
==== Front
Diabetes Ther
Diabetes Ther
Diabetes Therapy
1869-6953
1869-6961
Springer Healthcare Cheshire

39008234
1617
10.1007/s13300-024-01617-3
Original Research
ISIS 449884 Injection Add-On to Metformin in Patients with Type 2 Diabetes: A Randomized, Double-Blind, Placebo-Controlled, Phase II Clinical Study
Ji Linong jiln@bjmu.edu.cn

1
Gao Leili 1
Feng Zhikai 2
Chen Guoliang 2
Fu Jing 2
Morgan Erin 3
Bhanot Sanjay 3
Gao Shan 2
Zhang Hongyan 2
Liang Zicai 2
Gan Li-Ming li-ming.gan@ribocure.com

2456
1 https://ror.org/035adwg89 grid.411634.5 0000 0004 0632 4559 Department of Endocrinology and Metabolism, Peking University People’s Hospital, No. 11 Xi zhi men South Street, Xicheng District, Beijing, 100044 China
2 Ribo Life Science Co Ltd, Suzhou, Jiangsu China
3 https://ror.org/00t8bew53 grid.282569.2 0000 0004 5879 2987 Ionis Pharmaceuticals, Carlsbad, CA USA
4 Ribocure Pharmaceuticals AB, Gothenburg, Sweden
5 https://ror.org/04vgqjj36 grid.1649.a 0000 0000 9445 082X Department of Cardiology, Sahlgrenska University Hospital, Gothenburg, Sweden
6 https://ror.org/01tm6cn81 grid.8761.8 0000 0000 9919 9582 Department of Molecular and Clinical Medicine, Institute of Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden
15 7 2024
15 7 2024
10 2024
15 10 21832196
22 5 2024
26 6 2024
© The Author(s) 2024, corrected publication 2024
2024
https://creativecommons.org/licenses/by-nc/4.0/ Open Access This article is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License, which permits any non-commercial use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc/4.0/.
Introduction

ISIS 449884, a 2′-O-methoxyethyl antisense oligonucleotide that targets the glucagon receptor (GCGR), has demonstrated an ability to reduce hepatic glucose output and lower the blood glucose level. The primary objective of this study was to investigate the safety and efficacy of ISIS 449884 as an add-on to metformin in a population of Chinese patients with type 2 diabetes mellitus (T2DM).

Method

This was a multicenter, placebo-controlled (2:1), randomized, double-blind, parallel-enrollment, multiple-dose phase II study in Chinese patients with T2DM. A total of 90 patients who were uncontrolled by stable metformin monotherapy were randomized into three cohorts. Thirty subjects were enrolled in each cohort and received injections of ISIS 449884 (50 mg or 60 mg weekly or 100 mg every other week) or a corresponding volume of placebo (0.25 mL and 0.3 mL weekly or 0.5 mL every other week) subcutaneously in a 2:1 ratio for 16 weeks.

Results

The primary efficacy endpoint was analyzed in 88 subjects (ISIS 449884, n = 59; placebo, n = 29). The corrected LS mean change from baseline in glycated hemoglobin (HbA1c) at week 17 in the pooled ISIS 449884 treatment group was − 1.31% (95% CI − 1.66%, − 0.96%), and that in the pooled placebo group was 0.15% (95% CI − 0.37%, 0.66%). The LS mean difference between the two groups was − 1.46% (95% CI − 1.92%, − 1.00%, P < 0.001). Treatment-emergent adverse events (TEAEs) occurred in 53/60 subjects (88.3%) and 25/30 subjects (83.3%) in the pooled ISIS 449884 treatment group and the pooled placebo group, respectively, with similar incidences. Drug-related TEAEs occurred in 41/60 subjects (68.3%) and 9/30 subjects (30.0%), respectively. TEAEs of grade 3 or higher occurred in 5/60 (8.3%) subjects and 2/30 (6.7%) subjects, respectively, and none of them were drug related.

Conclusions

The ISIS 449884 injection add-on to metformin significantly reduced HbA1c in patients with T2DM uncontrolled by stable metformin monotherapy and showed an acceptable benefit/risk profile.

Clinical Trial Registration

www.chinadrugtrials.org.cn, CTR20191096.

Supplementary Information

The online version contains supplementary material available at 10.1007/s13300-024-01617-3.

Keywords

ISIS 449884
Glucagon receptor
Type 2 diabetes mellitus
Metformin
issue-copyright-statement© Springer Healthcare Ltd., part of Springer Nature 2024
==== Body
pmcKey Summary Points

Why carry out this study?	
Continuous research is required to explore novel mechanisms and develop drugs targeting new pathways in order to enhance blood glucose control and address the evolving clinical demands in the management of type 2 diabetes mellitus (T2DM).	
Efficacy and safety data on ISIS 449884 injection as an add-on to metformin in Chinese patients with T2DM who were uncontrolled by metformin monotherapy are still lacking.	
What was learned from this study?	
The ISIS 449884 injection add-on to metformin significantly reduced HbA1c in patients with T2DM uncontrolled by stable metformin monotherapy and showed an acceptable benefit/risk profile.	
This study provided clinical data on the application of ISIS 449884 injection in the Chinese T2DM population.	

Introduction

Diabetes mellitus (DM) is a collection of clinical syndromes characterized by an elevated blood glucose level. It results from a combination of genetic and environmental factors. The clinical symptoms of typical patients include polyuria, polydipsia, polyphagia and emaciation. Based on data from the International Diabetes Federation, the global prevalence of diabetes mellitus was estimated to be 537 million in 2021, with a projected increase to 783 million by the year 2045 [1]. China leads the world in the number of people with type 2 diabetes mellitus (T2DM), and the incidence of diabetes mellitus in China has increased significantly, highlighting the urgent need for effective diabetes management strategies in China [2]. The exact causes and mechanisms underlying the development of T2DM are still not fully understood, but it is characterized by a combination of insulin resistance and impaired insulin secretion, leading to an elevated blood glucose level [3].

T2DM treatment aims to maintain good glycemic control, which can prevent complications. Although the Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial and the Veterans Affairs Diabetes Trial (VADT) reported no significant effect on the rates of major cardiovascular events, the Action in Diabetes and Vascular Disease: Preterax and Diamicron Modified Release Controlled Evaluation (ADVANCE) trial indicated that intensive control of glucose has an important role in the prevention of microvascular complications of type 2 diabetes [4–6]. Metformin is the first-line drug, and optional combination therapies are based on individual conditions [7]. Data collected over the period of 2013 to 2018 in mainland China revealed that among adults with diabetes across the country, 36.7% were aware of their diabetes, 32.9% were receiving treatment for their diabetes, and the diabetes was under adequate control in 50.1%, all of which are low levels [8]. Thus, continuous research is needed to explore new mechanisms and develop drugs to enhance blood glucose control and meet the evolving clinical demands of T2DM management.

Glucagon, a hormone secreted by pancreatic alpha cells, antagonizes insulin action and plays a role in elevating the blood glucose level by binding to and activating glucagon receptor (GCGR). The dysregulation of glucagon secretion, which leads to the overproduction of hepatic glucose, contributes to hyperglycemia in type 2 diabetes [9]. In patients with T2DM, in addition to insufficient action of insulin itself, dysregulation of the glucagon/insulin balance leads to enhanced glucagon action and impaired postprandial glucagon suppression, which are important causes of elevated blood glucose [10]. Thus, blocking the production of GCGR protein could theoretically block the action of glucagon in patients with T2DM, thereby lowering the blood glucose level. ISIS 449884 is a second-generation 2′-O-(2-methoxyethyl) (2′-MOE) chimeric antisense nucleic acid inhibitor that targets GCGR. It is designed to be complementary to intron 1 of human GCGR and binds to 17 bases of GCGR mRNA through nucleic acid base pairing. This binding action recruits RNase H1, which leads to the degradation of GCGR mRNA and the consequent inhibition of GCGR protein formation. By preventing the production of GCGR protein, the goal of lowering blood glucose is achieved. Also, clinical studies indicated that ISIS 449884 may inhibit the action of glucagon in hepatic glycogen conversion by decreasing the expression level of GCGR in liver, thereby reducing hepatic glucose output and controlling the blood glucose level [11].

Currently, efficacy and safety data on ISIS 449884 injection added to metformin in Chinese patients with T2DM who were uncontrolled by metformin monotherapy are still lacking. This phase II clinical study was conducted in China to investigate the efficacy, safety, and tolerability of the ISIS 449884 injection add-on to metformin compared to metformin monotherapy in patients with T2DM who were uncontrolled by metformin monotherapy so as to provide clinical data on the use of ISIS 449884 injection in the T2DM population.

Methods

Study Design and Participants

This was a multicenter, placebo-controlled (2:1), randomized, double-blind, parallel-enrollment, multiple-dose phase II study to evaluate the efficacy, safety, and tolerability of the ISIS 449884 injection add-on to metformin in Chinese patients with T2DM who were uncontrolled by stable metformin monotherapy.

The trial was conducted in accordance with the 2013 Declaration of Helsinki and the International Conference on Harmonization’s Good Clinical Practice guidelines and was approved by the institutional review board (IRB) of Peking University People's Hospital (the main center; approval number: 2019PHA018-001) and the relevant institutional review board at each study site. A list of the institutional ethics committees (IECs)/IRBs that reviewed and approved the protocol are provided in Supplemental Table S2. Informed consent was obtained from all participants prior to enrollment in the study.

Eligible participants had to meet the following requirements: they voluntarily participated in the study and signed the informed consent form; aged 18–75 years (inclusive), male or female; diagnosed with T2DM according to the World Health Organization (WHO) 1999 criteria; male body weight ≥ 50 kg, female ≥ 45 kg, and body mass index (BMI) ≥ 18 kg/m2; receiving an ongoing stable dose of metformin monotherapy (≥ 1500 mg/day or the maximum tolerated dose as judged by the investigator) for 8 weeks prior to screening; 7.5% ≤ HbA1c ≤ 10.5% at screening and prior to randomization, and fasting glucose (serum) < 16.0 mmol/L at the screening visit; women of childbearing potential had a negative pregnancy test prior to dosing; the participant was willing and able to comply with the requirements of the protocol. Key exclusions included prior treatment with nucleotides or oligonucleotides (including small interfering ribonucleic acids (siRNAs)); three or more serious hypoglycemia events within 6 months prior to screening; the presence of proliferative diabetic retinopathy, severe diabetic peripheral neuropathy, or diabetic foot ulcers; uncontrolled hypertension, defined as a mean systolic blood pressure (SBP) ≥ 160 mmHg and/or a diastolic blood pressure (DBP) ≥ 100 mmHg at screening.

Procedures

A total of 90 subjects were randomized into three cohorts. The 30 subjects per dose cohort received ISIS 449884 injections of 50 mg or 60 mg weekly or 100 mg every other week in a 2:1 randomization ratio or corresponding volumes (0.25 mL, 0.3 mL, and 0.5 mL) of placebo by subcutaneous injection. The study was divided into four periods: a screening period, a run-in period, a treatment period, and a follow-up period. In the screening period, patients with T2DM who were receiving a stable dose of metformin monotherapy (≥ 1500 mg/day or the maximum tolerated dose as judged by the investigator) within 8 weeks prior to screening were screened. After signing the informed consent form, subjects completed the screening assessments within the screening period of up to 2 weeks. Glucose-lowering agents other than metformin and investigational medicinal products (except rescue medication given by the investigator) were prohibited, and metformin dose modification was prohibited throughout the study. Subjects entered the run-in period upon the completion of screening, during which all subjects received a placebo run-in, i.e., 0.25 mL of placebo was administered subcutaneously at each visit. At the end of the run-in period, upon re-verification, all subjects who met all the inclusion criteria and did not meet any exclusion criterion were randomized to each dose cohort and randomized in a 2:1 ratio at the cohort level to receive the corresponding dose of the investigational medicinal product added to metformin. The investigator was required to perform the specified sample collection and examination in accordance with the requirements specified in the protocol. Subjects were observed for an additional 16 weeks after a 16-week treatment period to collect safety data.

End Point Assessments

The primary efficacy endpoint was change from baseline in HbA1c at week 17 with the ISIS 449884 injection add-on to metformin versus placebo plus metformin. The secondary efficacy endpoints included the change from baseline in HbA1c at week 9 (100 mg cohort)/week 10 (50 mg or 60 mg cohorts), the changes from baseline in fasting plasma glucagon, fasting plasma active glucagon-like peptide 1 (GLP-1), fasting plasma total GLP-1, fasting glucose (serum), fasting insulin (serum), fasting serum C-peptide, and self-monitored blood glucose (SMBG) at week 17 and subsequent time points, the change from baseline in HbA1c at week 20, and the percentage of patients reaching the goal of HbA1c < 7%/6.5% at week 17 in the ISIS 449884 injection treatment groups versus the placebo group.

Safety

The investigator and the medical monitor from the sponsor monitored the safety of ISIS 449884 injection throughout the study. Safety assessments included any adverse events (AEs), laboratory tests, 12-lead electrocardiograms, vital signs, physical examinations, injection-site reactions and durations, hypoglycemia events, and concomitant medications that were administered during the clinical study.

Statistical Analysis

The primary analysis of the primary efficacy endpoint was performed in the full analysis set (FAS) population, and the change from baseline in HbA1c at week 17 was analyzed using an analysis of covariance (ANCOVA) model. The least-squares estimate of the change from baseline in HbA1c at week 17 with the 95% CI for each treatment group, the least-squares point estimate with the 95% CI for the difference between treatment groups at the same dose, and the least-squares point estimates with the 95% CIs and p values for the overall difference between treatment groups are presented (if the p value of the test for the difference between groups is ≤ 0.05, there is a significant difference between the two treatment groups in the primary efficacy endpoint). The analysis of the secondary efficacy endpoints in the FAS population was performed using similar analytical methods to those used for the primary efficacy endpoint. For the additional secondary efficacy endpoint (percentage of patients reaching the goal of HbA1c < 7%/6.5% at week 17 in the ISIS 449884 injection 50, 60, or 100 mg treatment group versus the placebo group), an estimate of the percentage of patients reaching the goal (n/N, where n is the number of subjects who met the response criterion and N is the total number of subjects in the overall analysis population) and the corresponding 95% CI are provided.

AEs were reported by system organ class (SOC) and preferred term (PT) by dose group using MedDRA V24.0 and graded by National Cancer Institute (NCI) common terminology criteria for adverse event (CTCAE) V5.0, and the number of AEs was summarized by subject. If at least one AE occurred in the same subject, it was calculated as once. The term, time to onset, end date, severity, and relationship to the investigational medicinal product of treatment-emergent adverse events (TEAEs) were summarized and analyzed. The frequency and percentage of TEAEs, investigational-medicinal-product-related TEAEs, CTCAE grade 3 or higher TEAEs, and treatment-emergent SAEs were statistically summarized by dose group.

The ratio of ISIS 449884 injection to placebo was 2:1, and the standard deviation of the change from baseline in HbA1c was estimated to be approximately 1.0%, the two-sided α was 0.05, and the power was 85% based on previous experience in the phase II clinical study of ISIS 449884 injection. Based on the results of previous Ionis studies, the difference in mean change from baseline in HbA1c was estimated to be 0.9% for the ISIS 449884 injection against placebo. It was estimated that the overall sample size was 45 subjects for ISIS 449884 injection and 23 for placebo, and considering that 25% of subjects may drop out during the course of the study, the total sample size was set at 90. There were 60 subjects in the ISIS 449884 injection group and 30 subjects in the placebo group.

Results

Patients

A total of 209 subjects were screened in this study, of whom 102 subjects did not enter the lead-in phase due to screening failure. Of the 107 subjects included in the lead-in phase of this study, a total of 90 subjects participated in randomization and received at least one dose of the investigational medicinal product; of these, 60 subjects were in the pooled ISIS 449884 treatment group (including 20 subjects each in the 50, 60, and 100 mg treatment groups) and 30 subjects were in the pooled placebo group (including 10 each in the 0.25-mL, 0.3-mL, and 0.5-mL placebo groups). Due to the COVID-19 pandemic, we considered that 25% of the subjects may drop out during the course of the study. However, in fact, a total of 10 patients dropped out, resulting in an overall actual dropout rate of 11.1% (10/90) in this study. The disposition of the subjects in this study is shown in Fig. 1.Fig. 1 Patient disposition in the study

A total of 88 of the 90 randomized subjects entered the full analysis set (FAS). Two subjects in the 0.5-mL placebo group in the 100-mg cohort were excluded due to the withdrawal of consent during the treatment period and a lack of postdose glycated hemoglobin records as a result of loss to follow-up, respectively. The baseline disease characteristics were generally comparable between the pooled ISIS 449884 treatment group and the pooled placebo group, as shown in Table 1. Table 1 Demographic and baseline characteristics of the subjects

	ISIS 449884 pooled treatment group
(N = 60)	Pooled placebo group
(N = 28)	
Age (years)	
 Mean (min, max)	57.4 (36, 73)	55.7 (31, 69)	
Age group, number (%)	
  < 60	30 (50.0)	16 (57.1)	
  ≥ 60	30 (50.0)	12 (42.9)	
Sex, number (%)	
 Male	28 (46.7)	17 (60.7)	
 Female	32 (53.3)	11 (39.3)	
Ethnicity, number (%)	
 East Asian	60 (100.0)	28 (100.0)	
 Others	0	0	
Body weight (kg)	67.7 (11.7)	69.8 (8.3)	
BMI (kg/m2)	25.4 (2.8)	25.7 (2.9)	
Duration of type 2 diabetes  mellitus (months)	56.3 (50.0)	51.4 (34.6)	
Disease duration group, number (%)	
 Less than 1 year	5 (8.3)	2 (7.1)	
 1–2 years	4 (6.7)	3 (10.7)	
 2–5 years	9 (15.0)	4 (14.3)	
 More than 5 years	10 (16.7)	8 (28.6)	
Baseline HbA1c (%)	8.7(0.75)	8.7 (0.82)	
HbA1c group, number (%)	
  < 8.5	25 (41.7)	12 (42.9)	
  ≥ 8.5	35 (58.3)	16 (57.1)	
Baseline fasting glucose (mmol/L)	9.7 (2.6)	9.8 (1.9)	
Baseline glomerular filtration rate (mL/min)	103.7 (14.7)	110.3 (17.5)	
Glomerular filtration rate group, number (%)	
 60–90	8 (13.3)	2 (7.1)	
 > 90	52 (86.7)	26 (92.9)	
Baseline systolic blood pressure (mmHg)	128.7 (15.5)	124.9 (13.2)	
Baseline diastolic blood pressure (mmHg)	80.4 (9.7)	79.7 (8.1)	
Lipid-lowering drugs, number (%)	
 Statins	3 (5.0)	1 (3.6)	
 Statins, fibrates	1 (1.7)	0	
 Fibrates	3 (5.0)	3 (10.7)	
Baseline was defined as the last non-missing value prior to the first dose of double-blind investigational medicinal product. Data are presented as n (%) or mean (SD) unless otherwise indicated. HbA1c glycated hemoglobin A1c

BMI body-mass index, LDL low-density lipoprotein, HDL high-density lipoprotein

Efficacy

There was a significant difference between the two groups in the primary efficacy endpoint (change from baseline in HbA1c at week 17). The mean ± SD of HbA1c at baseline was 8.7% ± 0.75% and 8.7% ± 0.82% in the pooled ISIS 449884 treatment group and the pooled placebo group, respectively. In the pooled ISIS 449884 treatment group, the mean ± SD of HbA1c at week 17 was 7.5% ± 0.96% (Fig. 2A); the corrected mean LS change from baseline was − 1.31% (95% CI − 1.66%, − 0.96%). In the pooled placebo group, the mean ± SD of HbA1c at week 17 was 9.1% ± 1.35%; the corrected LS mean change from baseline was 0.15% (95% CI − 0.37%, 0.66%). The LS mean difference in the pooled ISIS 449884 treatment group compared to the pooled placebo group was − 1.46% (95% CI − 1.92%, − 1.00%; P < 0.001). Compared with placebo plus metformin, the ISIS 449884 injection add-on to metformin significantly reduced HbA1c after 16 weeks of treatment (Fig. 2B).Fig. 2 The primary efficacy endpoint. The change from baseline in HbA1c level over time (A); each error bar indicates the SD. Least-squares mean change in HbA1c from baseline to week 9/10, week 17, and week 20 (B); each error bar indicates the 95% confidence interval (CI). Proportion of patients achieving the HbA1c targets; the values above the bars represent n (%) (C)

There was a significant reduction in the change from baseline in HbA1c at week 9 (100-mg cohort)/week 10 (50-mg and 60-mg cohorts). The mean ± SD of HbA1c at week 9/10 was 7.8% ± 0.93% in the pooled ISIS 449884 treatment group (Fig. 2A); the LS mean of the corrected change from baseline was − 1.04% (95% CI − 1.37%, − 0.71%). The mean ± SD of HbA1c at week 9/10 was 8.8% ± 1.32% in the pooled placebo group; the LS mean of the corrected change from baseline was − 0.19% (95% CI − 0.67%, 0.29%). The difference in LS mean between the pooled ISIS 449884 treatment group and the pooled placebo group was − 0.85% (95% CI − 1.28%, − 0.42%; P < 0.001) (Fig. 2B). Similarly, there was also a decrease in the change in HbA1c from baseline at week 20, which was the 4 weeks off-treatment. The mean ± SD HbA1c at week 20 was 7.6% ± 0.94% in the pooled ISIS 449884 treatment group; the LS mean of the corrected change from baseline was − 1.25% (95% CI − 1.62%, − 0.89%) (Fig. 2A). The mean ± SD of HbA1c at week 20 was 9.311% ± 1.4807% in the pooled placebo group; the LS mean of the corrected change from baseline was 0.39% (95% CI − 0.15%, 0.92%). The LS mean difference in the pooled ISIS 449884 treatment group compared to the pooled placebo group was − 1.64% (95% CI − 2.12%, − 1.16%; P < 0.001) (Fig. 2B).

As shown in Fig. 2A, HbA1c significantly decreased in the ISIS 449884 pooled treatment group after the start of treatment, and the level of HbA1c stabilized after week 9/10; the HbA1c in the pooled placebo group decreased slightly but remained essentially stable throughout the study period and remained significantly higher than that in the ISIS 449884 pooled treatment group at week 20. Overall, the mean HbA1c was lower in the ISIS 449884 treatment group than in the corresponding placebo group throughout the study. In addition, there was no significant difference in HbA1c between week 17 and week 20, which means the effect lasts for several weeks after the treatment.

The percentage of patients reaching the goal of HbA1c < 7%/6.5% at week 17 was higher in the ISIS 449884 injection 50-mg, 60-mg, and 100-mg treatment groups than in the corresponding placebo groups (Fig. 2C). The percentage of patients reaching the goal of HbA1c < 7%/6.5% at week 17 in the ISIS 449884 treatment groups was 25.0% (5/20 subjects)/5.0% (1/20 subjects), 50.0% (10/20 subjects)/15.0% (3/20 subjects), and 20.0% (4/20 subjects)/15.0% (3/20 subjects), respectively; the percentage of patients reaching the goal of HbA1c < 7%/6.5% at week 17 in the corresponding placebo groups was 0.

The changes in fasting plasma glucagon from baseline are shown in Fig. 3A. The mean fasting glucagon increased gradually over time in the pooled ISIS 449884 treatment group (n = 56), peaked at week 15/16, gradually decreased after the end of treatment, and remained above baseline at week 32. The mean fasting glucagon remained essentially stable throughout the study in the pooled placebo group (n = 23), with no significant change. ISIS 449884 significantly increased fasting glucagon levels compared to placebo at week 17 (LS difference, 14.52 pM (95% CI 7.75 pM, 21.28 pM), P < 0.001) (Fig. 3B).Fig. 3 Selected secondary efficacy endpoints. Time courses for the mean changes from baseline in fasting glucagon (A), total GLP-1 (C), fasting glucose (E), and fasting SMBG (G). Least-squares mean changes in fasting glucagon (B), GLP-1 (D), fasting glucose (F), and fasting SMBG (H) from baseline at week 17. Each error bar indicates the 95% CI

The changes in fasting plasma total GLP-1 are shown in Fig. 3C. After 16 weeks of the treatment add-on to metformin, the fasting plasma total GLP-1 increased in the pooled ISIS 449884 treatment group compared to the pooled placebo group. The mean fasting plasma total GLP-1 increased gradually over time in the pooled ISIS 449884 treatment group (n = 56), peaked at week 15/16, gradually decreased after the end of treatment, and remained above baseline at week 32. The mean fasting plasma total GLP-1 remained generally stable throughout the study in the pooled placebo group, with no significant change (n = 23). ISIS 449884 significantly increased the fasting plasma total GLP-1 compared to placebo at week 17 (LS difference, 23.58 pM (95% CI 13.63 pM, 33.54 pM), P < 0.001) (Fig. 3D).

As shown in Fig. 3E, the mean fasting serum glucose (n = 56) in the pooled ISIS 449884 treatment group showed an overall decreasing trend after the start of treatment and gradually increased after the end of treatment; the mean fasting serum glucose values in the pooled placebo group fluctuated after the start of treatment but were generally stable. The mean fasting serum glucose values measured during the treatment period in the pooled ISIS 449884 treatment group were lower than those in the pooled placebo group during the study (n = 28), with significant differences observed between the two groups. ISIS 449884 significantly decreased fasting serum glucose compared to placebo at week 17 (LS difference, − 0.81 mmol/L, (95% CI − 1.58 mmol/L, − 0.04 mmol/L), P = 0.039) (Fig. 3F).

For the change from baseline in self-monitored blood glucose (SMBG), during the treatment period, the reduction in fasting SMBG in the pooled ISIS 449884 treatment group was significantly better than that in the pooled placebo group, with significant differences observed between the groups (Fig. 3G). Compared with the pooled placebo group, the fasting SMBG decreased after 16 weeks of treatment in the pooled ISIS 449884 treatment group. After 16 weeks of the treatment add-on to metformin, the fasting SMBG decreased in the pooled ISIS 449884 treatment group compared to the pooled placebo group (LS difference, − 1.08 mmol/L, [95% CI − 1.66 mmol/L, − 0.50 mmol/L; P < 0.001) (Fig. 3H).

There were no significant differences in the changes from baseline in fasting active GLP-1, fasting serum insulin, and fasting serum C-peptide between the two groups during the study.

Although the blood lipids (low-density lipoprotein, cholesterol, and triglycerides), body weight, and blood pressure of the 90 participants in this study fluctuated throughout the study, there were no significant changes, and no significant differences were observed between the groups. Lipids neither increased nor decreased in the pooled ISIS 449884 treatment group compared to the pooled placebo group, and comparisons between the groups across cohorts gave similar results.

Safety

As shown in Table 2, TEAEs occurred in 53/60 subjects (88.3%) and 25/30 subjects (83.3%) in the pooled ISIS 449884 treatment group and pooled placebo group, respectively. Drug-related TEAEs occurred in 41/60 subjects (68.3%) and 9/30 subjects (30.0%), respectively. TEAEs of CTCAE grade 3 or higher were reported in 5/60 subjects (8.3%) and 2/30 subjects (6.7%), respectively, and none of them were drug related. In the pooled ISIS 449884 treatment group, serious TEAEs occurred in 4/60 subjects (6.7%), and TEAEs leading to drug discontinuation were reported in 3/60 subjects (5.0%). Only 1/20 subjects (5.0%) in the 100-mg group was confirmed to have a cardiovascular event as a TEAE. In the pooled ISIS 449884 treatment group, 7 of 60 subjects (11.7%) experienced alanine aminotransferase (ALT) ≥ 3 × ULN (where ULN is the upper limit of normal), and 6 of 60 (10.0%) subjects had AST ≥ 3 × ULN during the study. One of 20 subjects (5.0%) in the 50-mg treatment group had both ALT ≥ 5 × ULN and aspartate aminotransferase (AST) ≥ 5 × ULN, and the event was related to the study drug. After stopping the study-drug treatment, the outcome of this subject was resolution without sequelae. In addition, only 1 subject in the 50-mg group had TBIL (total bilirubin) ≥ 1.5 ×  ULN. Table 2 Summary of all TEAEs

Adverse event type, No (%)	ISIS 449884 pooled treatment group
(N = 60)	Placebo group
(N = 30)	
All TEAEs	53 (88.3)	25 (83.3)	
All study drug-related TEAEs	41 (68.3)	9 (30.0)	
Study-drug-related injection site TEAEs	13 (21.7)	0	
CTCAE grade 3 or higher TEAEs	5 (8.3)	2 (6.7)	
CTCAE grade 3 or higher study drug-related TEAEs	0	0	
Serious TEAEs (STEAEs)	4 (6.7)	0	
Study-drug-related STEAEs	1 (1.7)	0	
TEAEs leading to death	0	0	
Study-drug-related TEAEs leading to death	0	0	
TEAEs leading to study drug interruption	3 (5.0)	0	
TEAEs leading to reduced frequency of drug use	1 (1.7)	0	
TEAEs leading to drug discontinuation	3 (5.0)	0	
TEAEs associated with an injection reaction	13 (21.7)	0	
TEAEs for confirmed cardiovascular events	1 (1.7)	0	
Other AESIs	
 ALT	
  ≥ 3 ×  ULN	7 (11.7)	0	
  ≥ 5 ×  ULN	1 (1.7)	0	
 AST	
   ≥ 3 ×  ULN	6 (10.0)	0	
   ≥ 5 ×  ULN	1 (1.7)	0	
COVID-19 infections among the subjects	0	0	
Data are presented as no. (%). Confirmed cardiovascular events: non-fatal myocardial infarction, non-fatal stroke, and cardiovascular-related death

TEAEs treatment-emergent adverse events, ALT alanine aminotransferase, AST aspartate aminotransferase, AESI adverse event of special interest, ULN upper limit of normal

The changes in ALT and AST over time were analyzed in this study. As depicted in Fig. 4A, B and Table S1, the mean values of ALT and AST in the pooled ISIS 449884 treatment group were 20.48 U/L and 18.03 U/L at baseline. These values increased post-dose, peaked at week 16 (at 48.72 U/L and 39.42 U/L, respectively), gradually decreased after the end of treatment, and returned to baseline levels at week 32 (27.41 U/L and 23.78 U/L, respectively). In contrast, ALT and AST in the pooled placebo group remained at baseline levels throughout the study; they showed slight fluctuations but were generally stable. The levels of ALT and AST were 18.41 U/L and 17.59 U/L at baseline in the placebo group, and the levels were 19.99 U/L and 19.02 U/L at week 32.Fig. 4 Mean ALT levels over time (A), and mean AST levels over time (B). Each error bar indicates the 95% CI

Of note, 13/60 subjects (21.7%) in the pooled ISIS 449884 treatment group had infusion-reaction-related TEAEs, all of which were CTCAE grade 1 or 2, and no serious infusion-reaction-related TEAEs were observed. No injection-site reactions led to discontinuation. The incidence of hypoglycemia was higher in the pooled ISIS 449884 treatment group (10/60 subjects, 16.7%) than in the pooled placebo group (2/30 subjects, 6%). There were no serious hypoglycemia events during the study, and the drug did not increase the risk of serious hypoglycemia.

Discussion

This was a multicenter, placebo-controlled (2:1), randomized, double-blind, parallel-enrollment, multiple-dose phase II study to evaluate the efficacy and safety of the ISIS 449884 injection add-on to metformin in Chinese patients with T2DM who were uncontrolled by metformin monotherapy. In patients with T2DM, treatment with the ISIS 449884 injection add-on to metformin for 16 weeks notably improved HbA1c levels compared to placebo plus metformin while demonstrating an acceptable benefit/risk profile.

We also observed almost similar changes in HbA1c level at week 17 and week 20, which demonstrated that the reduction in HbA1c in the ISIS 449884 treatment group after treatment was stable and long in duration. The trend in HbA1c levels observed across different cohorts within the ISIS 449884 treatment group generally aligned with the trend observed in the overall pooled ISIS 449884 treatment group. While more patients in the medium-dose cohort than in the 100-mg dose cohort reached their target HbA1c, we think that the small sample size (20 patients in each cohort) and the big variability (1, 0, and 2 patients discontinued the treatment, respectively, in the 50-mg, 60-mg, and 100-mg cohorts) may have contributed to this bias.

Additionally, other glycemic control variables, such as the change from baseline in fasting serum glucose, confirmed the efficacy of subcutaneous injections of the ISIS 449884 add-on to metformin compared to placebo plus metformin over a 16-week period.

Furthermore, during the treatment period, ISIS 449884 exhibited a notable elevation in fasting glucagon levels. This observation aligns with the glucose-lowering mechanism of ISIS 449884, which functions by impeding the synthesis of GCGR protein and consequently inhibiting the binding of glucagon to its receptor. As a result, a glucose-lowering effect is achieved. Simultaneously, due to the blockade of the action of glucagon, the human body compensates by increasing the level of glucagon, which led to a substantial increase in glucagon within the treatment groups. This finding is consistent with previous scholarly studies [11].

The rises in GLP-1 were expected, given the mechanism of action of ISIS 449884. However, we found that elevated total GLP-1 concentrations, which paralleled those of glucagon, were most likely the consequence of alpha-cell hyperplasia. In this case, GLP-1 would have the sequence GLP-1 (1–37 amide) and not (7–36 amide), as if from the gut.

Reversible and dose-dependent increments in ALT and AST were observed when small-molecule antagonists targeting GCGR and the human glucagon receptor monoclonal antibody LY2786890 were used [12], and similarly, we witnessed similar changes in ALT/AST during ISIS 449884 treatment. However, these elevations in ALT and AST did not cause any increase in total bilirubin. Previous studies showed that these increases after treatment with an antisense inhibitor of GCGR—IONIS GCGRRx (also called ISIS 449884)—in the trial NCT02824003 [11] are accompanied by increases in hepatic fat content and hepatic lipids, though we did not measure the hepatic fat and lipids in the current study. In addition, when other glucagon receptor antagonists were used [13–17], increases in body weight, blood pressure, and blood lipid levels were observed; however, these changes were not observed with ISIS 449884 treatment in the current study.

In the world of diabetes management, the creation of a GCGR antagonist sparks continued interest. A multitude of antagonists have undergone evaluation in human trials, with each one revealing unique non-glycemic impacts [18]. Distinct outcomes can be obtained by using the diverse molecules and antibodies that interact with GCGR. These are due to drug-specific and mechanism-based variations in specific bindings such as the binding affinities to GCGR and other related receptors, especially the analogously structured GLP receptors within the GCGR subcategory. Due to their distinct binding specificities, differential effects can be observed among small molecules and antibodies that bind to GCGR along with other G-protein-coupled receptors, notably the closely related GLP receptors within the GCGR subfamily [19]. The inhibitory action of ISIS 449884 on GCGR is highly specific, with its efficacy limited to hepatic cells; it has no impact on structurally similar proteins [11]. Importantly, even at high doses of glucagon, ISIS 449884 does not induce a plateau effect despite the partial inhibition of GCGR [11]. A partial attenuation of GCGR expression is suggested by these findings. In contrast to GCGR knockout mice [20], administration of ISIS 449884 does not induce a compensatory rise in sympathetic activity, thereby avoiding a marked enhancement blood pressure. In conclusion, these findings indicate that ISIS 449884 could potentially serve as a promising therapeutic agent for patients suffering from conditions associated with GCGR deregulation.

The study has certain limitations, including a small sample size, the absence of hepatic lipid values, and the absence of data on hepatic fat content. In addition, other glycemic targets such as continuous glucose monitors (CGMs) which may show the postprandial glucose response were not measured, and an oral glucose/mix test was not performed to evaluate the GLP-1 and glycemic responses, which would add value given that GLP-1 is a gut hormone secreted after meal ingestion. Besides, insulin sensitivity, which can be impacted by the blocking of glucagon receptors, was not measured. These factors may be considered in a future trial. Despite some limitations, the results align with the positive impacts on blood sugar levels and heightened ALT levels observed with other drugs that target GCGR [11, 21–23]. These findings underscore the potential benefits of such agents in managing blood sugar control. Also, other combinations with ISIS 449884 (other than the combination with metformin) could be better and should be explored.

Conclusion

In summary, this study provided clinical data on the application of ISIS 449884 injection in the T2DM population. It indicated that ISIS 449884 injection shows potential in the treatment of T2DM and has good clinical application prospects, although these need to be validated by further clinical studies with larger sample sizes.

Supplementary Information

Below is the link to the electronic supplementary material.Supplementary file1 (PDF 70 KB)

Medical Writing/Editorial Assistance

No external medical writing or editorial assistance was received during the writing of this article.

Author Contributions

Linong Ji and Leili Gao were the investigators who contributed to the conception and study design, recruited patients, and collected data. The first draft of the manuscript was written by Guoliang Chen. Linong Ji, Leili Gao, Zhikai Feng, Guoliang Chen, Jing Fu, Erin Morgan, Sanjay Bhanot, Shan Gao, Hongyan Zhang, Zicai Liang, and Li-Ming Gan participated in the interpretation of data and the critical revision of the manuscript, and they approved the final version. All authors agreed to be accountable for all aspects of the work and to ensure that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Funding

This study and the journal’s rapid service fee was sponsored by Suzhou Ribo Life Science Co. Ltd.

Data Availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Conflict of Interest

Zhikai Feng, Guoliang Chen, Jing Fu, Shan Gao, Hongyan Zhang, Zicai Liang, and Li-Ming Gan are employees of Suzhou Ribo Life Science Co. Ltd. Erin Morgan and Sanjay Bhanot are employees of Ionis Pharmaceuticals, Inc. (the original company name was ISIS Pharmaceuticals, Inc.). Linong Ji reports receiving consulting and lecture fees from Eli Lilly and Company, Novo Nordisk, Merck, Bayer, Sanofi–Aventis, Roche, Merck Sharp & Dohme, Metronics, AstraZeneca, Boehinger Ingelheim, and Abbott. Leili Gao has nothing to disclose.

Ethical Approval

The trial was conducted in accordance with the 2013 Declaration of Helsinki and the International Conference on Harmonization’s Good Clinical Practice guidelines, and it was approved by the institutional review board of Peking University People's Hospital (the main center; approval number: 2019PHA018-001) and the relevant institutional review board at each study site. A list of the IECs/IRBs that reviewed and approved the protocol are provided in Supplemental Table S2. Informed consent was obtained from all participants prior to enrollment in the study.

Linong Ji and Leili Gao contributed equally to this study.

Change history

8/29/2024

A Correction to this paper has been published: 10.1007/s13300-024-01633-3
==== Refs
References

1. IDF. Diabetes atlas, 10th edition, 2021. https://diabetesatlas.org/en/. Accessed May 2024.
2. Ma RCW Epidemiology of diabetes and diabetic complications in China Diabetologia 2018 61 6 1249 1260 10.1007/s00125-018-4557-7 29392352
Ma RCW. Epidemiology of diabetes and diabetic complications in China. Diabetologia. 2018;61(6):1249–60.29392352
3. Galicia-Garcia U Benito-Vicente A Jebari S Pathophysiology of type 2 diabetes mellitus Int J Mol Sci 2020 21 17 6275 10.3390/ijms21176275 32872570
Galicia-Garcia U, Benito-Vicente A, Jebari S, et al. Pathophysiology of type 2 diabetes mellitus. Int J Mol Sci. 2020;21(17):6275.32872570
4. The Action to Control Cardiovascular Risk in Diabetes Study Group Effects of intensive glucose lowering in type 2 diabetes N Engl J Med 2008 358 24 2545 2559 10.1056/NEJMoa0802743 18539917
The Action to Control Cardiovascular Risk in Diabetes Study Group. Effects of intensive glucose lowering in type 2 diabetes. N Engl J Med. 2008;358(24):2545–59.18539917
5. Duckworth W Abraira C Moritz T Glucose control and vascular complications in veterans with type 2 diabetes N Engl J Med 2009 360 2 129 139 10.1056/NEJMoa0808431 19092145
Duckworth W, Abraira C, Moritz T, et al. Glucose control and vascular complications in veterans with type 2 diabetes. N Engl J Med. 2009;360(2):129–39.19092145
6. The ADVANCE Collaborative Group Intensive blood glucose control and vascular outcomes in patients with type 2 diabetes N Engl J Med 2008 358 24 2560 2572 10.1056/NEJMoa0802987 18539916
The ADVANCE Collaborative Group. Intensive blood glucose control and vascular outcomes in patients with type 2 diabetes. N Engl J Med. 2008;358(24):2560–72.18539916
7. Marathe PH, Gao HX, Close KL. American Diabetes Association standards of medical care in diabetes 2017. J Diabetes. 2017;9(4):320–4.
8. Wang L Peng W Zhao Z Prevalence and treatment of diabetes in China, 2013–2018 JAMA 2021 326 24 2498 2506 10.1001/jama.2021.22208 34962526
Wang L, Peng W, Zhao Z, et al. Prevalence and treatment of diabetes in China, 2013–2018. JAMA. 2021;326(24):2498–506.34962526
9. Defronzo RA Banting Lecture. From the triumvirate to the ominous octet: a new paradigm for the treatment of type 2 diabetes mellitus Diabetes 2009 58 4 773 795 10.2337/db09-9028 19336687
Defronzo RA. Banting Lecture. From the triumvirate to the ominous octet: a new paradigm for the treatment of type 2 diabetes mellitus. Diabetes. 2009;58(4):773–95.19336687
10. Petersen KF Sullivan JT Effects of a novel glucagon receptor antagonist (Bay 27–9955) on glucagon-stimulated glucose production in humans Diabetologia 2001 44 11 2018 2024 10.1007/s001250100006 11719833
Petersen KF, Sullivan JT. Effects of a novel glucagon receptor antagonist (Bay 27–9955) on glucagon-stimulated glucose production in humans. Diabetologia. 2001;44(11):2018–24.11719833
11. Morgan ES Tai LJ Pham NC Overman JK Watts LM Smith A Antisense inhibition of glucagon receptor by IONIS-GCGR(Rx) improves type 2 diabetes without increase in hepatic glycogen content in patients with type 2 diabetes on stable metformin therapy Diabetes Care 2019 42 4 585 593 10.2337/dc18-1343 30765435
Morgan ES, Tai LJ, Pham NC, Overman JK, Watts LM, Smith A, et al. Antisense inhibition of glucagon receptor by IONIS-GCGR(Rx) improves type 2 diabetes without increase in hepatic glycogen content in patients with type 2 diabetes on stable metformin therapy. Diabetes Care. 2019;42(4):585–93.30765435
12. Kelly RP, Garhyan P, Reynolds VL, et al. Glucagon receptor antibody LY2786890 reduced glucose levels in type 2 diabetes mellitus patients (abstract 106-LB). Diabetes. 2015;64(Suppl. 1A):LB27.
13. Guzman CB Zhang XM Liu R Regev A Shankar S Garhyan P Treatment with LY2409021, a glucagon receptor antagonist, increases liver fat in patients with type 2 diabetes Diabetes Obes Metab 2017 19 11 1521 1528 10.1111/dom.12958 28371155
Guzman CB, Zhang XM, Liu R, Regev A, Shankar S, Garhyan P, et al. Treatment with LY2409021, a glucagon receptor antagonist, increases liver fat in patients with type 2 diabetes. Diabetes Obes Metab. 2017;19(11):1521–8.28371155
14. Engel SS, Xu L, Andryuk PJ, et al. Efficacy and tolerability of MK-0893, a glucagon receptor antagonist (GRA), in patients with type 2 diabetes (T2DM) (abstract 309-OR). Diabetes. 2011;60(Suppl. 1):A85.
15. Ruddy M Pramanik B Lunceford J Inhibition of glucagon-induced hyperglycemia predicts glucose lowering efficacy of a glucagon receptor antagonist, MK-0893, in type 2 diabetes (T2DM) (Abstract 311-OR) Diabetes 2011 60 Suppl. 1 A85 A86
Ruddy M, Pramanik B, Lunceford J, et al. Inhibition of glucagon-induced hyperglycemia predicts glucose lowering efficacy of a glucagon receptor antagonist, MK-0893, in type 2 diabetes (T2DM) (abstract 311-OR). Diabetes. 2011;60(Suppl. 1):A85–6.
16. Engel SS, Teng R, Edwards RJ, et al. Efficacy and safety of the glucagon receptor antagonist, MK-0893, in combination with metformin or sitagliptin in patients with type 2 diabetes mellitus (abstract 191). In: 47th Annual Meeting of the EASD; 2011 Sept 13–16; Lisbon, Portugal.
17. Vajda EG, Logan D, Lasseter K, et al. Pharmacokinetics and pharmacodynamics of the glucagon receptor antagonist LGD-6972 in a multi-dose clinical trial (abstract 1193-P). Diabetes. 2015;64(Suppl. 1):A308.
18. Nunez DJ D'Alessio D Glucagon receptor as a drug target: a witches' brew of eye of newt (peptides) and toe of frog (receptors) Diabetes Obes Metab 2018 20 2 233 237 10.1111/dom.13102 28842950
Nunez DJ, D’Alessio D. Glucagon receptor as a drug target: a witches’ brew of eye of newt (peptides) and toe of frog (receptors). Diabetes Obes Metab. 2018;20(2):233–7.28842950
19. Sa Z Zhou J Zou Y Su Z Gu X Paralog-divergent features may help reduce off-target effects of drugs: hints from glucagon subfamily analysis Genomics Proteomics Bioinform 2017 15 4 246 254 10.1016/j.gpb.2017.03.004
Sa Z, Zhou J, Zou Y, Su Z, Gu X. Paralog-divergent features may help reduce off-target effects of drugs: hints from glucagon subfamily analysis. Genomics Proteomics Bioinform. 2017;15(4):246–54.
20. Gelling RW Du XQ Dichmann DS Romer J Huang H Cui L Lower blood glucose, hyperglucagonemia, and pancreatic alpha cell hyperplasia in glucagon receptor knockout mice Proc Natl Acad Sci USA 2003 100 3 1438 1443 10.1073/pnas.0237106100 12552113
Gelling RW, Du XQ, Dichmann DS, Romer J, Huang H, Cui L, et al. Lower blood glucose, hyperglucagonemia, and pancreatic alpha cell hyperplasia in glucagon receptor knockout mice. Proc Natl Acad Sci USA. 2003;100(3):1438–43.12552113
21. Kazda CM Ding Y Kelly RP Garhyan P Shi C Lim CN Evaluation of efficacy and safety of the glucagon receptor antagonist LY2409021 in patients with type 2 diabetes: 12- and 24-week phase 2 studies Diabetes Care 2016 39 7 1241 1249 10.2337/dc15-1643 26681715
Kazda CM, Ding Y, Kelly RP, Garhyan P, Shi C, Lim CN, et al. Evaluation of efficacy and safety of the glucagon receptor antagonist LY2409021 in patients with type 2 diabetes: 12- and 24-week phase 2 studies. Diabetes Care. 2016;39(7):1241–9.26681715
22. Kazierad DJ Bergman A Tan B Erion DM Somayaji V Lee DS Effects of multiple ascending doses of the glucagon receptor antagonist PF-06291874 in patients with type 2 diabetes mellitus Diabetes Obes Metab 2016 18 8 795 802 10.1111/dom.12672 27059951
Kazierad DJ, Bergman A, Tan B, Erion DM, Somayaji V, Lee DS, et al. Effects of multiple ascending doses of the glucagon receptor antagonist PF-06291874 in patients with type 2 diabetes mellitus. Diabetes Obes Metab. 2016;18(8):795–802.27059951
23. Kostic A King TA Yang F Chan KC Yancopoulos GD Gromada J A first-in-human pharmacodynamic and pharmacokinetic study of a fully human anti-glucagon receptor monoclonal antibody in normal healthy volunteers Diabetes Obes Metab 2018 20 2 283 291 10.1111/dom.13075 28755409
Kostic A, King TA, Yang F, Chan KC, Yancopoulos GD, Gromada J, et al. A first-in-human pharmacodynamic and pharmacokinetic study of a fully human anti-glucagon receptor monoclonal antibody in normal healthy volunteers. Diabetes Obes Metab. 2018;20(2):283–91.28755409
