2–27) 18 (3–50) 
Enza monotherapyAnastrozole (1 mg/d) + enzaExemestane (25 mg/d) + enzaExemestane (50 mg/d) + enzaFulvestrant (500 mg/mo) + enza
(n = 14)(n = 20)(n = 16)(n = 23)(n = 11)
Patients with ≥1 postbaseline tumor assessment, n (%) 13 (93) 16 (80) 15 (94) 20 (87) 10 (91) 
CBR 16, n 
 % (95% CI) 14 (3–42) 10 (2–32) 13 (2–37) 22 (9–43) 46 (20–74) 
CBR 24, n 
 % (95% CI) 7 (0.3–31) 5 (0.3–24) 6 (0.3–30) 9 (2–27) 18 (3–50) 

Abbreviations: CBR, clinical benefit rate defined as complete response, partial response, or stable disease; CBR 16, CBR at 16 weeks; CBR 24, CBR at 24 weeks; enza, enzalutamide.

This study represents the first investigation of enzalutamide in women with breast cancer. The purpose was to characterize the PK of enzalutamide alone and in combination with ETs, as well as to assess the safety and tolerability of these treatments in order to support future enzalutamide development in ER+/PgR+ breast cancer. Assessment of antitumor activity in this study was exploratory.

Enzalutamide is a strong CYP3A4 inducer, and ETs are commonly metabolized by CYP3A4. Therefore, this investigation also evaluated effects of enzalutamide on PK and PD of ETs that are potential CYP3A4 substrates: anastrozole, exemestane, and fulvestrant.

The PK of enzalutamide has been well characterized in men (27). Following oral administration in prostate cancer patients, enzalutamide has a mean terminal half-life of 5.8 days and takes about 28 days to reach plasma steady state with daily dosing. The estimated terminal half-life appeared to be slightly longer in women with breast cancer; however, these estimates of half-life are not considered robust, as the duration of PK sampling (approximately 7 days) was shorter than the estimated mean values (8.25 to 11.7 days), and the coefficients of variation were large (52.9% to 60.6%; Supplementary Table S1). Based on mean values from this study for Cmin, Cmax, and AUCtau for enzalutamide administered in the absence of ETs, plasma exposure to enzalutamide and its active metabolite at 160 mg/day is essentially the same in women and men (Supplementary Tables S1 and S2), supporting a dose of 160 mg/day in women.

PK data from the present investigation showed that concomitant enzalutamide led to an 89% decrease in mean plasma anastrozole exposure, a 43% to 57% decrease in mean exemestane exposure, and an 8% decrease in mean fulvestrant exposure. The reduced exposure to anastrozole and exemestane was considered quantitatively and possibly clinically significant, whereas the reduction in fulvestrant exposure was not (25). These results are consistent with prior data on the drug–drug interaction liabilities of anastrozole, exemestane, and fulvestrant (15, 17, 28). In addition, the approximate 50% decrease in exemestane exposure with concomitant enzalutamide is consistent with a 54% decrease reported with concomitant exemestane and a strong CYP3A4 inducer, rifampin (17). The U.S. product label for exemestane recommends doubling the dose when taken concomitantly with a strong CYP3A4 inducer (17). In our study, doubling the dose of exemestane from 25 mg/day to 50 mg/day with concomitant enzalutamide restored exposures consistent with those of exemestane monotherapy. The lack of a clinically meaningful change in fulvestrant exposure during concomitant enzalutamide treatment is also consistent with prior data showing that concomitant rifampin did not affect fulvestrant exposure (15), and a previous analysis that concluded fulvestrant is unlikely to be the subject of significant CYP3A4-mediated drug interactions (29).

Recently published experiments on in vitro metabolism of exemestane suggest that CYP3A4-mediated metabolites may be more potent than exemestane at suppressing the in vitro growth of breast cancer cells (30, 31). Importantly, however, there is no indication that these findings translate to clinical effects. As reported in the exemestane prescribing information (17), a radiolabeled mass balance study in patients showed that exemestane is extensively metabolized, and each metabolite accounts for only a limited amount of drug-related material. Correspondingly, plasma exposure to any given metabolite is not considered quantitatively important. While it is possible that the CYP3A4 enzyme–inducing effects of enzalutamide may increase the rate of exemestane metabolite formation, it is also possible that the metabolites themselves are substrates of CYP3A4 or any number of other enzymes induced by enzalutamide. Therefore, increased metabolic turnover of exemestane does not necessarily lead to increased exposure to biologically active exemestane metabolites in patients. Accordingly, the CYP3A4 enzyme–inducing effects of enzalutamide may or may not cause increased exposure to exemestane metabolites in patients, and it is uncertain whether these metabolites have in vivo pharmacologic effects that are relevant to breast cancer treatment.

In postmenopausal women, estrogens are predominantly produced by aromatase, which converts adrenal androgens to estradiol and estrone. Aromatase is mainly expressed in peripheral tissues and can also be expressed in tumors, where estrogens may promote tumor growth (32, 33). Suppression of estrogen biosynthesis is achieved with aromatase inhibition. The PD effects of AIs are commonly assessed through circulating concentrations of estradiol and estrone. Because of significant local estrogen production in some tumors, plasma estrogens may not reflect estrogen levels in breast cancer tumors (34). Nevertheless, plasma concentrations of estradiol and estrone present a rough estimate of total body production of estrogen (35) and are useful for assessing gross changes in the PD effects of AIs. As expected, administration of anastrozole or exemestane was generally associated with estradiol plasma concentrations less than 2.0 pg/mL, which is assumed to indicate a therapeutic level of aromatase inhibition (36).

In patients in the anastrozole cohort, concomitant use of enzalutamide led to a 40% and 68% relative increase in mean plasma concentrations of estradiol and estrone, respectively (Table 2). Given the magnitude of these changes, and that the 90% CI upper bounds for the geometric means did not include the value 1.00, the apparent decrease in the PD effects of anastrozole is considered quantitatively significant. However, it is less certain whether enzalutamide led to a clinically meaningful change in the pharmacologic effects of anastrozole. There was only a weak association between plasma exposure to anastrozole and increases in estradiol greater than 1 pg/mL (3.7 pmol/L), and there was no clear trend to support that lower exposure to anastrozole was associated with an increase in estrone (Supplementary Fig. S1; ref. 28). PD data from the two exemestane cohorts (25 mg/day and 50 mg/day) showed that concomitant use of enzalutamide led to an 8% to 13% relative increase in the mean plasma estradiol concentrations, whereas mean estrone increased 16% in the 25 mg/day cohort and increased 3% in the 50 mg/day cohort. None of these changes was considered quantitatively or clinically significant, as the value of 1.00 was included in the 90% CI.

Enzalutamide monotherapy or in combination with ETs was generally well tolerated; although AEs appeared similar across the combination therapies, a rigorous comparison of particular combination treatments was precluded by the small number of patients in each cohort. The addition of enzalutamide to AIs appeared to be well tolerated, as the incidence of AEs associated with enzalutamide was similar to those reported in prostate cancer patients receiving enzalutamide monotherapy (6). Importantly, no seizures or new safety signals were observed. The most common AEs were those commonly reported with ETs.

Evaluation of AR expression by IHC was an exploratory endpoint in this study. The rate of AR positivity (defined as AR ≥10%) was consistent with published data (2). Given the small sample size in this study, the adequacy of IHC as a companion diagnostic for enzalutamide treatment in ER+/PgR+ breast cancer remains to be determined.

Although there were limited efficacy data, the safety, PK, and PD results support further evaluation of oral enzalutamide 160 mg/day either as monotherapy or in combination with either exemestane 50 mg/day or intramuscular fulvestrant 500 mg/month in ER+/PgR+ breast cancer. The PK profile of enzalutamide in women is similar to that in men. PD effects of AI therapy, as assessed by changes in circulating hormone levels, were preserved in the presence of enzalutamide when given in combination with exemestane. Enzalutamide monotherapy and in combination with AIs (either anastrozole, exemestane, or fulvestrant) appears to be well tolerated. Our study results provide support for continued enzalutamide development in breast cancer patients and directly informed the design of the ongoing randomized, double-blind, multicenter phase II study of exemestane 25 mg/day plus placebo versus exemestane 50 mg/day plus enzalutamide 160 mg/day in patients with advanced ER+/PgR+ breast cancer (NCT02007512; ref. 37).

M. Patel reports receiving speakers bureau honoraria from Medivation. A. Gucalp reports receiving commercial research grants from Innocrin and Pfizer. T.A. Traina is a consultant/advisory board member for Bayer and Medivation, and reports receiving commercial research grants from Innocrin and Medivation. No potential conflicts of interest were disclosed by the other authors.

Conception and design: L.S. Schwartzberg, A.D. Elias, P. LoRusso, H.A. Burris, A. Gucalp, A.C. Peterson, M.E. Blaney, J.L. Steinberg, J.A. Gibbons, T.A. Traina

Development of methodology: A.D. Elias, H.A. Burris, A. Gucalp, A.C. Peterson, M.E. Blaney, J.L. Steinberg, J.A. Gibbons, T.A. Traina

Acquisition of data (provided animals, acquired and managed patients, provided facilities, etc.): L.S. Schwartzberg, D.A. Yardley, A.D. Elias, M. Patel, P. LoRusso, H.A. Burris, A. Gucalp, A.C. Peterson, M.E. Blaney, T.A. Traina

Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis): L.S. Schwartzberg, D.A. Yardley, A.D. Elias, M. Patel, P. LoRusso, H.A. Burris, A.C. Peterson, M.E. Blaney, J.A. Gibbons, T.A. Traina

Writing, review, and/or revision of the manuscript: L.S. Schwartzberg, D.A. Yardley, A.D. Elias, M. Patel, P. LoRusso, H.A. Burris, A. Gucalp, A.C. Peterson, M.E. Blaney, J.L. Steinberg, J.A. Gibbons, T.A. Traina

Administrative, technical, or material support (i.e., reporting or organizing data, constructing databases): A.C. Peterson, M.E. Blaney

Study supervision: L.S. Schwartzberg, A.D. Elias, M. Patel, A.C. Peterson, M.E. Blaney, T.A. Traina

This study was funded by Medivation, Inc., and Astellas Pharma, Inc., the codevelopers of enzalutamide (Medivation, Inc., was acquired by Pfizer, Inc., in September 2016). Data analysis was provided by Iulia Cristina Tudor, PhD, and Rachel Wei, PhD, both employees of Medivation, Inc. Medical writing and editorial support funded by both sponsor companies were provided by Stephanie Vadasz, PhD, and Paula Stuckart of Ashfield Healthcare Communications.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1.
Coss
CC
,
Jones
A
,
Dalton
JT
. 
Selective androgen receptor modulators as improved androgen therapy for advanced breast cancer
.
Steroids
2014
;
90
:
94
100
.
2.
Collins
LC
,
Cole
KS
,
Marotti
JD
,
Hu
R
,
Schnitt
SJ
,
Tamimi
RM
. 
Androgen receptor expression in breast cancer in relation to molecular phenotype: Results from the Nurses' Health Study
.
Mod Pathol
2011
;
24
:
924
31
.
3.
Chen
CD
,
Welsbie
DS
,
Tran
C
,
Baek
SH
,
Chen
R
,
Vessella
R
, et al
Molecular determinants of resistance to antiandrogen therapy
.
Nat Med
2004
;
10
:
33
9
.
4.
Tran
C
,
Ouk
S
,
Clegg
NJ
,
Chen
Y
,
Watson
PA
,
Arora
V
, et al
Development of a second-generation antiandrogen for treatment of advanced prostate cancer
.
Science
2009
;
324
:
787
90
.
5.
Guerrero
J
,
Alfaro
IE
,
Gomez
F
,
Protter
AA
,
Bernales
S
. 
Enzalutamide, an androgen receptor signaling inhibitor, induces tumor regression in a mouse model of castration-resistant prostate cancer
.
Prostate
2013
;
73
:
1291
305
.
6.
Xtandi [package insert].
Astellas Pharma US, Inc.
,
San Francisco, CA
; 
2014
.
7.
De Amicis
F
,
Thirugnansampanthan
J
,
Cui
Y
,
Selever
J
,
Beyer
A
,
Parra
I
, et al
Androgen receptor overexpression induces tamoxifen resistance in human breast cancer cells
.
Breast Cancer Res Treat
2010
;
121
:
1
11
.
8.
Rechoum
Y
,
Rovito
D
,
Iacopetta
D
,
Barone
I
,
Ando
S
,
Weigel
NL
, et al
AR collaborates with ERalpha in aromatase inhibitor-resistant breast cancer
.
Breast Cancer Res Treat
2014
;
147
:
473
85
.
9.
Cochrane
DR
,
Bernales
S
,
Jacobsen
BM
,
Cittelly
DM
,
Howe
EN
,
D'Amato
NC
, et al
Role of the androgen receptor in breast cancer and preclinical analysis of enzalutamide
.
Breast Cancer Res
2014
;
16
:
R7
.
10.
Ciupek
A
,
Rechoum
Y
,
Gu
G
,
Gelsomino
L
,
Beyer
AR
,
Brusco
L
, et al
Androgen receptor promotes tamoxifen agonist activity by activation of EGFR in ERalpha-positive breast cancer
.
Breast Cancer Res Treat
2015
;
154
:
225
37
.
11.
Gallicchio
L
,
Macdonald
R
,
Wood
B
,
Rushovich
E
,
Helzlsouer
KJ
. 
Androgens and musculoskeletal symptoms among breast cancer patients on aromatase inhibitor therapy
.
Breast Cancer Res Treat
2011
;
130
:
569
77
.
12.
Campagnoli
C
,
Pasanisi
P
,
Castellano
I
,
Abba
C
,
Brucato
T
,
Berrino
F
. 
Postmenopausal breast cancer, androgens, and aromatase inhibitors
.
Breast Cancer Res Treat
2013
;
139
:
1
11
.
13.
Elias
A
,
Burris
H
,
Patel
M
,
Schwartzberg
L
,
Richer
J
,
Kavalerchik
E
, et al
Abstract P1-16-05: MDV3100-08: A phase 1 study evaluating the safety and pharmacokinetics of enzalutamide plus fulvestrant in women with advanced hormone receptor-positive breast cancer
.
Cancer Res
2016
;
76
:
P1-16-05
.
14.
Wakeling
AE
,
Dukes
M
,
Bowler
J
. 
A potent specific pure antiestrogen with clinical potential
.
Cancer Res
1991
;
51
:
3867
73
.
15.
Faslodex [package insert]
.
AstraZeneca Pharmaceuticals LP
,
Wilmington, DE
; 
2012
.
16.
Milani
M
,
Jha
G
,
Potter
DA
. 
Anastrozole use in early stage breast cancer of post-menopausal women
.
Clin Med Ther
2009
;
1
:
141
56
.
17.
Aromasin [package insert]
.
Pfizer, Inc.
,
New York, NY
; 
2014
.
18.
Gucalp
A
,
Tolaney
S
,
Isakoff
SJ
,
Ingle
JN
,
Liu
MC
,
Carey
LA
, et al
Phase II trial of bicalutamide in patients with androgen receptor-positive, estrogen receptor-negative metastatic breast cancer
.
Clin Cancer Res
2013
;
19
:
5505
12
.
19.
Bonnefoi
H
,
Grellety
T
,
Tredan
O
,
Saghatchian
M
,
Dalenc
F
,
Mailliez
A
, et al
A phase II trial of abiraterone acetate plus prednisone in patients with triple-negative androgen receptor positive locally advanced or metastatic breast cancer (UCBG 12-1)
.
Ann Oncol
2016
;
27
:
812
8
.
20.
Ingle
JN
,
Buzdar
AU
,
Schaid
DJ
,
Goetz
MP
,
Batzler
A
,
Robson
ME
, et al
Variation in anastrozole metabolism and pharmacodynamics in women with early breast cancer
.
Cancer Res
2010
;
70
:
3278
86
.
21.
Corona
G
,
Elia
C
,
Casetta
B
,
Diana
C
,
Rosalen
S
,
Bari
M
, et al
A liquid chromatography-tandem mass spectrometry method for the simultaneous determination of exemestane and its metabolite 17-dihydroexemestane in human plasma
.
J Mass Spectrom
2009
;
44
:
920
8
.
22.
Bennett
D
,
Gibbons
JA
,
Mol
R
,
Ohtsu
Y
,
Williard
C
. 
Validation of a method for quantifying enzalutamide and its major metabolites in human plasma by LC-MS/MS
.
Bioanalysis
2014
;
6
:
737
44
.
23.
Balaram
VM
,
Parmar
D
,
Teja
BB
,
Rathnam
S
,
Rao
JV
,
Dasandi
B
. 
Sensitive and rapid high-performance liquid chromatography tandem mass spectrometry method for estimation of fulvestrant in rabbit plasma
.
Biomed Chromatogr
2010
;
24
:
863
7
.
24.
National Cancer Institute (NCI). Common Terminology Criteria for Adverse Events v4.0
.
Bethesda (MD)
:
NCI, National Institutes of Health (NIH), Department of Health and Human Services
;
May 29, 2009. NIH Publication No.: 09-7473
.
25.
Guidance for industry. Drug interaction studies - study design, data analysis, implications for dosing, and labeling recommendations [Internet]
.
Rockville (MD)
:
U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research
;
[cited 2017 May 5]. Available from
https://www.fda.gov/downloads/drugs/guidances/ucm292362.pdf.
26.
Gibbons
JA
,
de Vries
M
,
Krauwinkel
W
,
Ohtsu
Y
,
Noukens
J
,
van der Walt
JS
, et al
Pharmacokinetic drug interaction studies with enzalutamide
.
Clin Pharmacokinet
2015
;
54
:
1057
69
.
27.
Gibbons
JA
,
Ouatas
T
,
Krauwinkel
W
,
Ohtsu
Y
,
van der Walt
JS
,
Beddo
V
, et al
Clinical pharmacokinetic studies of enzalutamide
.
Clin Pharmacokinet
2015
;
54
:
1043
55
.
28.
Arimidex [package insert]
.
AstraZeneca Pharmaceuticals LP,
Wilmington, DE
; 
2011
.
29.
Robertson
JF
,
Erikstein
B
,
Osborne
KC
,
Pippen
J
,
Come
SE
,
Parker
LM
, et al
Pharmacokinetic profile of intramuscular fulvestrant in advanced breast cancer
.
Clin Pharmacokinet
2004
;
43
:
529
38
.
30.
Amaral
C
,
Lopes
A
,
Varela
CL
,
da Silva
ET
,
Roleira
FM
,
Correia-da-Silva
G
, et al
Exemestane metabolites suppress growth of estrogen receptor-positive breast cancer cells by inducing apoptosis and autophagy: A comparative study with Exemestane
.
Int J Biochem Cell Biol
2015
;
69
:
183
95
.
31.
Varela
CL
,
Amaral
C
,
Tavares da Silva
E
,
Lopes
A
,
Correia-da-Silva
G
,
Carvalho
RA
, et al
Exemestane metabolites: Synthesis, stereochemical elucidation, biochemical activity and anti-proliferative effects in a hormone-dependent breast cancer cell line
.
Eur J Med Chem
2014
;
87
:
336
45
.
32.
Reed
MJ
. 
The role of aromatase in breast tumors
.
Breast Cancer Res Treat
1994
;
30
:
7
17
.
33.
Brueggemeier
RW
,
Hackett
JC
,
Diaz-Cruz
ES
. 
Aromatase inhibitors in the treatment of breast cancer
.
Endocr Rev
2005
;
26
:
331
45
.
34.
Geisler
J
. 
Breast cancer tissue estrogens and their manipulation with aromatase inhibitors and inactivators
.
J Steroid Biochem Mol Biol
2003
;
86
:
245
53
.
35.
Lonning
PE
,
Geisler
J
. 
Aromatase inhibitors: Assessment of biochemical efficacy measured by total body aromatase inhibition and tissue estrogen suppression
.
J Steroid Biochem Mol Biol
2008
;
108
:
196
202
.
36.
Geisler
J
,
Haynes
B
,
Anker
G
,
Dowsett
M
,
Lonning
PE
. 
Influence of letrozole and anastrozole on total body aromatization and plasma estrogen levels in postmenopausal breast cancer patients evaluated in a randomized, cross-over study
.
J Clin Oncol
2002
;
20
:
751
7
.
37.
ClinicalTrials.gov [Internet].
Bethesda (MD):
National Library of Medicine (US)
.
Identifier NCT02007512
, 
Efficacy and safety study of enzalutamide in combination with exemestane in patients with advanced breast cancer
.
[cited 2017 May 5]. Available from:
https://clinicaltrials.gov/ct2/show/NCT02007512.

Supplementary data