This study was a multi-institutional single-arm prospective trial designed in collaboration with the principal investigator and Translational Research Center for Medical Innovation, Foundation for Biomedical Research and Innovation at Kobe (TRI), and funded by Astellas Pharma, the developer of enzalutamide. It was conducted according to the Declaration of Helsinki and Clinical Trials Act, the Japanese law outlining the ethical conduct of clinical trials. The ethics review committee of Kagawa University Hospital, TRI, the individuals, and institutional review boards of all the participating facilities approved this study (trial registration: UMIN000018964, first registration: September 10, 2015). The clinical trials review board of Kagawa University Hospital also approved and certified this study in line with the provisions of the Clinical Trials Act (trial registration: CRB6180007). Informed consent was obtained from all patients. The protocol was listed at a Additional files 1 and 2.

The inclusion criteria were as follows: (1) Patients aged 20 years or older with histologically confirmed PC; (2) patients with a history of radical prostatectomy or radiation therapy for radical treatment; (3) patients who received continuous ADT using either luteinizing hormone-releasing hormone (LHRH) agonist, antagonist, or surgical castration; (4) patients with a serum testosterone level of 1.73 nmol/L (0.50 ng/dL) or lower; (5) patients with a history of bicalutamide or flutamide use after the confirmation of the first recurrence since the completion of radical treatment; (6) patients with three test results showing increased PSA levels, which were measured consecutively at an interval of at least 1 week during ADT; (7) patients with a serum PSA level of 1 ng/mL or higher (2 ng/mL or more before August 23, 2017); (8) patients with no confirmed remote metastasis on CT and bone scintigraphy after the diagnosis of PC (excluding lymph node metastasis with a minor axis of less than 15 mm, which was considered non-measurable in the Response Evaluation Criteria In Solid Tumors version 1.1); (9) patients with asymptomatic PC; (10) patients with an Eastern Cooperative Oncology Group performance status of 0–1; and (11) patients with a life expectancy of at least 12 months.

The exclusion criteria were as follows: (1) patients with a history of any chemotherapy (including estramustine phosphate sodium hydrate and docetaxel) or treatment with enzalutamide or abiraterone acetate; (2) patients with a history of steroid use as a treatment for PC; (3) patients with a history of 5-alpha-reductase inhibitor, estrogen, or steroidal antiandrogen therapy within 4 weeks before the initial administration of enzalutamide; (4) patients with a history of malignant tumor other than PC within the past 3 years; (5) patients with a history of seizures or pre-disposed to seizures; (6) patients with severe liver dysfunction; and (7) patients with a history of hypersensitivity to any component of the drugs administered in this study.

All patients received enzalutamide 160 mg orally once daily. The treatment was started at visit 0 within 1 week after enrollment. Visit 1 occurred 2 weeks after the start of treatment; clinical assessments were conducted for adverse events (AEs) by using the Japanese version of the Functional Assessment of Cancer Therapy-Prostate (FACT-P) scales. PSA was measured every 3 months, and CT and bone scintigraphy was performed every 6 months. In the case of AEs of ≥ grade 3, enzalutamide could not be ruled out as a causative agent and was temporarily discontinued. However, after the AE severity reduced to grade 1 or less, enzalutamide was resumed at half the original dose (80 mg) initially. Eligible patients in this study continued the treatment until the 12th-week visit (counted from the administration of the initial dose) until they met the withdrawal or protocol treatment discontinuation criteria. The outcomes for these patients were evaluated for 2 years from the final patient enrollment.

The primary endpoint was PSA-progression-free survival (PSA-PFS), defined as the time from enrollment to PSA-based progression or death from any cause. PSA-based progression was defined as an increase in PSA levels of > 25% relative to the nadir PSA level and higher than 2 ng/mL [15].

The secondary endpoints included OS, defined as the time from enrollment to death from any cause, PFS, and MFS. PFS was defined as the time from enrollment to investigator-assessed disease progression (based on imaging or symptom development) or death from any cause. MFS was defined as the time from enrollment to disease progression assessed by CT and bone scintigraphy by investigator review. Other secondary endpoints were a 50% PSA response rate (less than half the baseline PSA concentration), chemotherapy-free survival, and quality of life (QOL) assessment using the Japanese version of the FACT-P scales. A safety assessment of the frequency and severity of AEs was performed using the Common Terminology Criteria for Adverse Events (CTCAE) version 4.0 [16].

Information on patients’ demographic and disease characteristics; interventions; laboratory tests; and outcome evaluations, including PSA, imaging, and survival, were obtained by investigators at each participating institute via a filled-in form on the website prepared by TRI. The PSA response rates at 2 and 12 weeks after the start of treatment were reported as the number and percent of patients with 95% confidence intervals (CI). The health-related QOL (assessed using the FACT-P scales) data at baseline and at 2 and 60 weeks after the start of treatment were collected.

We carried out univariate analyses for time-to-event data in primary and secondary endpoints by using the Kaplan–Meier method; the median survival time and 95% CI were estimated. We also summarized patient characteristics with frequencies and percentages for categorical variables or with medians and ranges for continuous variables.

As an ad-hoc analysis, we investigated the PSA concentrations in the follow-up period as potential predictors of OS or PFS using a Cox proportional hazard model with PSA as a time-dependent variate and baseline age as a covariate. Furthermore, we performed a restricted cubic spline regression (RCSR) with three knots, assuming the Cox proportional hazard ratio could be nonlinearly changed according to the changes in PSA concentrations [17]. This model evaluated PSA concentrations not only at baseline but also at other each patient’s visit until the patient experienced the outcome event or was censored. The aim of this approach was to test the following null hypothesis: PSA concentration curves over time were identical between censored and event-experienced patients. If the null hypothesis is rejected, it would indicate that event-experienced patients had characteristic changes in PSA concentrations over time, which were different from those of censored or survived patients. We used SAS statistical software (version 9.3, SAS Institute Inc., Cary, NC, USA), R4.2.1 (R Foundation), and R library rms for the RCSR.

The sample size was determined to be 60 evaluable subjects for the feasibility of this study. However, with a null hypothesis that states the practical lowest effective limit of the median PSA-PFS time was 13 months and considering that the intervention could achieve 24 months of the median PSA-PFS time, the sample size could reject the null hypothesis with a power of 0.87 and 5% significance level in a two-sided test.