A randomized trial comparing LY01011, biosimilar candidate, with the reference product denosumab (Xgeva®) in healthy Chinese subjects.

Introduction

Malignant tumor, one of the leading causes of human death, is a major public health concern worldwide. Bone is the most common site of metastasis in advanced malignancies. The commonly reported tumors prone to bone metastasis include breast cancer (70%), prostate cancer (85%), lung cancer (30%-40%), renal cancer (20%-25%) and multiple myeloma (95%) [1] . Bone metastasis, involving tumor-induced osteoclast activation, often result in skeletal tumor progression, severe bone pain and other multiple complications including skeletal-related events (SREs) and skeletal disorders. Skeletal lesions and SREs attributed to bone metastases not only greatly affect the patients’ functional status and quality of life, but also threaten to patients’ survival to some extents.

The common treatments for bone metastasis in advanced malignant tumors include bisphosphates, pain medications, steroid, chemotherapy, hormone therapy, targeted therapy, radiation therapy and surgery. Of which, bone modifying medications such as bisphosphate and denosumab are recommended as the preferred options for the treatment of bone metastasis in patients with prostate cancer [2] . However, in comparison with bisphosphates, denosumab (Xgeva®) takes the advantages of superior effectiveness, no nephrotoxicity and relatively more convenient dose administration, and therefore it has been recommended by the National Comprehensive Cancer Network (NCCN) for the treatment of bone metastasis in patients with malignant tumors [3] .

The receptor activator of nuclear Kappa-B ligand (RANKL) is an essential cytokine taking responsible for the osteoclast activation and differentiation. Aberrant expression of RANKL induced by metastatic tumor cells would primarily contribute to the pathological conditions characterized by excessive bone resorption in patients with bone metastasis [4] . Hence, numerous recombinant fully human anti-RANKL monoclonal antibodies have been developed to block the activation of osteoclasts and is used for the treatment of patients with bone metastasis.

Denosumab (Xgeva®) was approved by FDA in 2010 for the prevention of SREs in patients with multiple myeloma and in patients with bone metastasis from solid tumors [5] . In addition to that, it is also indicated for the treatment of treatment of hypercalcemia of malignancy refractory to bisphosphate therapy. In China, it was approved in 2020 for the same indication in bone metastasis patients as the reference listed drug does [6] . Denosumab acts as an inhibitor to block the RANKL from binding to its receptor RANK, which expresses on the surface of osteoclasts, for reduction of bone resorption. However, in some countries, access to the reference product is highly limited due to cost, and therefore biosimilar products are provided at lower costs, offering patients access to more treatment options by overcoming the cost barriers to some extent.

LY01011, a recombinant fully human anti-RANKL monoclonal antibody injection, is a biosimilar of the reference product denosumab (Xgeva®) and was developed by Shandong Boan Biotechnology Co, LTD. Preclinical studies, including PK, PD and pharmacological toxicities, showed no statistically significant differences between LY01011 and denosumab, which further supported the clinical development of LY01011.

Material and Methods

Study population

Healthy Chinese male or female subjects aged 18–50 years, inclusive, weighing ≥ 50 kg for males and weighing ≥ 45 kg for females with body mass index (BMI) of 18–28 kg/m 2 were eligible for the enrollment. All subjects are required to be assessed as no abnormality or no clinical significance regarding vital signs, physical examination, laboratory tests, 12- lead electrocardiogram (ECG), urinalysis screening and chest radiography at screening period. The main exclusion criteria were as follows: patients suffering from or having osteomyelitis or osteonecrosis of jaw, or planning to undergo dental or jaw surgery during the study, or those with unhealed dental or oral surgical wounds; fractures within the past 6 months; use of drugs that may affect bone turnover prior to screening or planned to be used during the study; subjects with abnormal serum calcium; any prior or current presence of clinically significant serious or chronic diseases or any disease conditions that would interfere with the study outcomes; participated in other clinical studies within 3 months prior to screening.

The single-center clinical study was conducted in compliance with the provisions of the Declaration of Helsinki, the International Conference on Harmonization E6 Guidelines on Good Clinical Practice, and local regulatory requirements. Written informed consents were provided to all subjects at the screening visits prior to initiation of any further procedure. All mandatory documents such as protocol and informed consent documents were reviewed and approved by the Institutional Review Board of the First Hospital of Jilin University.

Study design and treatment

It was a single-center, randomized, double-blind, single-dose, active-comparator, parallel-controlled conducted in the First Hospital of Jilin University. The study was registered at clinicaltrial.gov ( NCT04198636 ).

The study is to compare the PK, PD, safety and immunogenicity of LY01011 to denosumab following a single dose administration in healthy Chinese subjects. Six hundred and twenty five healthy Chinese subjects were screened, and 168 eligible subjects, stratified by baseline body weight (≥66 kg and＜66 kg), were randomized to receive a single subcutaneous administration of 120 mg LY01011 or denosumab in a 1:1 ratio with 85 subjects in LY01011 group and 83 subjects in denosumab group. PK, PD, safety, and immunogenicity data were collected for approximately 9 months.

Pharmacokinetic/Pharmacodynamics evaluation

PK parameters were analyzed using Phoenix WinNonlin (WNL) Version 8.1 software, and the non-compartmental analysis (NCA) method was used for estimating PK parameters. The PK parameters included in the study were C max, AUC 0→∞ , AUC 0→t , T max , CL/F, λ z , t 1/2 and Vz/F, where C max and AUC 0→ t were primary parameters and others were considered as secondary parameters. The plasma concentration of the study drug were measured at each collected time point in both groups.

PD parameters were analyzed using Statistical Analysis System (SAS) Version 9.4 software. The PD parameters included in the study were percentage change in serum C-telopeptide of type I collagen (sCTX-1) from baseline, AUEC 0-t , E max , and TE max . The plasma concentration of the study drug were measured at each collected time point in both groups.

Blood samples for PK/PD evaluations from each subject were collected at pre-dose (within 1 h prior to dosing), and at 6 h ± 10 min (D1), 24 h ± 1 h (D2), 72 h ± 1 h (D4), 120 h ± 4 h (D6), 168 h ± 4 h (D8), 216 h ± 4 h (D10), 264 h ± 4 h (D12), 312 h ± 4 h (D14), 480 h ± 4 h (D21), 648 h ± 1 d (D28), 984 h ± 2 d (D42), 1320 h ± 2 D (D56), 1656 h ± 3 d (D70), 1992 h ± 4 d (D84), 2328 h ± 4 d (D98), 2664 h ± 5 d (D112), 3000 h ± 6 d (D126), 3336 h ± 6 d (D140), 4008 h ± 7 d (D168), 4680 h ± 7 d (D196), 5352 h ± 7 d (D224), 6024 h ± 7 d (D252) following the study drug administration. 4 mL of whole blood were collected for each time point. The plasma concentration of LY01011 and denosumab as well as sCTX-1 were quantified by a validated electrochemiluminescence immunoassay (ECLIA).

Safety evaluation

Safety profiles were any adverse events (AEs) including TEAEs, SAEs, etc., vital signs, physical examination, laboratory tests, and 12-lead electrocardiogram (ECG). All AEs were coded via MedDRA Version 24.0 and assessed by Investigators for severity and relationship to the study drugs. All AEs were monitored throughout the study.

Immunogenicity evaluation

Blood samples were collected from each subject for immunogenicity evaluation at pre-dose (within 1 h prior to dosing) and at 312 h ± 4 h (D14), 648 h ± 1 d (D28), 1320 h ± 2 d (D56), 1992 h ± 4 d (D84), 2664 h ± 5 d (D112), 3336 h ± 6 d (D140), 4680 h ± 7 d (D196), and 6024 h ± 7 d (D252) following the study drug administration. The presence of anti-drug antibody (ADA) was detected through a validated Bridging-Electrochemiluminescent Immunoassay (ECLIA), and a validated competitive ligand binding assay technique (CLBA) and affinity capture elution (ACE) method was further performed to measure the neutralizing antibodies (Nabs).

Sample size and statistical Analysis

Assuming that the coefficient of variation between subjects was approximately 35%, geometric mean ratio of LY01011 to denosumab was 1.05 and the pre-specified acceptance range was 80%-125%. Estimation of 168 subjects enrolled in the study with 84 subjects per group would achieve a 90% of power at a 5% significance level by two one-sided tests, considering the dropout rate was 20%. PK similarity between LY01011 and denosumab was met if the 90% CIs of GMRs for primary PK parameters, AUC 0→t and C max , were in the range of 80%-125%. Other statistical analysis were conducted by SAS Version 9.4. Safety and anti-drug antibody sets were used for the evaluation of safety and immunogenicity of the study drug, respectively.

Results

Subject disposition, demographic and baseline characteristics

Following the screening, 168 eligible healthy male subjects were enrolled and randomized in a 1:1 ratio to receive either LY01011 or denosumab with 85 subjects in LY01011 group and 83 subjects in denosumab group. Eighty subjects completed the study in LY01011 group, whereas 78 subjects completed throughout the study in denosumab group ( Fig. 1 ). In Full Analysis Set (FAS), 38 (44.7%) male subjects and 47 (55.3%) female subjects were in LY01011 group, whereas 35 (42.2%) male subjects and 48 (57.8%) female subjects were in denosumab group, respectively. Majority of the subjects were Han Chinese (96.5% in LY01011 vs 97.6% in denosumab). The mean age (SD), height (SD), body weight (SD) and BMI (SD) in LY01011 group were 36.6 (8.20) years, 162.69 (8.067) cm, 62.46 (8.524) kg, and 23.5 (2.42) kg/m 2 . In denosumab group, the mean age (SD), height (SD), body weight (SD) and BMI (SD) were 37.1 (8.05) years, 162.75 (8.104) cm, 63.04 (8.460) kg, and 23.8 (2.20) kg/m 2 , respectively ( Table 1 ). Fig. 1 Subject Disposition. Table 1 Demographic and Baseline Characteristics. Metrics LY01011 Group (N = 85) Denosumab Group (N = 83) Total (N = 168) Age (years) N 85 83 168 Mean ± SD 36.6 ± 8.20 37.1 ± 8.05 36.9 ± 8.11 Q1 ～ Q3 31.0 to 43.0 32.0 ～ 44.0 31.0 to 43.5 Min to Max 20–50 18–50 18–50 Median 36.0 39.0 38.0 Sex n (%) M 38 (44.7) 35 (42.2) 73 (43.5) F 47 (55.3) 48 (57.8) 95 (56.5) Total 85 83 168 Races n (%) Asian 85 (100.0) 83 (100.0) 168 (100.0) Others 0 (0) 0 (0) 0 (0) Total 85 83 168 Ethnicity n (%) Han Chinese 82 (96.5) 81 (97.6) 163 (97.0) Others 3 (3.5) 2 (2.4) 5 (3.0) Total 85 83 168 Height (cm) N 85 83 168 Mean ± SD 162.69 ± 8.067 162.75 ± 8.104 162.72 ± 8.061 Q1 ～ Q3 156.70 to 169.00 157.50 to 169.20 156.95 to 169.10 Min to Max 145.8 to 179.2 146.8 to 178.2 145.8 to 179.2 Median 162.60 162.30 162.45 Body weight (kg) N 85 83 168 Mean ± SD 62.46 ± 8.524 63.04 ± 8.460 62.75 ± 8.472 Q1 ～ Q3 56.00 ∼ 68.60 57.30 ∼ 67.40 56.30 ～ 67.85 Min to Max 46.1 to 83.3 46.5 to 85.2 46.1–85.2 Median 62.40 62.00 62.30 Stratified by body weight ≥ 66 kg 30 (35.3) 26 (31.3) 56 (33.3) < 66 kg 55 (64.7) 57 (68.7) 112 (66.7) Total 85 83 168 BMI (kg/m2) N 85 83 168 Mean ± SD 23.5 ± 2.42 23.8 ± 2.20 23.7 ± 2.31 Q1 ～ Q3 22.0 ～ 25.0 22.0–26.0 22.0–25.5 Min to Max 18 to 28 20–28 18 to 28 Median 24.0 24.0 24.0 BMI = body mass index. The percentage (%) was based upon the number of subjects in each treatment group from the full analysis set.

All the healthy subjects in the study were analyzed for demographic and baseline characteristics per FAS, and the baselines were comparable between LY01011 group and denosumab group.

Pharmacokinetic/pharmacodynamics evaluations

As shown in Fig. 2 , the mean (±SD) plasma concentration–time profiles of denosumab exhibited a similar pattern over the course of sampling following a single-dose subcutaneous administration of either LY01011 or the reference drug, denosumab in the study. Fig. 2 Mean (±SD) plasma concentration–time profiles of denosumab following a single 120 mg subcutaneous administration of LY01011 or denosumab in linear and semi-log scales.

The primary and secondary PK parameters were comparable between LY01011 group and denosumab group following a single 120 mg dose administration ( Table 2 ). The GMRs for C max , AUC 0–t and AUC 0–∞ ,in the study were 98.13%, 100.32% and 100.39%, respectively. 90% CIs of GMRs for primary PK parameters, C max , AUC 0–t and AUC 0–∞ , were within predefined acceptance range of 80%-125%. In line with primary PK parameters, the secondary PK parameters such as AUC 0–∞ with GMR (100.39%) also fell within the range of 80%-125% ( Table 3 ). Table 2 PK Parameters Analysis Set (PKPS). PK Parameters (units) LY01011 Group (N = 85) Denosumab Group (N = 83) Cmax (μg/mL) 11.05 ± 4.21 (38.1) 11.46 ± 4.22 (36.8) AUC0-t (day * μg/mL) 540.48 ± 189.71 (35.1) 554.87 ± 200.37 (36.1) AUC0- ∞ (day* μg/mL) 546.30 ± 191.01 (35.0) 560.13 ± 201.47 (36.0) Tmax (day) 9.03 (5.00 ～ 27.06) 9.00 (3.00 ～ 27.03) T1/2 (day) 19.30 ± 5.80 (30.0) 20.20 ± 6.87 (34.0) AUC_% Extrap (%) 1.09 ± 0.69 (63.3) 1.01 ± 0.96 (95.1) Vz/F (mL) 6537.21 ± 2148.66 (32.9) 6899.69 ± 3533.02 (51.2) CL/F (mL/day) 247.79 ± 90.73 (36.6) 254.17 ± 149.89 (59.0) Note: For subjects with missing three or more consecutive plasma concentrations (subjects No. 1015、1024、2014、2016、2027、2043、2048 in LY01011 group; subjects No. 1007、1014、1016、1017、1021、2003、2015、2026、2041、2042 in denosumab group) after T max , only C max and T max were included in PKPS and other parameters would be precluded from PKPS. T max was median (min–max), other parameters were mean ± SD (%CV). Table 3 C max 、AUC 0-t 、AUC 0-∞ by Log-Transformed PK Similarity Analysis. LY01011 Group /Denosumab Group PK Parameter (Units) Treatment Groups N Geometric LS Mean 95% CI Ratio (%) 90% CI Cmax (μg/mL) LY01011 group 85 9.92 9.12 to 10.80 98.13 88.96 ～ 108.26 Denosumab group 83 10.11 9.27 to 11.03 AUC0-t (day * μg/mL) LY01011 group 78 487.97 449.26 to 530.00 100.32 91.04 to 110.54 Denosumab group 73 486.40 445.79 to 530.72 AUC0- ∞ (day* μg/mL) LY01011 group 78 493.43 454.46 to 535.74 100.39 91.15 ～ 110.57 Denosumab group 73 491.51 450.64 to 536.08 Note: CI = Confidence Interval; LS = least square. For subjects with missing three or more consecutive plasma concentrations (subjects No. 1015、1024、2014、2016、2027、2043、2048 in LY01011 group; subjects No. 1007、1014、1016、1017、1021、2003、2015、2026、2041、2042 in denosumab group) after T max , only C max was included in PKPS and other parameters would be precluded from PKPS.

As shown in Fig. 3 , the mean (±SD) sCTX-1 concentration–time profiles and change-rate time profiles of LY01011 and denosumab displayed similar trending over the course of time following a single-dose subcutaneous administration. Fig. 3 Mean (±SD) sCTX-1 concentration–time profiles and change rate-time profiles following a single 120 mg subcutaneous administration of LY01011 or denosumab.

The PD parameters between LY01011 group and denosumab group following a single 120 mg dose administration were comparable ( Table 4 ). The GMRs for AUEC 0-t and E max in the study were 98.71% and 99.80%. 90% CIs of GMRs for PD parameters were within pre-specified acceptance range of 80%-125%. ( Table 5 ). Table 4 PD Parameters Analysis Set (PDPS). PD Parameters (units) LY01011 Group (N = 85) Denosumab Group (N = 83) AUEC0-t (day* %) − 19376.432 ± 2395.4933 (-12.4) − 19634.136 ± 2243.7914 (-11.4) Emax (%) − 87.654 ± 6.1445 (-7.0) − 87.759 ± 5.2627 (-6.0) TEmax (day) 69.000 (7–167.01) 83.000 (5–167.04) Note: For subjects with missing three or more consecutive PD data (Subjects No. 2014, 2016, 2027, 2043, 2048 in LY01011 group; subjects No. 1016, 1017, 2003, 2041 and 2042 in denosumab group), only E max and TE max were included in PDPS, while AUEC0-t was excluded from PDPS. TE max was median (min–max), other parameters were mean ± SD (%CV). Table 5 AUEC 0-t , and E max by Log-Transformed PD Similarity Analysis. LY01011 Group /Denosumab Group Parameter (Units) Treatment Groups N Geometric LS Mean 95% CI Ratio (%) 90% CI AUEC0-t (day*%) LY01011 group 80 − 19134.77 − 19687.14 to −18597.90 98.71 95.48 to 102.05 Denosumab group 78 − 19384.79 − 19962.28 to −18824.00 Emax (%) LY01011 group 85 − 87.44 − 88.75 ～ −86.15 99.80 98.08 ～ 101.54 Denosumab group 83 − 87.62 − 88.97 ～ −86.29 Note: CI = Confidence Interval; LS = least square. For subjects with missing three or more consecutive PD data (Subjects No. 2014, 2016, 2027, 2043, 2048 in LY01011 group; subjects No. 1016, 1017, 2003, 2041 and 2042 in denosumab group), only E max was included in PDPS, while AUEC0-t was excluded from PDPS.

Safety evaluations

All AEs per safety set occurred throughout the study were summarized in Table 6 . Table 6 Summary of Treatment-Emergent Adverse Events-Safety Set. SOC PT LY01011 Group (N = 85) Denosumab Group (N = 83) Total (N = 168) N (%) Events N (%) Events N (%) Events Study drug with at least one TEAEs Study drug related TEAEs 83 (97.6) 82 (96.5) 383 301 80 (96.4) 78 (94.0) 343 286 163 (97.0) 160 (95.2) 726 587 Investigations 78 (91.8) 167 70 (84.3) 142 148 (88.1) 309 Blood parathyroid hormone increased 77 (90.6) 139 68 (81.9) 113 145 (86.3) 252 Alanine aminotransferase increased 9 (10.6) 9 6 (7.2) 8 15 (8.9) 17 Aspartate aminotransferase increased 5 (5.9) 5 6 (7.2) 8 11 (6.5) 13 Blood bilirubin increased 3 (3.5) 3 3 (3.6) 3 6 (3.6) 6 White blood cell count increased 2 (2.4) 2 2 (2.4) 2 4 (2.4) 4 Neutrophil count increased 2 (2.4) 2 2 (2.4) 2 4 (2.4) 4 White blood cell count decreased 1 (1.2) 1 1 (1.2) 2 (1.2) 3 Blood creatinine increased 1 (1.2) 1 1 (1.2) 1 (1.2) 2 Neutrophil count decreased 1 (1.2) 1 1 (1.2) 1 (1.2) 2 Low density lipoprotein increased 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Lymphocyte count decreased 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Electrocardiogram T wave abnormal 1 (1.2) 2 0 (0) 0 1 (0.6) 2 Blood thyroid stimulating hormone decreased 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Tri-iodothyronine free increased 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Metabolism and nutrition disorders 54 (63.5) 102 55 (66.3) 110 109 (64.9) 212 Hypophosphataemia 47 (55.3) 83 49 (59.0) 93 96 (57.1) 176 Hypertriglyceridemia 6 (7.1) 7 4 (4.8) 4 10 (6.0) 11 Hypocalcemia 3 (3.5) 3 6 (7.2) 6 9 (5.4) 9 Hyperuricaemia 3 (3.5) 3 2 (2.4) 2 5 (3.0) 5 Hyperglycaemia 2 (2.4) 6 1 (1.2) 2 3 (1.8) 8 Hypoalbuminaemia 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Hyperkalemia 0 (0) 0 1 (1.2) 2 1 (0.6) 2 Renal and urinary disorders 6 (7.1) 6 10 (12.0) 11 16 (9.5) 17 Haematuria 3 (3.5) 3 6 (7.2) 6 9 (5.4) 9 Proteinuria 3 (3.5) 3 5 (6.0) 5 8 (4.8) 8 Infections and infestations 8 (9.4) 9 7 (8.4) 8 15 (8.9) 17 Urinary tract infection 7 (8.2) 7 7 (8.4) 8 14 (8.3) 15 Herpes zoster 1 (1.2) 2 0 (0) 0 1 (0.6) 2 Cardiac disorders 4 (4.7) 5 3 (3.6) 3 7 (4.2) 8 Supraventricular extrasystoles 1 (1.2) 1 2 (2.4) 2 3 (1.8) 3 Arrhythmia supraventricular 1 (1.2) 2 0 (0) 0 1 (0.6) 2 Ventricular extrasystole 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Palpitations 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Sinus tachycardia 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Gastrointestinal disorders 5 (5.9) 8 0 (0) 0 5 (3.0) 8 Noninfective gingivitis 2 (2.4) 2 0 (0) 0 (1.2) 2 Stomatitis 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Vomiting 1 (1.2) 2 0 (0) 0 1 (0.6) 2 Toothache 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Eructation 1 (1.2) 2 0 (0) 0 1 (0.6) 2 Blood and lymphatic system disorders 1 (1.2) 1 4 (4.8) 6 5 (3.0) 7 Anaemia 1 (1.2) 1 4 (4.8) 6 5 (3.0) 7 General disorders and administration site conditions 0 (0) 0 3 (3.6) 3 3 (1.8) 3 Pyrexia 0 (0) 0 3 (3.6) 3 3 (1.8) 3 Respiratory, thoracic and mediastinal disorders 1 (1.2) 2 1 (1.2) 1 (1.2) 3 Cough 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Oropharyngeal pain 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Rhinorrhoea 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Skin and subcutaneous tissue disorders 1 (1.2) 1 1 (1.2) 1 (1.2) 2 Alopecia areata 1 (1.2) 1 0 (0) 0 1 (0.6) 1 Pruritus 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Nervous system disorders 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Headache 0 (0) 0 1 (1.2) 1 1 (0.6) 1 Note: SOC = System Organ Class; PT = Preferred Term; TEAE = Treatment-Emergent Adverse Event. The percentages (%) were based upon the number of subjects in each treatment group from the safety set.

All the reported AEs in the study were TEAEs, and in total, 163 subjects (97.0%) had experienced 726 TEAEs. Of whom, 83 subjects (97.6%) experienced 383 TEAEs in LY01011 group and 80 subjects (96.4%) with 343 TEAEs were reported in denosumab group. Among 83 subjects in LY01011 group, 82 subjects (96.5%) were reported to have 301 TEAEs, which were deemed as study drug related TEAEs. Likewise, 78 subjects (94.0%) in denosumab group had 286 TEAEs related to the study drug.

Majority of the TEAEs occurred in the study were categorized as Grade 1 in severity.Most of the subjects experiencing TEAEs were fully recovered. 62 (72.9%) and 54 (65.1%) subjects were reported to have 344 and 305 Grade 1 TEAEs in LY01011 group and denosumab group. 34 and 36 Grade 2 TEAEs were observed in 17 (20.0%) and 24 (28.9%) subjects in LY01011 group and denosumab group. 4 (4.7%) and 2 (2.4%) subjects had experienced 5 and 2 TEAEs of Grade 3 or above in LY01011 group and denosumab group. The severities of TEAEs between the two treatment groups were comparable.

Majority of the study drug related TEAEs occurred in the study were categorized as Grade 1 in severity, which most of them were fully recovered. 68 (80.0%) and 64 (77.1%) subjects were reported to have 282 and 266 Grade 1 study drug related TEAEs in LY01011 group and denosumab group. 19 and 20 study drug related TEAEs of Grade 2 were observed in 14 (16.5%) and 14 (16.9%) subjects in LY01011 group and denosumab group. No Grade 3 or above study drug related TEAEs were reported in both groups, and the severities of study drug related TEAEs between the two treatment groups were comparable.

The TEAEs, defined as ≥ 5% of the subjects experienced in either group (LY01011 vs. denosumab) included blood parathyroid hormone increased (90.6% vs. 81.9%), hypophosphatemia (55.3% vs. 59.0%), alanine aminotransferase increased (14.1% vs. 8.4%), urinary tract infection (11.8% vs. 13.3%), aspartate aminotransferase increased (10.6% vs. 7.2%), hypertriglyceridemia (9.4% vs. 8.4%), hematuria (4.7% vs. 8.4%), hypocalcemia (3.5% vs. 7.2%), anemia (3.5% vs. 7.2%), proteinuria (3.5% vs. 6.0%), blood bilirubin increased (3.5% vs. 6.0%), white blood cell count increased (3.5% vs. 6.0%) and neutrophil count increased (3.5% vs. 6.0%). Blood parathyroid hormone increased and hypophosphatemia were the most commonly reported TEAEs in LY01011 group and denosumab group.

The TEAEs of Grade 3 or above in LY01011 group and denosumab group were blood parathyroid hormone increased (1.2% vs. 0%), alanine aminotransferase increased (1.2% vs. 0%), aspartate aminotransferase increased (1.2% vs. 0%), lumbar vertebral fracture (1.2% vs. 0%), scrotal injury (0% vs. 1.2%) and syncope (0% vs. 1.2%), which were unlikely related to the study drug at the discretion of the Investigator.

Blood parathyroid hormone increased (90.6% vs. 81.9%), hypophosphatemia (55.3% vs. 59.0%), alanine aminotransferase increased (10.6% vs. 7.2%), aspartate aminotransferase increased (5.9% vs. 7.2%), hypertriglyceridemia (7.1% vs. 4.8%), hypocalcemia (3.5% vs. 7.2%), hematuria (3.5% vs. 7.2%), proteinuria (3.5% vs. 6.0%), and urinary tract infection (8.2% vs. 8.4%) were regarded as study drug related TEAEs, which were defined as ≥ 5% of the subjects experienced in either group (LY01011 vs. denosumab).

A total of 3 (1.8%) subjects experiencing 6 TEAEs were reported as SAEs, which led to the subject withdrawal from the study. Of 6 SAEs, contusion (Grade 1), neck injury (Grade 1), head injury (Grade 1) and lumbar vertebral fracture (Grade 3) were reported in LY01011 group, whereas ectopic pregnancy (Grade 1) and scrotal injury (Grade 3) were observed in denosumab group. All SEAs in the study were considered to be unlikely related to the study drug.

Laboratory tests, vital signs, physical examination, and 12-lead ECG results were generally similar with no clinically meaningful differences between LY01011 group and denosumab group.

No TEAEs leading to death were reported in either group throughout the study.

Immunogenicity evaluations

In the study, all the subjects were tested ADA negative at baseline prior to the study drug administration. Only one subject (1.2%) with ADA positive was reported on Day 196 (approximately at 4680 h) in denosumab group. No subjects were detected Nab positive, irrespective of the study drug administration.

Discussion

LY01011, a recombinant fully human anti-RNAKL monoclonal antibody, was developed by Shandong Boan Biotechnology Co. Ltd, and was processed as a biosimilar to the reference product dneosumab (Xgeva®). LY01011 has the same active pharmaceutical ingredient as denosumab along with dose strength and dose administration for demonstration of biosimilar between the two products. LY01011 is similar to denosumab regarding the manufacturing process, quality control, pharmacological studies and data from non-clinical studies.

A single 120 mg dose of LY01011 was administered to the subject subcutaneously, which was in line with what it was referred in the reference drug labeling. The mean elimination half-life of denosumab was 28 days. The sampling time collected in the study should be 3–5 times half-lives per the guidance for clinical PK studies enacted by National Medical Products Administration, and hence the data was collected until Day 252 (greater than5 times half-lives) [7] . Biomarkers of bone metabolism are the key indicators in support of the clinical diagnosis and treatment effect. Despite many other available biochemical markers, sCTX is still recommended as the preferred biomarker for the evaluation of bone resorption [8] .

To minimize the confounding effects resulted from the comorbidities and concomitant medication, eligible healthy Chinese subjects were selected to undergo the randomization process. A randomized, double-blind, single-dose, active-comparator, controlled phase 1 study was conducted to compare PK/PD, safety and immunogenicity of biosimilar product, LY01011, to its reference drug denosumab following a single dose of 120 mg subcutaneous administration in healthy Chinese population. All the PK parameters in LY01011 group and denosumab group were comparable as represented in Fig. 2 and Table 2 . PK similarity in terms of the primary PK parameters was achieved since GMRs of C max and AUC 0→t were 98.13% (90% CI: 88.96%- 108.26%) and 100.32% (90% CI: 91.04%-110.54%), which fell within the pre-defined acceptance range of 80%-125%. Similarly, GMRs of the secondary PK parameters such as AUC 0–∞ (100.39%; 90% CI: 91.15%-110.57%) were also within the acceptance range of 80%-125%, demonstrating LY01011 was biosimilar to denosumab.

In addition, PK parameters in the study, including but not limited to, AUC 0–t , C max , T max and t 1/2 , were consistent with those reported in the previous clinical studies. A randomized, double-blind, active-controlled study was to compare the PK/PD, safety and immunogenicity of a single 120 mg dose of denosumab biosimilar (MW032) to denosumab (Xgeva®) in healthy Chinese population [9] . Following a single dose administration, the mean C max, AUC 0→t , the median T max and the mean t 1/2 were 10.8 μg/mL (vs. 11.05 μg/mL in this study), 561.9 day*μg/mL (vs. 540.48 day*μg/mL in this study), 7 days (vs. 9.03 days in this study) and 20.01 days (vs. 19.3 days in this study), respectively. Another randomized, placebo-controlled study that was reported to assess the PK/PD and safety of a single 60 mg and 120 mg of denosumab in healthy Chinese subjects also showed the PK results following a 120 mg dose were congruent with the ones in this study [10] .

All the PD parameters in LY01011 group and denosumab group were comparable as represented in Table 4 . The mean sCTX-1concentration-time curves measured in LY01011 group and denosumab group were greatly similar following a single subcutaneous dose administration in healthy Chinese subjects ( Fig. 3 ). Mean sCTX-1 concentrations declined rapidly by 6 h with more than 80% reduction from baseline on Day 6 following the dose administration in both groups. The reduction of mean sCTX-1concentration continued and lasted until Day 140. The maximum mean percentage reductions were 85.27% and 85.02% in LY01011 group and denosumab group during the study. The levels of mean sCTX-1 decreased in a similar pattern between the two groups, which were consistent with the historical data of the reference drug denosumab. The GMRs of PD parameters such as AUEC 0-t and E max were 98.71% (90% CI: 95.48%- 102.05%) and 99.80% (90% CI: 98.08%-101.54%). PD similarity was concluded given the data fell within the range of 80.00% to 125.00%.

The incidences of TEAEs and study drug related TEAEs during the study were similar between the two groups ( Table 6 ). TEAEs in the study were comparable with those listed in the reference drug labeling, for instance, hypophosphatemia, vomiting and headache. In LY01011 group, 82 subjects (96.5%) had 301 TEAEs, which were deemed to be related to LY01011. Meanwhile, 78 subjects (94.0%) in denosumab group had 286 TEAEs that were treated as study drug related TEAEs. Majority of TEAEs were assessed as Grade 1 in severity. 4 (4.7%) and 2 (2.4%) subjects had experienced 5 and 2 TEAEs of Grade 3 or above in LY01011 group and denosumab group, which were considered to be unlikely related to the study drug. In LY01011 group, one subject (1.2%) experienced 4 SAEs and 2 subjects (2.4%) in denosumab group had 2 SAEs. Only one Grade 3 or above SAE was reported in either group, which was fully recovered momentarily. All SAEs, leading to the subject withdrawal from the study, were deemed to be unlikely related to the study drug.

Moreover, the common study drug related TEAEs were similar to those reported in a previous randomized, placebo-controlled study in healthy Chinese volunteers. Between the two treatment groups, there were no clinically significant differences in terms of vital signs, physical examination, other relevant lab tests and 12-lead ECG, demonstrating LY01011 and denosumab both were acceptable and tolerable in healthy Chinese subjects with similar safety profiles.

Prior to the study drug administration, all subjects were tested ADA negative at baseline, and only one positive ADA result was reported in one subject in denosumab group. Nab was negative regardless of study drug administration in either group. ADA had no impact on the PK and PD of denosumab.

Stratification by sex has not been done in the study due to relatively small coefficient of variation. Plus, per labeling of the reference drug denosumab, PK was not affected by age, gender and race [5] . Since LY01011 is biosimilar to denosumab, sex stratification indeed was not necessary to be warranted. Several clinical studies pertaining to the development of denosumab biosimilar in healthy and bone metastasis population from other biopharmaceutical companies were either completed or are currently ongoing. A randomized, double-blind, active comparator controlled phase 1 study, comparing the PK/PD, safety and immunogenicity of MB09 (denosumab biosimilar) with EU/US-sourced Xgeva® in Chinese healthy male subjects, was completed and registered at Clinicaltrial.gov ( NCT05299073 ). A randomized, single-dose phase 1 study was conducted to evaluate the biosimilarity of QL1206 (denosumab biosimilar) to Xgeva® among Chinese healthy subjects ( NCT03651947 ). Besides that, a randomized, double-blind, active-controlled phase 3 study (CTR20223114) is ongoing in China to assess the efficacy, safety and immunogenicity of HL05 (denosumab biosimilar) in patients with bone metastasis from solid tumors. Recently, Sandoz has announced that FDA has already accepted the Biologics License Application for the proposed denosumab biosimilar.

Furthermore, Boan’s another product LY06006, the first global biosimilar to denosumab (Prolia®), was approved by National Medical Products Administration in 2022 for the treatment of osteoporosis in postmenopausal women with high risk of bone fracture. An ongoing phase 3 study of LY01011 is conducted to compare the efficacy, safety and immunogenicity of LY01011 with that of the reference product denosumab (Xgeva®) in bone metastasis patients with solid tumors.

Conclusion

The phase 1 study demonstrated that the PK and PD profiles of denosumab biosimilar, LY01011, were comparable to its reference product, denosumab in healthy Chinese subjects following a single dose of 120 mg subcutaneous administration. Overall, the safety profiles were tolerable and comparable between the two treatment groups. No new, unexpected study drug related safety signals were identified in either group. Immunogenicity of LY01011 was similar to that of denosumab.

Funding

This work was supported by the sponsor, Shandong Boan Biotechnology Co., Ltd. The sponsor fulfilled the responsibilities according to the Good Clinical Practice without interfering with implementing clinical trials.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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