Safety and tolerability of bireociclib for the treatment of advanced solid tumors as monotherapy and in combination with endocrine therapy: a multicenter, open-label, phase 1 clinical trial

Abstract

Background

Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors have been approved for the treatment of hormone receptor-positive (HR⁺) and human epidermal growth factor receptor-2 negative (HER2⁻) breast cancers. The study aims were to evaluate the safety, tolerability, and antitumor activity of the novel selective small molecule CDK4/6 inhibitor bireociclib in advanced solid/breast cancer (ABC) patients.

Methods

A multicenter, open-label, phase 1 trial which consisted of two parts. Part 1 evaluated bireociclib monotherapy at doses ranging from 20 mg once-daily (QD) to 480 mg twice-daily (BID), and part 2 evaluated bireociclib at the recommended dosage in combination with endocrine therapy for ABC patients, including bireociclib plus non-steroidal aromatase inhibitor (cohort A), bireociclib plus fulvestrant as first-line (cohort B), or second-line therapy (cohort C). The endpoints included observing dose-limiting toxicities (DLTs) and the maximum tolerated dose (MTD), as well as determining recommended phase 2 dosage for a single regimen (RP2D-S) and recommended phase 2 dosage for combination therapy (RP2D-C) and assessment of the general safety and efficacy of therapy.

Results

Thirty-five patients were included in the part 1 MTD analysis and no DLTs occurred. Bireociclib 480 mg BID had more stable blood concentration fluctuations as well as a superior tumor response rate (objective response rate [ORR], 17.5%), which was identified as RP2D-S. In part 2, 6 patients each were enrolled in the combination study to assess MTD. One DLT occurred (a grade 3 hepatic enzyme increase), so RP2D-C was determined to be 360 mg BID. The highest incidence of any grade and grade 3 or 4 treatment-emergent adverse events in both part 1 and part 2 were diarrhea, neutropenia, and leukopenia. During the dose expansion phase in part 2, the ORR reached 57.1%, 57.1%, and 46.3% in cohorts A, B, and C, respectively.

Conclusions

Bireociclib demonstrated favorable efficacy and an acceptable safety profile both as monotherapy (RP2D-S of 480 mg BID) and in combination therapy (RP2D-C of 360 mg BID) for treating HR⁺/HER2⁻ ABC patients.

Trial registration

Registered with ClinicalTrials.gov, identification ID: NCT04539496, registration date: 03/09/2020 (retrospectively registered).

Supplementary Information

The online version contains supplementary material available at 10.1186/s12916-025-04364-9.

Background

In solid tumors, mutations in genes related to cyclin-dependent kinase/retinoblastoma-protein (CDK/Rb) signaling are relatively common and lead to dysregulation of the cell cycle [1]. Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors, developed to target this aberrant signaling pathway in cancer, have been approved for the treatment of hormone receptor-positive (HR⁺) and human epidermal growth factor receptor-2 negative (HER2⁻) breast cancers [2, 3]. Furthermore, since estrogen receptor-positive (ER⁺) breast cancers typically feature high expression of the ER target gene CCND1 and functional Rb, making them particularly sensitive to CDK4/6 inhibitors, which block cell cycle progression by preventing Rb phosphorylation downstream of the ER-CyclinD1 axis [4]. Preclinical studies have shown strong synergistic effects when CDK4/6 inhibitors were added to standard anti-estrogen therapy, delaying and reversing endocrine resistance [5, 6]. CDK4/6 inhibitors (palbociclib, ribociclib, abemaciclib, dalpiciclib) combined with endocrine therapy, specifically non-steroidal aromatase inhibitors (NSAIs) in the first-line setting and fulvestrant (FULV) in the second-line setting, have received approval for HR⁺/HER2⁻ advanced or metastatic breast cancer treatment, although the specific indications and approved combinations may vary among the individual drugs. These combinations have produced significant improvements in both progression-free survival (PFS) and overall survival (OS) in patients with HR⁺/HER2⁻ advanced breast cancer (ABC) [7–9].

Bireociclib is an orally selective small molecule inhibitor of CDK4 (half maximal inhibitory concentration [IC₅₀], 3 nM) and CDK6 (IC₅₀, 56 nM), and its activity against cyclin-dependent kinase subtypes is comparable to that of abemaciclib [10]. Consistent with its activity against CDK4/6, bireociclib decreases Rb protein phosphorylation at the Ser 780 site and reduces total Rb protein levels in tumors [11–13]. The preclinical characterization of pharmacokinetic and pharmacodynamic (PK/PD) analyses in plasma and tumor tissues revealed that bireociclib was widely distributed including in brain tissue [14]. In humans, the plasma protein binding rate of bireociclib was determined to be 81.9%, underscoring its higher free drug concentration.

The phase 1 portion of the phase 1/2 trial NCT04539496 investigated bireociclib in patients with advanced solid tumors, particularly those with HR⁺/HER2⁻ ABC, and was followed by a phase 2 monotherapy cohort [15]. Here we report the safety and efficacy of bireociclib for the treatment of advanced solid tumors as monotherapy and for HR⁺/HER2⁻ ABC in combination with endocrine therapy in the phase 1 trial.

Methods

Study design

The present open-label, multicenter, multiple-dose phase 1 clinical trial (NCT04539496) consisted of two parts: monotherapy dose escalation and dose expansion in patients with advanced solid tumors and combination therapy expansion in patients with advanced HR⁺/HER2⁻ breast cancer. The trial was conducted in 15 hospitals across 10 cities, with 1 to 55 patients enrolled at each hospital. Detailed information is presented in Additional file 1: Table S1.

The study design is shown in Fig. 1, part 1 for monotherapy (Fig. 1a) and part 2 for combination therapy (Fig. 1b). The initial dose of bireociclib monotherapy was 20 mg once-daily (QD) based on the preclinical data. The dose-escalation scheme included accelerated titration from 20 mg QD towards 160 mg QD and then the traditional “3 + 3” design based on a modified Fibonacci scheme. If either dose-limiting toxicity (DLT) occurred or the same type of adverse events (AEs; ≥ grade 2 and related to treatment) recurred during the accelerated titration period, the dosing scheme was switched to the traditional “3 + 3” design. After completing the DLT evaluation in the 560 mg QD cohort, the administration schedule was switched to BID dosing, with dose escalation still following the “3 + 3” design. Given the equivalent daily dose of 560 mg QD would be 280 mg BID, the BID regimen was initiated at a conservative dose of 240 mg BID, followed by 360 mg BID and 480 mg BID dose groups (Fig. 1a). For the 240, 360, and 480 mg twice-daily (BID) dose levels, only HR⁺/HER2⁻ ABC patients were enrolled.

Fig. 1.

Phase 1 study design for bireociclib treatment. a) part 1 for monotherapy; b) part 2 for combination with endocrine therapy. Abbreviations:ABC advanced breast cancer; AE adverse event; ATD accelerated titration design; BID twice-daily; DLT dose-limiting toxicity; ECOG Eastern Cooperative Oncology Group; ET endocrine therapy; FULV fulvestrant 500 mg intramuscularly (on day 1 and day 15 of the first cycle and then on day 1 of each subsequent 4-week cycle); HR⁺/HER2^- hormone receptor-positive/human epidermal growth factor receptor-2 negative; NSAI non-steroidal aromatase inhibitor (letrozole 2.5 mg or anastrozole 1 mg oral QD); PK pharmacokinetic; QD once-daily; RECIST Response Evaluation Criteria in Solid Tumors; RP2D-C recommended phase 2 dosage for combination therapy; RP2D-S recommended phase 2 dosage for a single regimen

In part 2, which evaluated combination therapy, dose escalation commenced at one dose level below the established recommended phase 2 dosage for a single regimen (RP2D-S) of bireociclib monotherapy, combined with a fixed dose of endocrine therapy. Confirmation of this level as the recommended phase 2 dosage for combination therapy (RP2D-C) required the observation of ≤ 1 DLT among 6 treated patients. The confirmed RP2D-C was then utilized for the combination expansion cohort. The patients with HR⁺/HER2⁻ ABC were assigned to receive bireociclib plus NSAI or bireociclib plus FULV, based on the investigator’s decision, to determine the RP2D-C in each group. Following the determination of RP2D-C, the sample size was expanded in both treatment groups and patients were divided into cohorts A, B, and C, based on prior systemic therapy.

Combination therapy was administered in cycles of 28 days. For cohort A, patients received orally bireociclib and NSAI (letrozole 2.5 mg or anastrozole 1 mg oral QD). Cohort B received bireociclib with the same schedule as cohort A plus FULV 500 mg intramuscularly (on day 1 and day 15 of the first cycle and then on day 1 of each subsequent 4-week cycle) as first-line treatment. Cohort C patients received the same schedule as cohort B as second-line treatments, after confirmed recurrence or disease progression (PD) during or after previous endocrine therapy. The administration of bireociclib followed the RP2D-C (Fig. 1b).

Treatment continued until PD, consent withdrawal, unacceptable toxicity, investigator’s decision, initiation of new anticancer therapy, completion of the trial, or death, whichever occurred first.

All premenopausal or perimenopausal women were required to take a gonadotropin-releasing hormone analog during the trial, starting at least 14 days before the first dose of the study medication was administered.

Blood sample collections for PK analysis were acquired from all patients who participated in the dose escalation of part 1. In part 2, PK blood samples were also collected from 27 patients in the combination therapy cohort, including 12 in the NSAI group and 15 in the FULV group.

The study protocol and all amendments were approved by the Ethics Committee of the National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College (approval number: 17–163/1419), as well as by the ethics committees of all other participating hospitals. All procedures were conducted in accordance with Good Clinical Practice guidelines and the Declaration of Helsinki. All patients provided written informed consent before being enrolled in the trial.

Patient eligibility

Eligible patients for part 1 were individuals aged 18–70 years with pathologically or cytologically confirmed locally advanced, recurrent or metastatic malignant solid tumors, who had experienced either treatment failure or lacked access to standard treatment options, including those who had received neoadjuvant or adjuvant chemotherapy.

For part 2, eligible patients were women aged 18–70 years with HR⁺/HER2⁻ ABC, where HR⁺ was defined as ER⁺ and/or progesterone receptor positive with at least 1% of tumor cells showing positive staining. Patients in cohorts A and B had disease recurrence more than 12 months after completion of (neo)adjuvant therapy, or initial diagnosis of ABC and had not received any previous systemic therapy during the advanced stage. Patients in cohort C had PD that occurred ≥ 6 months after initiation of endocrine therapy for metastatic disease, or progression during adjuvant endocrine therapy, or within 12 months after the end of (neo)adjuvant endocrine therapy. Patients were allowed to receive ≤ 1 line of prior chemotherapy for advanced disease in cohort C. Both part 1 and part 2 patients who were ineligible for curative resection or radiotherapy must have had at least 1 measurable lesion according to the Response Evaluation Criteria in Solid Tumors (RECIST) v1.1 definition [16] and an Eastern Cooperative Oncology Group performance status of 0 or 1. Prior chemotherapy in the (neo)adjuvant setting was allowed.

For all monotherapy dose groups or combination cohorts, the key exclusion criteria were: the presence of brain or central nervous system metastases; previous treatment with CDK4 and CDK6 inhibitors; Impaired heart function or clinically significant heart disease; and clinically significant abnormalities of the gastrointestinal tract. The details of inclusion and exclusion criteria are presented in Additional file 2.

Study endpoints

The primary endpoints were safety, DLT, maximum tolerated dose (MTD) and RP2D-S of bireociclib monotherapy in patients with advanced solid tumors in dose-escalation and dose-expansion studies, and an RP2D-C of bireociclib combination regimen for HR⁺/HER2⁻ ABC patients.

The secondary endpoints were the objective response rate (ORR), progression-free survival (PFS), disease control rate (DCR), clinical benefit rate (CBR), duration of response (DoR) and overall survival (OS) in patients undergoing part 1 and part 2 studies.

DLT and safety assessments

In part 1 and part 2, the dose escalation continued until 2 or more patients reached DLT at one dose level during the DLT assessment period in the first cycle. DLT was defined as one of the following AEs related to study treatment: grade 4 hematological toxicity lasting for ≥ 5 consecutive days; grade ≥ 3 neutropenia with infection and fever (defined as a neutrophil count < 1.0 × 10⁹/L with fever (body temperature ≥ 38.5℃) or infection); grade ≥ 3 thrombocytopenia with bleeding; grade ≥ 3 vomiting, nausea, and electrolyte disturbances that continued for more than 2 days despite optimal management; grade ≥ 3 diarrhea despite optimal antidiarrhea treatment; confirmed grade ≥ 3 QTcF prolongation (QTcF > 500 ms) that persisted after correction of other potential causes such as electrolyte imbalances or hypoxia; any grade ≥ 3 non-hematologic AEs (including biochemical laboratory abnormalities, with renal dysfunction defined as creatinine clearance < 30 mL/min/1.73 m²), except for alopecia; and any additional levels of toxicity that required early termination of the trial after discussion by the investigator and sponsor.

Safety was monitored concerning AEs, laboratory parameters, vital signs, and the 12-lead electrocardiogram. AEs were graded by the investigator according to the National Cancer Institute Common Terminology Criteria for Adverse Events ver. 4.03 for part 1 and ver. 5.0 for part 2.

PK assessments

Part 1: Blood samples for bireociclib PK assessments were collected pre-dose on day 1 and post-dose at 1, 2, 4, 6, 8, 12, and 24 h on day 1, and on day 2 and day 3 during the single dose period. Patients entered the multiple-dose phase after completing the 72-h post-single-dose PK blood sampling on day 4. The first treatment cycle was calculated from the fourth day onwards, in patients receiving continuous dosing and blood sample collections at 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, and 24 h. This protocol fulfilled the PK blood collection requirements for both single and multiple doses.

Part 2: Blood samples for bireociclib PK assessments were collected before dosing on day 1 of cycle 1 and at 1, 2, 4, 6, 8, and 12 h post-dose on day 1 for single-dose PK characterization. Patients received continuous dosing and underwent PK blood sample collections at 1 h, 2 h, 4 h, 6 h, 8 h, and 12 h post-dose during the multiple-dose phase. This PK sampling strategy also met the requirements for both single- and multiple-dose assessments.

The main PK parameters measured were area under the curve from zero to the final quantifiable concentration, half-life (t_1/2), time to peak, maximum plasma concentration, apparent clearance, area under the curve between 0 and 12 h (AUC_0-12), volume of distribution for single-dose administration and minimum steady-state plasma concentration, maximum steady-state plasma concentration (C_ss,max), area under the curve at steady-state over a dosing interval, area under the curve at steady state from 0 to 12 h (AUC_ss,0–12), and the accumulation ratio for multi-dose administration. Bireociclib plasma concentrations in monotherapy or combination therapy patients were measured using liquid chromatography–mass spectrometry tandem mass spectrometry.

Efficacy assessments

CT or MRI scans were conducted at baseline and every other cycle during treatment. Tumor response assessments were performed according to RECIST v1.1.

Statistical analysis

The data collection was conducted using Medidata Classic Rave 2018.2.0 (EDC system, Medidata, NY, USA). The data cut-off date was June 30, 2022, for part 1 and May 19, 2023, for part 2. No formal hypotheses were tested in the present trial. Efficacy and safety were analyzed in the intention-to-treat population who had received at least one dose of the trial medication. ORR, PFS, DCR, CBR, DoR, and OS were evaluated. PK was analyzed in patients who received at least one dose and had evaluable post-treatment PK data. The median PFS, DoR, and OS were estimated using the Kaplan–Meier method. ORR, DCR, and CBR were estimated, with the corresponding 95% confidence intervals (CIs). All statistical analyses were carried out using SAS ver. 9.4 (SAS Institute Inc., Cary, NC).

Results

Baseline characteristics

Part 1, involving monotherapy with bireociclib, screened 225 patients across 15 hospitals in China, with 141 enrolled between April 27, 2018, and February 3, 2021. Part 2, which focused on combination therapy with bireociclib and endocrine therapy, screened 172 patients in 12 hospitals, with 130 enrolled between July 13, 2020, and August 8, 2021, and was comprised of three cohorts of patients: 35 in cohort A (bireociclib plus NSAI), 28 in cohort B (bireociclib plus FULV first-line) and 67 in cohort C (bireociclib plus FULV second-line) (Fig. 1). Patient baseline characteristics are summarized in Additional file1: Table S2. The median age of all patients given bireociclib monotherapy for advanced solid tumors was 53 years (range, 28–72) and for all bireociclib combination therapy patients confirmed with HR⁺/HER2⁻ ABC was 55 years (range, 26–70). In part 2, 70% of patients with ABC had received chemotherapy during neoadjuvant/adjuvant treatment. In part 2 cohort A, 3 patients had received prior systemic therapy in the advanced setting (2 with chemotherapy and 1 with endocrine therapy), representing protocol violations as this cohort required treatment-naïve patients in advanced disease.

Additionally, 209 patients (77.1%) exhibited visceral metastasis, with 119 given monotherapy with bireociclib and 90 patients given combination therapy with bireociclib. A total of 144 patients had ≥ 3 metastatic lesions.

At the data cut-off in part 1, 134 patients (95%) discontinued the treatment, with 7 still undergoing treatment, with the most common reasons being PD (71.6%) and AEs (9.7%). In part 2, 73 patients (56.2%) discontinued the treatment due to PD (78.1%) and AEs (11.0%).

MTD and RP2D

Eleven dose levels were explored from 20 mg QD to 480 mg BID for monotherapy (Fig. 1a) and 35 patients were included in the MTD analysis set. Three patients were excluded from the MTD analysis set because they received < 75% of the planned bireociclib dose, namely 2 patients in the 360 mg BID group and 1 patient in the 160 mg QD group. No DLTs were observed, and the MTD was not reached (NR) during dose escalation. Due to efficacy signals observed at higher dose levels during the dose escalation phase and toxicity concerns, coupled with PK considerations, further dose escalation was discontinued and dose expansion was conducted in higher dosage groups to evaluate further PK, safety, and efficacy across different dose levels. Considering the clinically acceptable safety profile at 480 mg BID, together with efficacy data and the moderate to high inter-individual PK variability (detailed data presented in subsequent sections of the article), the 480 mg BID regimen is expected to provide therapeutic benefit in a broader patient population. Therefore, 480 mg BID was selected as the RP2D-S.

As for combination therapy with FULV or NSAI (letrozole or anastrozole), 6 patients were enrolled and received bireociclib 360 mg BID and FULV or NSAI (letrozole or anastrozole) per standard dose in each cohort to evaluate DLTs. Among the 6 patients administered bireociclib plus FULV, no DLT was observed. However, among the 6 patients who received bireociclib plus letrozole or anastrozole, 1 patient experienced a DLT event of a grade 3 hepatic enzyme increase. Therefore, bireociclib 360 mg BID was determined as the RP2D-C, and an expansion study including cohorts A, B, and C was conducted to evaluate further the safety and efficacy of the combination regimens.

Safety

To ensure safety of monotherapy, the median exposure to bireociclib was 115 days (IQR, 54.0–282.0) and every patient had at least one treatment-emergent adverse event (TEAE). Specifically, 7.1% of serious AEs were related to bireociclib. Diarrhea was the most common TEAE, occurring in 83.7% of patients in any grade and 21.3% at grade 3. Additionally, neutropenia and leukopenia occurred in 77.3% of patients at any grade, with 23.4% and 14.9% at grade 3 or 4. The incidences of grade 3 elevated aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were 3.5% and 2.8%, respectively. In this population, grade 2 AEs with an incidence > 10% were leukopenia (46.8%), diarrhea (38.3%), neutropenia (38.3%), and anemia (25.5%) (Table 1). Additionally, in the 480 mg BID cohort, vomiting was reported in 8 patients (20%) and increased creatinine concentrations in 6 patients (15%). None of these grade 2 AEs occurred consecutively during the DLT observation period.

Table 1.

TEAEs that occurred in ≥ 20% of patients (part 1, dose escalation and expansion)—safety set

	Grade	QD		BID			Total
		20–420 mg	560 mg	240 mg	360 mg	480 mg
		N = 27	N = 43	N = 12	N = 19	N = 40	N = 141
Diarrhea	Any	18 (66.7)	37 (86.0)	10 (83.3)	15 (78.9)	38 (95.0)	118 (83.7)
	1	7 (25.9)	13 (30.2)	6 (50.0)	2 (10.5)	6 (15.0)	34 (24.1)
	2	9 (33.3)	16 (37.2)	2 (16.7)	11 (57.9)	16 (40.0)	54 (38.3)
	≥ 3	2 (7.4)	8 (18.6)	2 (16.7)	2 (10.5)	16 (40.0)	30 (21.3)
Leukopenia	Any	19 (70.4)	35 (81.4)	7 (58.3)	17 (89.5)	31 (77.5)	109 (77.3)
	1	8 (29.6)	5 (11.6)	2 (16.7)	5 (26.3)	2 (5.0)	22 (15.6)
	2	10 (37.0)	23 (53.5)	5 (41.7)	9 (47.4)	19 (47.5)	66 (46.8)
	≥ 3	1 (3.7)	7 (16.3)	0	3 (15.8)	10 (25.0)	21 (14.9)
Neutropenia	Any	17 (63.0)	37 (86.0)	8 (66.7)	16 (84.2)	31 (77.5)	109 (77.3)
	1	8 (29.6)	5 (11.6)	2 (16.7)	5 (26.3)	2 (5.0)	22 (15.6)
	2	5 (18.5)	23 (53.5)	5 (41.7)	6 (31.6)	15 (37.5)	54 (38.3)
	≥ 3	4 (14.8)	9 (20.9)	1 (8.3)	5 (26.3)	14 (35.0)	33 (23.4)
Vomiting	Any	10 (37.0)	31 (72.1)	4 (33.3)	7 (36.8)	31 (77.5)	83 (58.9)
	1	9 (33.3)	24 (55.8)	4 (33.3)	3 (15.8)	17 (42.5)	57 (40.4)
	2	0	3 (7.0)	0	3 (15.8)	8 (20.0)	14 (9.9)
	≥ 3	1 (3.7)	4 (9.3)	0	1 (5.3)	6 (15.0)	12 (8.5)
Anemia	Any	11 (40.7)	27 (62.8)	6 (50.0)	12 (63.2)	27 (67.5)	83 (58.9)
	1	6 (22.2)	11 (25.6)	3 (25.0)	7 (36.8)	11 (27.5)	38 (27.0)
	2	4 (14.8)	15 (34.9)	3 (25.0)	5 (26.3)	9 (22.5)	36 (25.5)
	≥ 3	1 (3.7)	1 (2.3)	0	0	7 (17.5)	9 (6.4)
Creatinine increased	Any	12 (44.4)	27 (62.8)	4 (33.3)	14 (73.7)	21 (52.5)	78 (55.3)
	1	11 (40.7)	23 (53.5)	3 (25.0)	11 (57.9)	15 (37.5)	63 (44.7)
	2	1 (3.7)	3 (7.0)	1 (8.3)	3 (15.8)	6 (15.0)	14 (9.9)
	≥ 3	0	1 (2.3)	0	0	0	1 (0.7)
Nausea	Any	10 (37.0)	22 (51.2)	3 (25.0)	7 (36.8)	24 (60.0)	66 (46.8)
	1	10 (37.0)	21 (48.8)	3 (25.0)	7 (36.8)	16 (40.0)	57 (40.4)
	2	0	1 (2.3)	0	0	8 (20.0)	9 (6.4)
	≥ 3	0	0	0	0	0	0
Hypertriglyceridemia	Any	6 (22.2)	25 (58.1)	4 (33.3)	10 (52.6)	14 (35.0)	59 (41.8)
	1	1 (3.7)	20 (46.5)	2 (16.7)	6 (31.6)	9 (22.5)	38 (27.0)
	2	4 (14.8)	3 (7.0)	2 (16.7)	2 (10.5)	3 (7.5)	14 (9.9)
	≥ 3	1 (3.7)	2 (4.7)	0	2 (10.5)	2 (5.0)	7 (5.0)
Thrombocytopenia	Any	4 (14.8)	21 (48.8)	3 (25.0)	6 (31.6)	21 (52.5)	55 (39.0)
	1	3 (11.1)	15 (34.9)	2 (16.7)	3 (15.8)	11 (27.5)	34 (24.1)
	2	0	4 (9.3)	0	2 (10.5)	4 (10.0)	10 (7.1)
	≥ 3	1 (3.7)	2 (4.7)	1 (8.3)	1 (5.3)	6 (15.0)	11 (7.8)
Elevated ALT	Any	10 (37.0)	17 (39.5)	5 (41.7)	6 (31.6)	16 (40.0)	54 (38.3)
	1	9 (33.3)	15 (34.9)	3 (25.0)	6 (31.6)	11 (27.5)	44 (31.2)
	2	1 (3.7)	2 (4.7)	1 (8.3)	0	2 (5.0)	6 (4.3)
	≥ 3	0	0	1 (8.3)	0	3 (7.5)	4 (2.8)
Elevated AST	Any	9 (33.3)	15 (34.9)	4 (33.3)	9 (47.4)	15 (37.5)	52 (36.9)
	1	9 (33.3)	13 (30.2)	2 (16.7)	8 (42.1)	9 (22.5)	41 (29.1)
	2	0	2 (4.7)	1 (8.3)	1 (5.3)	2 (5.0)	6 (4.3)
	≥ 3	0	0	1 (8.3)	0	4 (10.0)	5 (3.5)
Hypokalemia	Any	2 (7.4)	11 (25.6)	3 (25.0)	8 (42.1)	19 (47.5)	43 (30.5)
	1	2 (7.4)	11 (25.6)	3 (25.0)	7 (36.8)	12 (30.0)	35 (24.8)
	2	0	0	0	0	2 (5.0)	2 (1.4)
	≥ 3	0	0	0	1 (5.3)	5 (12.5)	6 (4.3)
Abdominal pain	Any	8 (29.6)	15 (34.9)	0	3 (15.8)	13 (32.5)	39 (27.7)
	1	8 (29.6)	14 (32.6)	0	2 (10.5)	10 (25.0)	34 (24.1)
	2	0	1 (2.3)	0	1 (5.3)	3 (7.5)	5 (3.5)
	≥ 3	0	0	0	0	0	0
Hypercholesteremia	Any	2 (7.4)	16 (37.2)	2 (16.7)	6 (31.6)	12 (30.0)	38 (27.0)
	1	1 (3.7)	15 (34.9)	1 (8.3)	5 (26.3)	11 (27.5)	33 (23.4)
	2	1 (3.7)	1 (2.3)	1 (8.3)	0	1 (2.5)	4 (2.8)
	≥ 3	0	0	0	1 (5.3)	0	1 (0.7)
Hypoalbuminemia	Any	1 (3.7)	10 (23.3)	4 (33.3)	7 (36.8)	13 (32.5)	35 (24.8)
	1	1 (3.7)	9 (20.9)	4 (33.3)	7 (36.8)	12 (30.0)	33 (23.4)
	2	0	1 (2.3)	0	0	1 (2.5)	2 (1.4)
	≥ 3	0	0	0	0	0	0
Urinary tract infection	Any	3 (11.1)	9 (20.9)	2 (16.7)	7 (36.8)	14 (35.0)	35 (24.8)
	1	0	5 (11.6)	2 (16.7)	5 (26.3)	11 (27.5)	23 (16.3)
	2	3 (11.1)	4 (9.3)	0	2 (10.5)	3 (7.5)	12 (8.5)
	≥ 3	0	0	0	0	0	0
Weight decrease	Any	4 (14.8)	7 (16.3)	1 (8.3)	7 (36.8)	13 (32.5)	32 (22.7)
	1	0	6 (14.0)	1 (8.3)	5 (26.3)	10 (25.0)	22 (15.6)
	2	3 (11.1)	1 (2.3)	0	1 (5.3)	3 (7.5)	8 (5.7)
	≥ 3	1 (3.7)	0	0	1 (5.3)	0	2 (1.4)
Hyperuricemia	Any	6 (22.2)	13 (30.2)	1 (8.3)	3 (15.8)	8 (20.0)	31 (22.0)
	1	6 (22.2)	13 (30.2)	1 (8.3)	3 (15.8)	8 (20.0)	31 (22.0)
	2	0	0	0	0	0	0
	≥ 3	0	0	0	0	0	0
Epigastric pain	Any	3 (11.1)	9 (20.9)	2 (16.7)	3 (15.8)	12 (30.0)	29 (20.6)
	1	3 (11.1)	8 (18.6)	2 (16.7)	3 (15.8)	9 (22.5)	25 (17.7)
	2	0	1 (2.3)	0	0	3 (7.5)	4 (2.8)
	≥ 3	0	0	0	0	0	0

Data are presented as number of patients (incidence). ALT alanine aminotransferase; AST aspartate aminotransferase; BID twice-daily; QD once-daily; TEAEs treatment-emergent adverse events

No deaths occurred in patients during the trial treatment period. However, 3 patients died within 30 days of the end of treatment due to AEs, all of which were deemed unrelated to bireociclib.

For the safety of combination therapy, the median exposure to bireociclib was 479.3 days (IQR, 195.0–728.0). Notably, similar to those in part 1, the TEAEs that occurred in part 2 mostly involved gastrointestinal and hematopoietic toxicities (Table 2). The most common all-grade AEs in the bireociclib plus NSAI group were leukopenia (34, 97.1%), diarrhea (33, 94.3%), and neutropenia (30, 85.7%). In addition, the most common grade 3 or 4 TEAEs were neutropenia (13, 37.1%), leukopenia (8, 22.9%), anemia (5, 14.3%), and diarrhea (4, 11.4%). While the most frequent all-grade and grade 3 or 4 TEAEs in the bireociclib plus FULV group were diarrhea (91, 95.8%; 18, 18.9%), leukopenia (90, 94.7%; 23, 24.2%) and neutropenia (88, 92.6%; 32, 33.7%). Serious TEAEs were reported in 15 (42.9%) and 14 (14.7%) patients in the bireociclib plus NSAI and bireociclib plus FULV groups, with the most common types being pneumonia (14.3%) and thrombocytopenia (5.3%), respectively. Elevated AST and ALT occurred in 25 (71.4%) patients in the bireociclib plus NSAI group with no grade ≥ 3 elevations. In the bireociclib plus FULV group, AST and ALT elevations occurred in 58 (61.1%) and 53 (55.8%) patients, respectively, with grade 3 or 4 increases also observed (6.4% vs. 5.3%). In both the NSAI and FULV combination groups, the most common grade 2 TEAEs that occurred in > 10% of patients were primarily hematologic, gastrointestinal, and metabolic toxicities, including leukopenia, diarrhea, neutropenia, anemia, vomiting, and hypertriglyceridemia (Table 2). Additionally, of the 5 patients (3.85%) who experienced venous thromboembolism, 1 permanently discontinued treatment, 1 required dose reduction, and 3 experienced temporary treatment interruption. No interstitial pneumonia was observed. No deaths occurred in patients during the trial treatment, though 4 deaths (3.1%) occurred within 30 days of a patient’s last dose, 2 (1.5%) due to PD. Two (1.5%) were deemed to be possibly associated with the study treatment by the investigator. One death occurred 26 days after discontinuation of bireociclib plus anastrozole due to PD; the patient died suddenly at home, with the cause of death remaining unknown. Another patient, who received bireociclib in combination with FULV, died peacefully in her sleep at home during cycle 10, with no ongoing AEs reported prior to her death. In both cases, the sponsor considered the deaths unlikely to be related to the investigational drugs.

Table 2.

TEAEs elicited by bireociclib combination therapy (part 2), with TEAEs occurring in ≥ 20% of all treated patients – safety set

	Bireociclib plus NSAI (N = 35)				Bireociclib plus FULV (N = 95)
	All grades	Grade 2	Grade 3	Grade 4	All grades	Grade 2	Grade 3	Grade 4
All	35 (100.0)	34 (97.1)	26 (74.3)	4 (11.4)	95 (100)	94 (98.9)	65 (68.4)	12 (12.6)
Leukopenia	34 (97.1)	17 (48.6)	8 (22.9)	0	90 (94.7)	57 (60.0)	23 (24.2)	0
Diarrhea	33 (94.3)	17 (48.6)	4 (11.4)	0	91 (95.8)	45 (47.4)	18 (18.9)	0
Neutropenia	30 (85.7)	12 (34.3)	13 (37.1)	0	88 (92.6)	49 (51.6)	27 (28.4)	5 (5.3)
Anemia	21 (60.0)	8 (22.9)	5 (14.3)	0	72 (75.8)	28 (29.5)	13 (13.7)	0
Hypertriglyceridemia	25 (71.4)	10 (28.6)	1 (2.9)	1 (2.9)	60 (63.2)	13 (13.7)	9 (9.5)	0
Elevated AST	25 (71.4)	1 (2.9)	0	0	58 (61.1)	12 (12.6)	5 (5.3)	1 (1.1)
Blood creatinine increased	23 (65.7)	8 (22.9)	0	0	56 (58.9)	9 (9.5)	0	0
Thrombocytopenia	20 (57.1)	7 (20.0)	2 (5.7)	0	58 (61.1)	9 (9.5)	8 (8.4)	4 (4.2)
Elevated ALT	25 (71.4)	4 (11.4)	0	0	53 (55.8)	5 (5.3)	4 (4.2)	1 (1.1)
Hypercholesterolemia	19 (54.3)	2 (5.7)	1 (2.9)	0	52 (54.7)	3 (3.2)	0	1 (1.1)
Hyperuricemia	20 (57.1)	0	0	0	48 (50.5)	1 (1.1)	0	0
Nausea	15 (42.9)	2 (5.7)	0	0	45 (47.4)	10 (10.5)	0	0
Hypokalaemia	19 (54.3)	0	4 (11.4)	1 (2.9)	36 (37.9)	8 (8.4)	7 (7.4)	1 (1.1)
Vomiting	15 (42.9)	4 (11.4)	0	0	38 (40.0)	16 (16.8)	1 (1.1)	0
Weight decrease	15 (42.9)	1 (2.9)	1 (2.9)	0	35 (36.8)	9 (9.5)	0	0
Urinary tract infection	10 (28.6)	7 (20.0)	0	0	34 (35.8)	21 (22.1)	2 (2.1)	0
Blood lactate dehydrogenase increased	15 (42.9)	0	0	0	29 (30.5)	0 (0)	0	0
Hypoalbuminemia	12 (34.3)	0	0	0	30 (31.6)	3 (3.2)	0	0
Fatigue	10 (28.6)	0	0	0	26 (27.4)	2 (2.1)	2 (2.1)	0
Hyperglycaemia	11 (31.4)	0	0	0	23 (24.2)	2 (2.1)	0	0
Gamma-glutamyltransferase increase	11 (31.4)	2 (5.7)	0	1 (2.9)	21 (22.1)	4 (4.2)	3 (3.2)	1 (1.1)
Hypophosphatemia	5 (14.3)	0	0	0	26 (27.4)	1 (1.1)	0	0
Hyponatremia	8 (22.9)	0	0	0	18 (18.9)	1 (1.1)	0	0

Data are presented as number of patients (incidence)ALT alanine aminotransferase; AST aspartate aminotransferase; FULV fulvestrant; NSAI non-steroidal aromatase inhibitor; TEAEs treatment-emergent adverse events

Pharmacokinetics

PK analysis of bireociclib monotherapy was based on blood samples obtained from all 85 patients. Samples were collected on cycle 1 day 1 (C1D1) for 20–560 mg QD and 240–480 mg BID following single-dose administration, and on C2D1 within 24 h after multiple-dose administration. These samples were used to evaluate the PK parameters of bireociclib based on the plasma concentration–time profiles for a single dose and multiple doses, as shown in Additional file 1: Fig. S1 and Fig. 2. The geometric mean t_1/2 was 12.3–27.0 h, and the exposure level in the plasma (AUC_0-12) was 351.5–3024.7 h × ng/mL following single-dose administration of bireociclib from 20–560 mg QD to 240–480 mg BID. When bireociclib was administered as QD in multiple doses ranging from 80 to 560 mg, the median value of the time to reach the maximum steady-state plasma concentration (T_ss,max) was 4–6 h. Additionally, when bireociclib was given at 240–480 mg as BID by multi-dose administration, the C_ss,max was achieved 276.0 (46.2)–403.6 (59.8) ng/mL in a median time (T_ss,max) of 4.0–5.0 h, and AUC_ss,0–12 was 2600.2–4019.1 h × ng/mL. The exposure was increased by raising the dose from 240 to 480 mg BID for multiple doses. The plasma concentration fluctuations in the bireociclib 480 mg BID (0.506) were the lowest among the doses examined, suggesting that the BID dosing regimen produced a more stable plasma concentration (Table 3, Additional file1: Table S3). Comparable systemic exposures of the parent drug and its metabolites were observed when bireociclib was administered at 360 mg BID in combination with either NSAIs or FULV, relative to the same dosing regimen administered as monotherapy in the present study (Additional file 1: Table S4).

Fig. 2.

Mean plasma concentration-time of bireociclib (part 1) BID administration – pharmacokinetic parameter set. a) single-dose BID oral administration; b) multiple-dose BID oral administration. Data are presented as the mean ± SD. BID twice-daily.

Table 3.

PK parameters of the parent drug bireociclib after multiple doses (geometric mean)—PK parameter set

	QD						BID
	80 mg (N = 2)	160 mg (N = 3)†	240 mg (N = 6)	320 mg (N = 4)	420 mg (N = 10)	560 mg (N = 20)	240 mg (N = 8)	360 mg (N = 11)	480 mg (N = 19)
Css,max (ng/mL)	110.8 (21.6)	323.9 (170.7)	273.9 (58.0)	435.0 (75.1)	326.6 (58.1)	394.1 (30.4)	276.0 (46.2)	282.2 (51.5)	403.6 (59.8)
Css,min (ng/mL)	24.3 (115.7)	167.0 (288.1)	65.2 (98.5)	165.7 (66.4)	107.2 (58.3)	139.3 (42.5)	142.8 (71.4)	156.9 (66.3)	257.5 (79.4)
AUCss,0-t (h × ng/mL)	1463.6 (50.2)	5868.3 (243.3)	3758.7 (68.6)	6841.5 (70.7)	4825.9 (57.8)	6321.7 (29.5)	4228.7 (55.7)	4243.5 (59.3)	7194.3 (72.4)
AUCss,0–12 (h × ng/mL)	961.5 (42.1)	3137.5 (184.2)	2368.3 (59.4)	3993.5 (72.7)	2977.4 (59.7)	3704.7 (29.6)	2600.2 (51.8)	2720.2 (55.2)	4019.1 (65.7)
AUCss, 0-tau (h × ng/mL)	1466.8 (51.0)	5869.4 (237.0)	3785.8 (68.8)	6861.9 (70.9)	4818.3 (62.8)	6225.7 (32.8)	2600.2 (51.8)	2720.2 (55.2)	4019.1 (65.7)
Tss, max (h)*	5.0 (4.0–6.0)	5.0 (4.1–6.0)	4.0 (4.0–6.0)	6.0 (4.0–6.1)	6.0 (4.0–7.9)	6.0 (3.9–8.1)	5.0 (3.7–8.0)	4.0 (2.0–6.0)	4.0 (0.0–8.0)
Swing	3.395 (116.2)	0.884 (78.4)	3.031 (91.6)	1.611 (24.8)	2.017 (31.9)	1.752 (54.1)	0.857 (67.7)	0.753 (51.3)	0.506 (70.9)

Data are presented as the geometric mean (geometric coefficient of variation). QD group: tau = 24 h; BID: tau = 12 h.*T_{ss, max} is presented as the median (min–max).^†The PK parameter values of only one patient in the dose group are shown. AUC_ss,0-t area under the curve from zero to the final quantifiable steady state concentration; AUC_ss,0-12 area under the curve at steady state between 0 and 12 h; AUC_ss,0-tau area under the curve at steady state over a dosing interval; BID twice-daily; C_ss,max maximum steady-state plasma concentration; C_ss,min minimum steady-state plasma concentration; PK pharmacokinetic; QD once-daily; T_ss,max time to reach the maximum steady-state plasma concentration; Swing (peak-to-trough fluctuation), quantifies the fluctuations in the extent of plasma drug concentration variability following multiple doses of bireociclib, Swing = (C_ss,maxC_ss,min)/C_ss,min.

Efficacy

For the efficacy of treatments in monotherapy, in the bireociclib QD administration study, no patient had a best overall response (BOR) of complete response (CR). Seven patients achieved partial responses (PRs), including 1 in the 80 mg cohort, 6 in the 560 mg cohort, and 35 had a BOR of stable disease (SD) (Additional file 1: Table S5). The 18-month PFS rate was observed in the 420 mg (12.7%) and 560 mg (12.6%) cohorts. The 18-month OS rate was 100% in both the 240 mg and 360 mg groups, 87.5% in the 420 mg group, and 62.8% in the 560 mg group.

Among patients who received bireociclib BID, 9 achieved PR and 37 had SD. The highest ORR was 17.5% (95% CI, 7.34–32.78) at 480 mg, with a median DoR of 7.4 months (95% CI, 5.22–not evaluable [NE]) among responders (Additional file 1: Table S5). The best percentage changes in target lesion size for patients who received bireociclib QD or BID are shown in Fig. 3.

Fig. 3.

Waterfall plot of best percentage change in the sum of diameters after baseline in part 1 – intention-to-treat population. a 560 mg QD dose, (b) 480 mg BID dose. Only patients with measurable lesions are shown in Fig. 3; in (a) 7 patients with a BOR of NE and 2 with PD are not shown; in (b) the corresponding numbers were 6 and 1, respectively. BOR best overall response; NE (N) not evaluable; PD (P) progressive disease; PR (R) partial response; SD (S) stable disease; SLD sum of all (target) lesion diameters

Therefore, 480 mg BID was established as the RP2D-S for monotherapy of bireociclib in patients with advanced solid tumors, based on PK, safety, and efficacy data confirmed by the sponsor and principal investigators.

For the efficacy of bireociclib combination therapy, in patients who had no prior systemic therapy for advanced disease (intention-to-treat population), the median follow-up times were 23.4 and 24.0 months in cohort A and cohort B, respectively. At the cut-off, the ORR was 57.1% (95% CI: 39.35–73.68) in cohort A and 57.1% (95% CI: 37.18–75.54) in cohort B (Table 4). This included 1 CR in cohort A and 35 PRs (19 in cohort A and 16 in cohort B) (Fig. 4a, b, d, and e). In this population, 21 PFS events occurred (9 [25.7%] in cohort A and 12 [42.9%] in cohort B). The median PFS was NR in cohort A and 26.3 months (95% CI, 9.99–NR) in cohort B. The 18-month PFS rate was 75.8% in cohort A and 60.2% in cohort B. The median DoR was NR in cohort A with 13 patients (37.1%) still receiving treatment at the time of analysis and 24.5 months (95% CI, 9.26–NR) in cohort B. The CBR was 71.4% (95% CI, 53.70–85.36) in cohort A and 78.6% (95% CI, 59.05–91.70) in cohort B (Table 4). At the time of data cut-off, 3 and 5 patients died in cohorts A and B, respectively. The median OS was maturing in cohort A and was 28.3 months (95% CI, 28.29–NR) in cohort B.

Table 4.

Responses according to RECIST v1.1 produced by bireociclib combination therapy (part 2) – intention-to-treat population

	Cohort A* (n = 35)	Cohort B* (n = 28)	Cohort C* (n = 67)
Best overall response, n (%)
CR	1 (2.9)	0	2 (3.0)
PR	19 (54.3)	16 (57.1)	29 (43.3)
SD	12 (34.3)	9 (32.1)	29 (43.3)
PD	0	3 (10.7)	7 (10.4)
Not evaluable	3 (8.6)	0	0
ORR (%, 95% CI)	57.1 (39.35–73.68)	57.1 (37.18–75.54)	46.3 (34.00–58.88)
DCR (%, 95% CI)	91.4 (76.94–98.20)	89.3 (71.77–97.73)	89.6 (79.65–95.70)
CBR (%, 95% CI)	71.4 (53.70–85.36)	78.6 (59.05–91.70)	71.6 (59.31–81.99)
Survival estimates
Median PFS (months, 95% CI)	NR (20.83–NR)	26.3 (9.99–NR)	20.1 (8.90–27.14)
Median DoR (months, 95% CI)	NR (NR–NR)	24.5 (9.26–NR)	21.8 (11.07–25.53)

*Cohort A: bireociclib plus letrozole/anastrozole group; Cohort B: bireociclib plus FULV first-line; Cohort C: bireociclib plus FULV second-line. Cohorts A and B included patients who had not received prior systemic treatment in the metastatic setting. Among these patients, de novo metastatic cases represented only a minority, as the majority had previously received adjuvant therapy. CBR clinical benefit rate; CI confidence interval; CR complete response; DCR disease control rate; DoR duration of response; FULV fulvestrant; NR not reached; ORR objective response rate; PD progressive disease; PFS progression-free survival; PR partial response; RECIST Response Evaluation Criteria in Solid Tumors; SD stable disease

Fig. 4.

Tumor response for bireociclib combination therapy in part 2 – efficacy analysis set. a, b and c. Waterfall plot of maximal percentage change in tumor size for the cohort of patients receiving 360 mg bireociclib plus NSAI (cohort A) (a), 360 mg bireociclib plus FULV for treatment naïve patients in metastatic setting (cohort B) (b), and 360 mg bireociclib plus FULV for patients who progressed on or after prior endocrine therapy (cohort C) (c). d, e and f. Duration of treatment for patients in cohort A (d), cohort B (e) and cohort C (f). BOR best overall response; CR complete response; FULV fulvestrant 500 mg intramuscularly (on day 1 and day 15 of the first cycle and then on day 1 of each subsequent 4-week cycle); NE not evaluable; NSAI non-steroidal aromatase inhibitor (letrozole 2.5 mg or anastrozole 1 mg oral QD); PD progressive disease; PR partial response; SD stable disease

Among the patients who progressed on or after prior endocrine therapy and continued FULV with the addition of bireociclib (cohort C), the median follow-up time was 24.0 months. At the cut-off, the BOR for 2 patients (3.0%) was CR and 29 (43.3%) achieved a PR (Fig. 4c and f). The ORR was 46.3% (95% CI, 34.00–58.88) and the CBR was 71.6% (95% CI, 59.31–81.99) in cohort C. The median DoR was 21.8 months (95% CI, 11.07–25.53). In cohort C, the median PFS reached 20.1 months (95% CI, 8.90–27.14) (Table 4).

In exploratory subgroup analyses of patients with visceral metastases, the ORRs for cohort A and cohort B were 52.2% (95% CI, 30.59%–73.18%) and 52.9% (95% CI, 27.81%–77.02%), respectively. On the other hand, the ORR was 40.0% (95% CI, 26.41%–54.82%) in cohort C for patients with visceral metastases who had failed to respond to prior ET (Table 5). For patients with visceral metastases, liver involvement was observed in 2 patients (5.7%) in cohort A and 5 (17.9%) in cohort B. In cohort C, liver metastases were present in 26 patients (38.8%).

Table 5.

Subgroup analyses of patients with visceral metastases and after previous endocrine therapy: intention-to-treat population

	Visceral metastases (n = 90)			Endocrine resistance (Cohort C, n = 67)
	Cohort A* (n = 23)	Cohort B* (n = 17)	Cohort C* (n = 50)	Primary endocrine resistance (n = 19)	Secondary endocrine resistance(n = 48)
Best of response, n (%)
CR	1 (4.3)	0	0	2 (10.5)	0
PR	11 (47.8)	9 (52.9)	20 (40.0)	9 (47.4)	20 (41.7)
SD	10 (43.5)	7 (41.2)	24 (48.0)	5 (26.3)	24 (50.0)
PD	0	1 (5.9)	6 (12.0)	3 (15.8)	4 (8.3)
Not evaluable	1 (4.3)	0	0	0	0
ORR (%, 95% CI)	52.2 (30.59–73.18)	52.9 (27.81–77.02)	40.0 (26.41–54.82)	57.9 (33.50–79.75)	41.7 (27.61–56.79)
DCR (%, 95% CI)	95.7 (78.05–99.89)	94.1 (71.31–99.85)	88.0 (75.69–95.47)	84.2 (60.42–96.62)	91.7 (80.02–97.68)
CBR (%, 95% CI)	69.6 (47.08–86.79)	82.4 (56.57–96.20)	64.0 (49.19–77.08)	57.9 (33.50–79.75)	77.1 (62.69–87.97)
Survival estimates
Median PFS (months, 95% CI)	NR (9.20–NR)	26.3 (7.43–NR)	12.7 (7.03–27.14)	8.9 (3.65–24.02)	25.7 (11.93–NR)

*Cohort A: bireociclib plus letrozole/anastrozole group; Cohort B: bireociclib plus FULV first-line; Cohort C: bireociclib plus FULV second-line. Cohorts A & B for treatment naïve patients in the metastatic setting; Cohort C for patients who progressed on or after prior endocrine therapy. CBR clinical benefit rate; CI confidence interval; CR, complete response; DCR, disease control rate; FULV fulvestrant; NR not reached; ORR, objective response rate; PD, progressive disease; PFS progression-free survival; PR, partial response; SD, stable disease.

In the subgroup of primary endocrine resistance in cohort C, the ORR was 57.9% (95% CI, 33.50–79.75), compared to the secondary endocrine resistance 41.7% (95% CI, 27.61–56.79). Additionally, in the subgroup of primary endocrine resistance in cohort C, the DCR was 84.2% (95% CI, 60.42–96.62), compared to secondary endocrine resistance of 91.7% (95% CI, 80.02–97.68). The median PFS was 8.9 months (95% CI, 3.65–24.02) for patients with primary endocrine resistance and 25.7 months (11.93-NR) for those with secondary endocrine resistance in cohort C (Table 5).

Discussion

This phase 1 clinical trial provides a preliminary evaluation of the safety and efficacy of bireociclib, a novel selective CDK4/6 inhibitor, administered continuously in patients with advanced solid tumors, particularly those with HR⁺/HER2⁻ ABC. The trial explored the appropriate dosage of bireociclib for use in patients with advanced solid tumors and revealed that 480 mg BID and 360 mg BID could be recommended as safe, suitable doses for monotherapy and in combination with endocrine therapy, respectively, for advanced HR⁺/HER2⁻ ABC patients. Furthermore, the results provided good evidence for the efficacy of bireociclib in combination with endocrine therapy for patients with HR⁺/HER2⁻ ABC.

In the QD dosing regimen for bireociclib monotherapy, doses ranging from 20 to 560 mg QD generally showed increased exposure with increasing dose levels. However, for certain dose levels, such as 420 mg QD vs. 320 mg QD, higher doses yielded lower drug exposure than lower doses. This inconsistency may be attributed to significant inter-patient variability influenced by disease factors, dose increments that were too small to overcome this variability, and small, uneven sample sizes across the dosing groups. PK data from bireociclib monotherapy at doses ranging from 240 to 480 mg BID revealed increases in drug exposure. The C_ss,max ranged from 276.0 (46.2) to 403.6 (59.8) h × ng/mL, occurring approximately 4.0–5.0 h post-dose. The AUC _ss,0–12 ranged from 2600.2 (51.8) to 4019.1 (65.7) h × ng/mL, with a t_1/2 of 12.3–12.7 h. Swing ranged from 0.51 to 0.86. These findings support the feasibility of the BID dosing regimen to maintain a stable plasma concentration. No dedicated drug-drug interaction (DDI) study between bireociclib and endocrine therapy has been conducted, as previous findings with other CDK4/6 inhibitors [17, 18] showed minimal risk of clinically relevant DDIs when combined with endocrine therapy. Moreover, PK comparisons were performed to evaluate potential exposure differences between monotherapy (part 1) and combination therapy (part 2) for bireociclib. The PK data in the present study revealed that after a single dose or multiple doses of bireociclib tablets at 360 mg BID in combination with NSAIs or FULV, the exposure levels of both the parent drug and its metabolites were comparable between combination therapy and monotherapy (Additional file 1: Table S4).

No DLT events were observed during the dose-escalation phase of bireociclib monotherapy and combined with PK properties and preliminary efficacy, 480 mg BID was selected as PR2D-S. In the dose-escalation phase of combination therapy with endocrine treatment, 1 patient experienced a DLT of a grade 3 hepatic enzyme increase at 360 mg BID during the first treatment cycle, setting this as PR2D-C. This dosing strategy, namely the combination therapy dose being one level lower than the monotherapy dose, aligns with the precedent set by abemaciclib, which shares a benzimidazole scaffold with bireociclib. In a phase 1 dose-escalation study of abemaciclib, 4 patients exhibited grade 3 fatigue as DLTs, leading to the establishment of 200 mg BID as the MTD and RP2D-S. Subsequently, a lower dose of 150 mg BID was evaluated and adopted as the recommended dose for combination with endocrine therapy [19]. More detailed information on bireociclib and abemaciclib regarding their chemical structures, in vitro activity against CDKs, dosing regimens, PK, and safety profiles in humans can be found in Additional file 1: Table S6 [19].

In the present trial, bireociclib produced predominantly hematologic and gastrointestinal toxicity (with common grade 2 AEs also in these categories), which were generally well-managed following appropriate symptomatic treatment and similar to other CDK4/6 inhibitors. During the dose escalation and expansion phases of bireociclib monotherapy, the incidence of grades 3–4 neutropenia was 23.4%. In previously reported phase 1 studies of other CDK4/6 inhibitors, including palbociclib [20], ribociclib [21], dalpiciclib [22] and abemaciclib [23], the reported incidences were 33.0%, 27.0%, 52.5%, and 32.0%, respectively. These differences should be interpreted with caution, as cross-trial comparisons are limited by differences in the study designs and patient populations. An unexpected finding was that the incidence of TEAEs was higher in the 420 mg QD group than in the 560 mg QD group. This discrepancy may be related to differences in patient baseline characteristics, sample size variations, or variation in dose-dependent tolerability. Some AEs occurred at a higher incidence at moderate doses, which could be attributed to nonlinear PK characteristics, adaptive physiological responses in the body, or inter-individual variability to drug exposure [24–26]. Given these observations and the limited sample sizes in each cohort, further evaluations in larger phase 2 or phase 3 studies are warranted to characterize more comprehensively the safety profile of bireociclib.

When combined with NSAI or FULV, bireociclib was associated with grade 3 or 4 neutropenia rates of 37.1% and 33.7%, respectively. The rate observed with NSAI (37.1%) appeared numerically lower than those reported for palbociclib plus letrozole in Asian (grade 3, 67.7%) [27], dalpiciclib plus letrozole or anastrozole (grade 3, 65.0%) [28], ribociclib plus letrozole in postmenopausal Chinese (grade ≥ 3, 63.6%) [29], ribociclib plus NSAI in premenopausal Chinese (grade ≥ 3, 62.0%) [29]. Similarly, the rate observed with FULV (33.7%) was numerically lower than that reported for palbociclib plus FULV in Asian (grade 3, 74.0%) [30], dalpiciclib plus FULV (grade 3, 65.0%) [31], and abemaciclib plus FULV than in East Asian populations (grade ≥ 3, 48.6%) [32], although cross-trial comparisons should be interpreted with caution. Despite thrombotic events that occurred in 5 patients in this study, all required dose modifications. The subsequent larger phase 3 trial with FULV combination showed minimal impact (only one discontinuation and virtually no dose reductions/interruptions due to thromboembolism), suggesting these events may be disease-related rather than representing a significant drug safety concern.

Previous studies have shown that Asians are more susceptible to myelosuppression, influenced by their racial background and genetic differences [33]. Our results suggest that bireociclib represents one of the potentially favorable treatment options for patients with myelopoietic insufficiency. Additionally, diarrhea was a common TEAE associated with bireociclib. However, no dose reductions due to diarrhea occurred during part 1 monotherapy, and only 1 patient in the part 2 combination therapy group required a dose reduction for grade 2 diarrhea. The incidence of grade 3 diarrhea (no grade 4 events observed) during bireociclib monotherapy dose escalation and expansion phases was 21.3%, while it was 11.4% and 18.9% when combined with NSAI and FULV, respectively. The most common grade 3 or 4 TEAEs in the bireociclib plus NSAI or FULV groups were also diarrhea, similar to the safety profile of abemaciclib [34–36], possibly due to the structural similarity between bireociclib and abemaciclib, which may have a certain inhibitory effect on CDK9 (IC₅₀, 53 nM). Hematologic abnormalities and gastrointestinal symptoms were adequately managed with standard supportive care, did not lead to complications, and resolved without cumulative effects. Increased creatinine concentrations were also common in the present study but primarily were grade 1. Previous studies with abemaciclib suggest that a modest elevation in the serum creatinine concentration may result from inhibition of OCT2, MATE1, and MATE2-K transporters [37]. Similarly, preclinical studies have demonstrated that bireociclib also inhibits OCT2, suggesting a consistent mechanism for the observed creatinine elevation. Notably, AEs related to cystatin-C, an alternative renal function marker, were infrequent among these patients, indicating that bireociclib likely affects only creatinine transport mechanisms without causing substantive renal impairment. This mechanism parallels that observed with abemaciclib. Comparing these four CDK4/6 inhibitors, Importantly, our current bireociclib phase 1 trial did not reveal any new AEs.

Our findings demonstrated, unequivocally, that bireociclib combined with endocrine therapy had significant clinical benefits for postmenopausal and premenopausal or perimenopausal women with HR⁺/HER2⁻ ABC. In patients who were treatment naïve in the advanced setting, the ORR reached 57.1% both in the bireociclib plus NSAI or FULV group, and the median PFS was NR for patients receiving bireociclib plus NSAI, being 26.3 months for those in the bireociclib plus FULV group. Other clinical trials have reported the efficacy of CDK4/6 inhibitors combined with NSAI as first-line treatment for patients with HR⁺/HER2⁻ ABC, with median PFS ranging from 20.2 to 28.2 months and ORR values between 54.5% and 62.5% [27, 38–41]. For patients who had PD on or after prior endocrine therapy for advanced disease, the median PFS was 20.1 months in the bireociclib plus FULV group. In previous clinical trials, a CDK4/6 inhibitor combination with FULV produced median PFS times ranging from 11.2 to 16.6 months [7, 31, 35]. These findings suggest that the combination of bireociclib and NSAI or FULV may have the potential to provide significant clinical benefits in patients who were treatment naïve in the advanced setting or have progressed on or after prior endocrine therapy, though further validation in larger, randomized trials is needed to confirm its comparative efficacy. Considering the patient baseline characteristics, 76.2% aged 41–64 years, 69.2% with visceral disease and 25.4% having received prior chemotherapy for advanced settings, the observed activity of bireociclib in combination with NSAI or FULV indicates potential use in younger patients with more aggressive disease features. While OS data are still immature, preliminary evidence of sustained benefits beyond the treatment period supports further investigation of bireociclib as a treatment option for patients with HR⁺/HER2⁻ ABC.

Conclusions

Taken together, the findings of the present trial have demonstrated that bireociclib has an acceptable safety profile and showed preliminary signs of clinical activity in combination with NSAI (letrozole/anastrozole) or FULV, findings that warrant further phase 2 and phase 3 clinical trials, especially for patients with HR⁺/HER2⁻ ABC.

Abbreviations

ABC

Advanced breast cancer

AE

Adverse event

ALT

Alanine aminotransferase

AST

Aspartate aminotransferase

AUC_0-12

Area under the curve between 0 and 12 h

AUC_ss,0-12

Area under the curve at steady-state from 0 to 12 h

BID

Twice-daily

BOR

Best overall response

C1D1

Cycle 1 day 1

CBR

Clinical benefit rate

CDK/Rb

Cyclin-dependent kinase/retinoblastoma-protein

CDK4/6

Cyclin-dependent kinase 4 and 6

CIs

Confidence intervals

CR

Complete response

C_ss,max

Maximum steady-state plasma concentration

DCR

Disease control rate

DLT

Dose-limiting toxicity

DoR

Duration of response

ER⁺

Estrogen receptor-positive

FULV

Fulvestrant

HER2

Human epidermal growth factor receptor-2 negative

HR⁺

Hormone receptor-positive

IC₅₀

Half maximal inhibitory concentration

MTD

Maximum tolerated dose

NE

Not evaluable

NR

Not reached

NSAI

Non-steroidal aromatase inhibitor

ORR

Objective response rate

OS

Overall survival

PD

Disease progression

PR

Partial response

PFS

Progression-free survival

PK/PD

Pharmacokinetic and pharmacodynamic

PR

Partial response

QD

Once-daily

RECIST

Response Evaluation Criteria in Solid Tumors

RP2D-C

Recommended phase 2 dosage for combination therapy

RP2D-S

Recommended phase 2 dosage for a single regimen

SD

Stable disease

t_1/2

Half-life

TEAE

Treatment-emergent adverse event

T_ss,max

Time to reach the maximum steady-state plasma concentration

Authors’ contributions

BHX had full access to all the data in the study and took responsibility for its integrity and the accuracy of data analysis. JYW contributed conceptualization, data collection, statistical analysis, data interpretation, writing – original draft, and writing – review and editing; TS and ZST contributed data collection, data interpretation, and writing – review and editing; XCH, WL, MY, YJL, QCO, XQL, JF, HPL, HL, WYC, CG, YET, LMX, MML, YPM, FL and LW contributed data collection and writing – review and editing; XHD contributed data collection, statistical analysis, and writing – review and editing; BHX contributed conceptualization, data collection, project administration, writing – original draft, and writing – review and editing. All authors read and approved the final manuscript.

Funding

The study was supported by the National Major Scientific and Technological Special Project for “Significant New Drugs Development” (No. 2018ZX09711002-011–027) and CAMS Innovation Fund for Medical Sciences (CIFMS, 2021-I2M-1–014 and 2023-12 M-2–004). The study was funded by Xuanzhu Biopharmaceutical Co., Ltd., a holding subsidiary of Sihuan Pharmaceutical Holdings Group Ltd., and was envisioned and carried out by the principal investigator in collaboration with the sponsor. Xuanzhu Biopharmaceutical Co., Ltd. was involved in data collection, analysis and interpretation of findings, and in writing the manuscript.

Data availability

Data collected within the study will be made available to researchers whose full proposal for their use of the data has been approved by the Trial Management Group, that includes a qualified statistician. The data required for the approved, specified purposes will be provided after completion of a data sharing agreement, that will be set up by the study sponsor. The data will be made available 2 years after publication. Please address requests for data to the corresponding author, BHX, at: xubh@cicams.ac.cn.

Declarations

Ethics approval and consent to participate

The study protocol and all amendments were approved by the Ethics Committees of all participating hospitals: the National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College (17–163/1419); the 307th Hospital of the Chinese People’s Liberation Army (2017–12-55–1); Peking University Cancer Hospital and Institute (2018YW16 and 2020YW126); the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Hunan Cancer Hospital (2019-Drug Review [23]); Fudan University Shanghai Cancer Center (1808189–1); the Affiliated Cancer Hospital of Zhengzhou University & Henan Cancer Hospital (2019087); the Fourth Hospital of Hebei Medical University, Hebei Tumor Hospital (2019036); the First Hospital of China Medical University (2020YL057); Institute of Oncology, Cancer Hospital of Dalian University of Technology, Liaoning Cancer Hospital (20190544); Cancer Center, the First Bethune Hospital of Jilin University (19Y081-001); Tianjin Medical University Cancer Institute and Hospital (E2019151); the Third Hospital of Nanchang (Drug (2020) Ethical No.32); Sichuan Cancer Hospital (SCCHEC-01A-2020–039); Sun Yat-sen Memorial Hospital, Sun Yat-sen University (2020-YW-093). All procedures were conducted in accordance with Good Clinical Practice guidelines and the Declaration of Helsinki. All patients provided written informed consent before being enrolled in the trial, which was registered with ClinicalTrials.gov (identification ID: NCT04539496).

Consent for publication

Not applicable.

Competing interests

LMX, XHD, MML, YPM, FL and LW are employees of Xuanzhu Biopharmaceutical Co., Ltd. BHX reported receiving consulting fees and payment for lectures from AstraZeneca and Novartis outside the submitted work. The remaining authors declare no potential competing interests.