Submucosal Injection of the RNA Oligonucleotide GUT-1 in Active Ulcerative Colitis Patients: A Randomized, Double-Blind, Placebo-Controlled Phase 2a Induction Trial Free

Abstract

Background and Aims

Carbohydrate sulfotransferase 15 [CHST15] biosynthesizes sulphated matrix glycosaminoglycans and is implicated in intestinal inflammation and fibrosis. Here, we evaluate the efficacy and safety of the double-stranded RNA oligonucleotide GUT-1, a specific blocker of CHST15, as induction therapy in patients with ulcerative colitis [UC].

Methods

In this randomized, double-blind, placebo-controlled, phase 2a study, we enrolled endoscopically active UC patients, refractory to conventional therapy, in five hospital centres across Germany. Patients were randomized 1:1:1 using a block randomized technique to receive a single dosing of 25 nM GUT-1, 250 nM GUT-1, or placebo by endoscopic submucosal injections. The primary outcome measure was improvement of endoscopic lesions at weeks 2 or 4. The secondary outcome measures included clinical and histological responses. Safety was assessed in all patients who received treatment.

Results

Twenty-eight patients were screened, 24 were randomized, and 21 were evaluated. Endoscopic improvement at weeks 2 or 4 was achieved by 71.4% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 28.6% in the placebo group. Clinical remission was shown by 57.1% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 14.3% in the placebo groups. Histological improvement was shown by 42.9% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 0% in the placebo groups. GUT-1 250 nM reduced CHST15 expression significantly and suppressed mucosal inflammation and fibrosis. GUT-1 application was well tolerated.

Conclusion

Single dosing by submucosal injection of GUT-1 repressed CHST15 mucosal expression and may represent a novel induction therapy by modulating tissue remodelling in UC.

1. Introduction

Ulcerative colitis [UC] can be characterized by progressive structural bowel damage due to persistent mucosal ulcerations and fibrosis, which are major manifestations of tissue remodelling leading to the formation of stenosis, and therapeutic failure of treatment options.^1,2

Integrated anti-tissue remodelling strategies that simultaneously target mediators of inflammation, profibrotic factors, and tissue intrinsic changes are needed to enable a rational therapeutic approach to treat this highly complex intestinal tissue transformation.^3–5

Extracellular matrix [ECM] molecules are considered as possible therapeutic targets for tissue remodelling, as they modulate the function of multiple mediators of tissue injury.⁶

Carbohydrate sulfotransferase 15 [CHST15] is a type II transmembrane Golgi protein that biosynthesizes highly sulphated disaccharide units [E-units] of chondroitin sulphate [CS], which binds to various functional proteins and pathogenic microorganisms.^7,8

CHST15-derived CS-E is reported to be centrally involved in the onset of fibrosis through activation of fibroblasts and formation of collagen fibrils.^7–9 CS-E acts as the co-receptor for Wnt ligands and as the direct receptor for receptor for advanced glycation end products [RAGE] on fibroblasts, both of which are responsible for activation of the fibrosis signalling pathway.^8,9 In addition, CS-E enhances fibril formation of collagen and of the extracellular molecule amyloid β [Aβ] peptide, which was shown to inhibit degradation of established fibrils,^10,11 contributing to the inhibition of fibrinolytic properties. Matrix CS-E thus accelerates the vicious cycle of fibrogenesis and strengthens fibrotic formations which represent an insuperable obstacle for tissue repair. However, several experimental models have shown that blockade of CHST15 prevented or repressed fibrosis in oesophageal strictures, cardiac and pulmonary fibrosis, experimental colitis, as well as in a Phase 1 trial in Crohn’s disease patients.^9,12–17

GUT-1 is a synthetic double-stranded RNA oligonucleotide that selectively inhibits the expression of CHST15.^15–17 Previous in vitro studies demonstrated that GUT-1 suppresses CHST15 mRNA in a dose-dependent manner as well as the mRNA expression of vimentin, α-SMA and Wnt3 in a TGF-β-stimulated human fibroblast cell line.⁹ Here, in a Phase 2a randomized, double-blind study, we investigated the efficacy, safety, and mode of action related to tissue remodelling of GUT-1 induction treatment in active UC patients.

2. Materials and Methods

2.1. Study design and participants

This was a Phase 2a, randomized, multi-centre, double-blind, placebo-controlled, parallel-group study which included adult patients [age 18–65 years] with moderate-to-severe UC, defined by a modified Mayo Score [mMS] of 5–9 and a Mayo endoscopic subscore [MES] of ≥2 [see Table 1].^18–20 The study was conducted in five hospital centres across Germany.

Patients had to be previously treated by conventional therapies and experienced an insufficient response. During the trial, patients could be taking 5-aminosalicylic acids, oral corticosteroids, or immunosuppressives in unchanged doses [14 days of unchanged doses prior to baseline]. Patients were excluded if they had a diagnosis of proctitis or pancolitis [local reading], a history of UC-related surgery, complications of UC, anal stenosis or perianal abscess with fever, concomitant local therapy, or a biological agent [wash-out period of 4 weeks]. Women who were pregnant or breast-feeding were excluded. The study protocol was reviewed and approved by the regional Independent Ethics Committees and by the competent German authority [Federal Institute for Drugs and Medical Devices]. This study is registered with EUCTR, EudraCT Number 2012-004768-23. Written informed consent was provided by all patients.

2.2. Procedures

Patients were randomized 1:1:1 using a block randomized technique, to receive a single dosing of 25 or 250 nM GUT-1 [synthesized by BioSpring], or placebo by endoscopic submucosal injections. These doses were determined by our previous results from a Phase 1 clinical trial for Crohn’s disease showing a reduction of CHST15 mRNA from baseline in the colon tissues of patients and from mouse colitis models which showed that silencing efficacy reached almost saturation at 250 nM.^9,16,17 The study medication [a water solution in a glass vial] and placebo had identical appearance, viscosity, smell, and labelling. Patients and investigators [including personnel administering the interventions, assessing outcome measures, and analysing data] were blinded to treatment assignment. The primary outcome measure was improvement of endoscopic lesions at weeks 2 or 4 [per evaluable population]. Eligible participants were admitted to the study site and GUT-1 was then injected submucosally [2 mL/site by one injection and two opposite injections per site at distances of 35, 25, 15, and 5 cm from the anal verge, thus eight injections] during endoscopy.¹⁷ Participants were discharged 24 h after administration. Patients underwent subsequent endoscopic examination at weeks 2 and 4. During all endoscopies [baseline and follow-up], three images indicative for the level of highest mucosal inflammation were taken for central reading, by experienced endoscopists blinded to all study data. No videorecording was taken for this early-stage study. However, as endoscopic scoring was done by focusing on the heaviest lesion; one image was taken from the area of the heaviest inflammation in the rectum, the sigmoid, and one additional from either the rectum or sigmoid in order for the central reader to be able to sufficiently assess the endoscopic disease activity of left-sided colitis. Biopsies were taken from the most inflamed mucosal area during all endoscopic examinations.

2.3. Outcome measures

The primary endpoint was improvement of endoscopic lesions by MES. Thus, changes from baseline of MES at week 2 or week 4 were evaluated. Endoscopic improvement was defined as a centrally read MES of 0 or 1^18–20 at week 2 or week 4. The endoscopic evaluation was done by central reading of the images as described before, in a blinded fashion. Secondary endpoints for efficacy evaluated the effects of GUT-1 on clinical and histological responses. Clinical response was evaluated based on the mMS. Clinical remission was defined as an mMS score of 0–2, including the following three components; [1] stool frequency subscore [SFS] of 0 or 1; [2] rectal bleeding subscore [RBS] of 0; and [3] centrally read endoscopy subscore [MES] of 0 or 1.^18–20 The percentage changes in mMS, SFS, RBS, and MES from baseline to week 2 or 4 were also evaluated, separately. Histological response was evaluated based on the Geboes Index.²¹ Histological improvement was defined as the composite of neutrophil infiltration in less than 5% of crypts [Grade 3 Geboes Index of 0 or 1] and no crypt destruction, erosions, ulcerations, or granulation tissue [Grade 4 Geboes Index of 0 and Grade 5 Geboes Index of 0]^21–23 at week 2 or 4. In addition, histological response was also evaluated by changes in the Nancy Histological Index [NHI]^23,24 from baseline to week 2 or 4. Histological assessment was performed by three independent pathologists, blinded to all data. In the case of a discrepancy between pathologists, the worst histological score assessed was taken. Qualitative histological analysis was performed using HE- and Masson Trichrome-stained sections.

Plasma drug concentrations 2 h after drug administration were measured by ELISA.^9,16 Intestinal biopsies were stained via immunohistochemistry for CHST15, as described elsewhere.²⁵ For quantitative analysis, bright-field images of CHST15-stained sections were captured using a digital camera at 200-fold magnification. To estimate the intensity of CHST15 staining in the cytoplasm, the positive areas, but not the number of positive cells, in four fields/section were measured using ImageJ software [National Institute of Health]. The mean of the percentage positive areas [from four fields/section] was reported.

Safety and tolerability were evaluated throughout the study. Safety was assessed by vital signs, 12-lead ECG, laboratory values, and incidence and type of treatment emergent adverse events [TEAEs] of all treated participants. The incidence and the type of TEAEs were summarized by treatment groups.

2.4. Statistical analysis

Sample size planning was based on the previous observation of a marked difference in endoscopic improvement between active treatment and placebo of 89% vs 11% in a previous study in Crohn’s disease patients using the same therapeutic regimen. Targeting seven patients per group and assuming a similar efficacy, thus a 6/7 vs 1/7 success rate, a Cohen’s effect size of 1.59 was determined. With a two-sided alpha of 0.05, we estimated the power to be 84.5% [‘pwr’ R package] for the comparison of the high-dose group and placebo regarded as the primary endpoint. Considering a drop-out rate of one patient per group as a realistic estimate, eight patients per group had to be recruited [eight for GUT-1 250 nM, eight for the low-dose group GUT-1 25 nM ,and eight for placebo], i.e. 24 participants in total. Missing values were tabulated but not be included in the calculation of percentages, and thereby were assumed to be missing completely at random.

Demographic and other baseline characteristics were summarized by tabulating frequencies or providing descriptive statistics stratified for each treatment group. The number of participants in whom TEAEs occurred and the number of events were summarized by treatment group. The safety population included all participants who received the study drug. A per-protocol population was defined including all participants who received the study drug, the pre-treatment drug, and without any major protocol deviation. Efficacy was assessed by evaluable population.

Descriptive statistical analysis was performed using GraphPad Prism v.9 and the R software package. Exact confidence intervals for proportions were calculated according to the method of Clopper-Pearson; in case of zero counts, the upper confidence limit was calculated with an adapted method.²⁶ For MES and NHI, the changes from baseline were calculated in the evaluable population [n = 21]. For mMS and its three components, MES, SFS, and RBS, percentage changes from baseline were calculated. The proportions of participants with endoscopic improvement, clinical remission, and histological improvement were calculated. The main focus was on complete descriptive statistics.

2.5. Role of the funding source

The funder of the study, Stelic Institute & Co., had a role in study design, data collection, data analysis, data interpretation, and writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit.

3. Results

3.1. Patient population and baseline characteristics

A total of 28 patients were screened from December 2013 to March 2016, in five hospital centres in Germany. The eligible 24 patients were randomized to receive a single dosing of 250 or 25 nM GUT-1, or placebo by endoscopic submucosal injections. All 24 patients completed the study. All patients were included in the safety analysis population. One patient who received placebo was excluded from the per-protocol population, as the patient was accidentally taking vedolizumab without sufficient wash-out time. Two patients who received GUT-1 25 nM [n = 1] or 250 nM [n = 1] and showed baseline endoscopic subscores of 1 were excluded from the clinical evaluation. Thus, 21 patients were included in the efficacy analysis as the evaluable population [Figure 1].

Patients in the three study arms were similar regarding demographic data and concomitant medication, although a tendency towards longer disease duration could be observed in the GUT-1 250 nM group. There were at least 50% anti-tumour necrosis factor [anti-TNF]-exposed patients in each group. In all patients who had received prior anti-TNF therapy, treatments had to be stopped due to anti-TNF refractory disease. All patients who received vedolizumab had prior anti-TNF refractory treatment and were demonstrated to be refractory to subsequent vedolizumab therapy as well [Table 1].

3.2. Efficacy evaluation in centrally read endoscopy

The mean baseline endoscopic subscores in the evaluable population [n = 7 each] were 2.5 (95% confidence interval [CI]: 1.9–2.9) in the GUT-1 250 nM, 2.6 [95% CI: 2.1–3.1] in the GUT-1 25 nM, and 2.5 [95% CI: 1.9–2.9] in the placebo groups. The primary endpoint, endoscopic improvement, was evaluated by centrally read MES. Endoscopic improvement at the end of this induction study [at week 2 or week 4] was achieved by 71.4% [95% CI: 29.0–96.3%; n = 5/7], 0.0% [95% CI: 0–34.8%; n = 0/7], and 28.6% [95% CI: 3.4–71.0%; n = 2/7] in the GUT-1 250 nM, GUT-1 25 nM, and placebo groups, respectively [Figure 2A].

The mean endoscopic subscores at week 2 [n = 7/each] were 1.6 [95% CI: 0.7–2.5] in the GUT-1 250 nM, 2.3 [95% CI: 1.8–2.7] in the GUT-1 25 nM, and 2.3 [95% CI: 1.6–3.0] in the placebo groups. At week 4, the mean endoscopic subscores were 1.2 [n = 6, 95% CI: 0.4–2.0] in the GUT-1 250 nM, 2.3 [n = 4, 95% CI: 1.5–3.0] in the GUT-1 25 nM, and 1.8 [n = 5, 95% CI: 0.8–2.8] in the placebo groups. The mean changes at the end of this induction study from baseline were −1.43 [95% CI: −2.16 to −0.70] in the GUT-1 250 nM, −0.29 [95% CI: −0.74 to 0.17] in the GUT-1 25 nM, and −0.43 [95% CI: −0.92 to 0.07] in the placebo groups [Figure 2B].

3.3. Efficacy evaluation in mMS

Clinical remission at week 2 or week 4 was shown in 57.1% [95% CI: 18.4–90.1%; n = 4/7], 0.0% [95% CI: 0–34.8%; n = 0/7], and 14.3% [95% CI: 0.4–57.9%; n = 1/7] in the GUT-1 250 nM, GUT-1 25 nM, and the placebo group, respectively [Figure 3A].

Percentage changes from baseline of the mMS and its three components, centrally read MES, SFS, and RBS, were also evaluated [Figure 3B]. The mean percentage change of mMS at week 2 from baseline was −50.6 % [95% CI: −83.9 to −17.3] in the GUT-1 250 nM, −1.6% [95% CI: −21.3 to 18.1] in the GUT-1 25 nM, and −18.1 % [95% CI: −39.5 to 3.3] in the placebo group. The mean percentage change of mMS at week 4 from baseline was −55.9 % [95% CI: −99.0 to −12.8] in the GUT-1 250 nM, −14.2% [95% CI: −57.1 to 28.8] in the GUT-1 25 nM, and −38.6% [95% CI: −80.8 to 3.6] in the placebo groups [Figure 3B].

Regarding mean percentage changes from baseline of the MES, the GUT-1 250 nM group showed −38.1% [95% CI: −69.8 to −6.4] at week 2 and −52.8% [95% CI: −88.5 to −17.1] at week 4. The GUT-1 25 nM group values were −9.5% [95% CI: −24.6 to 5.5] at week 2 and −16.7% [95% CI: −47.3 to 14.0] at week 4. The placebo group showed −4.8% [95% CI: −33.9 to 24.4] at week 2 and −26.7% [95% CI: −58.1 to 4.7] at week 4 [Figure 3B].

Concerning the mean percentage change of the SFS, the GUT-1 250 nM group showed −59.4% [95% CI: −97.1 to −21.8] at week 2 and −58.3% [95% CI: −109.9 to −6.7] at week 4. The GUT-1 25 nM group showed −16.7% [95% CI: –24.0 to 57.4] at week 2 and 0.0% [95% CI: –65.0 to 65.0] at week 4. The placebo group showed –21.4% [95% CI: –47.8 to 4.9] at week 2 and –46.6% [95% CI: −91.5 to −1.7] at week 4 [Figure 3B].

Regarding the mean percentage change of the RBS, the GUT-1 250 nM group showed −57.1% [95% CI: −106.6 to −7.7] at week 2 and −58.3% [95% CI: −109.9 to −6.7] at week 4. The GUT-1 25 nM group showed 0.0% [95% CI: not calculable] at week 2 and −22.2% [95% CI: −117.8 to 73.4] at week 4. The placebo group showed −33.3% [95% CI: −84.4 to 17.8] at week 2 and −46.7% [95% CI: −123.3 to −29.9] at week 4 [Figure 3B].

3.4. Efficacy evaluation in histology

The Geboes score was investigated to evaluate histological improvement.²¹ Histological improvement at the end of our study [at week 2 or week 4] was shown in 42.9% [95% CI: 9.9–81.6%; n = 3/7], 0.0% [95% CI: 0–34.8%; n = 0/7], and 0.0% [95% CI: 0–34.8%; n = 0/7] in the GUT-1 250 nM, GUT-1 25 nM, and placebo groups, respectively [Figure 4A]. Evaluation of the NHI showed that the mean baseline value in the evaluable population [n = 7/each] was 3.6 [95% CI: 2.8–4.3] in the GUT-1 250 nM, 3.9 [95% CI: 3.5–4.2] in the GUT-1 25 nM, and 3.6 [95% CI: 3.5–4.2] in the placebo groups. The mean changes at the end of the induction study [at week 2 or week 4] from baseline were −1.71 [95% CI: −2.60 to −0.83] in the GUT-1 250 nM, −0.86 [95% CI: −1.85 to 0.13] in the GUT-1 25 nM and −0.43 [95% CI: −0.92 to 0.07] in the placebo group [Figure 4B].

3.5. Mode of action: Expression of CHST15

We then performed immunohistochemical analyses to investigate changes of CHST15 expression [Figure 5]. The mean percentage positive areas of CHST15 in the GUT-1 250 nM group were 19.7% [95% CI: 17.0–22.5%] at baseline [n = 7], 11.5% [95% CI: 6.5–16.5%] at week 2 [n = 7], and 8.4% [95% CI: 3.1–13.7%] at week 4 [n = 6]. Those in the GUT-1 25 nM group were 19.6% [95% CI: 15.2–24.1%] at baseline [n = 7], 14.7% [95% CI: 10.6–18.7%] at week 2 [n = 7], and 14.0% [95% CI: 7.7–20.2%] at week 4 [n = 4]. Those in the placebo group were 16.3% [95% CI: 14.6–18.0%] at baseline [n = 7], 18.8% [95% CI: 16.1–21.4%] at week 2 [n = 7], and 18.8% [95% CI: 15.3–22.2%] at week 4 [n = 5].

3.6. Safety

GUT-1 25 or 250 nM administered submucosally was generally well tolerated. The proportion of participants who experienced at least one TEAE was as follows: GUT-1 25 nM 50.0% [95% CI: 15.7–84.3%; n = 4/8], 250 nM 62.5% [95% CI: 24.5–91.5%; n = 5/8], and placebo 50.0% [95% CI: 15.7–84.3%; n = 4/8]; of note, the safety population for placebo comprises all eight recruited participants. The severity was mild to moderate in all AEs. Drug-related TEAEs were shown in the GUT-1 25 nM and placebo groups, but not in the GUT-1 250 nM group [Table 2]. The plasma concentration of GUT-1 2 h after injection was below the lower limit of quantification [LLOQ 0.5 ng/mL] in all treatment groups [Supplementary material].

4. Discussion

In this Phase 2a induction trial, we found that CHST15 was highly expressed in the inflamed colon at baseline in UC patients who did not respond sufficiently to prior conventional treatment including biologics and had active endoscopic lesions with respect to an MES of ≥2. Single dosing of 250 nM GUT-1, an oligonucleotide that specifically suppresses CHST15, revealed 71.4% achieved endoscopic improvement at the end of this induction trial [at week 2 or week 4]. A total of 57.1% achieved clinical remission with rapid reduction of the MES and the SFS at week 2 with 250 nM GUT-1. Exploratory effects of GUT-1 on tissue remodelling were evaluated and discussed by histological examination as fibrosis/tissue remodelling has been demonstrated to link to the severity of mucosal inflammation and clinical symptoms such as loose stools.^2,27–31

The rates of endoscopic improvement were evaluated by the centrally read MES using images of the heaviest inflammation, which were provided from the local investigators in a blinded fashion. Although from a small sample size, we consider that the rate of endoscopic improvement of GUT-1 250 nM shows a sign of efficacy. First, a >1 point decline in MES in this relatively short period [weeks 2 or 4] is considered to be clinically meaningful and this was only achieved by GUT-1 250 nM, but not by placebo. Second, when considering previous reports showing that the rates of ‘mucosal healing’, which is equivalent to current ‘endoscopic improvement’, ranged from 11.6% to 31.3% in the placebo groups and from 27.0% to 41.1% in the active drug groups,^22,32–37 28.6% of placebo responses in this study were in the similar range as reported, while 71.2% in the GUT-1 250 nM was beyond the range of reported active drugs. In this regard, GUT-1 25 nM showed 0.0% of response in the MES. However, qualitative endoscopic findings in colonoscopy reports by local investigators showed that the numbers and sizes of ulcerated lesions were reduced in five patients who received 25 nM GUT-1. In addition, spontaneous bleeding at baseline was shown in four of seven patients but disappeared and turned into ‘light touch’ bleeding from week 2 in three of four patients. Since the number and size of ulcers as well as the bleeding condition was not reflected by the MES, MES did not change as long as small ulcers were present even if there was no spontaneous bleeding. We thus consider that the sign of healing was also found in the GUT-1 25 nM group although on a weak level and was not reflected by the MES.

Currently, the importance of histopathological assessment is increasing to evaluate reduction of mucosal inflammation as well as fibrosis in UC. Histological remission is an emergent treatment target, yet it is still hard to achieve as conventional biologics including infliximab and vedolizumab often fail to achieve histological improvement even after inducing endoscopic improvement.^38,39 GUT-1 250 nM showed histological improvement in 42.9% [0.0% for placebo], largely due to a reduction of neutrophils in both the epithelium and lamina propria. As CHST15-derived CS-E is a functional receptor for platelet factor 4/CXCL4 on the neutrophils for activation,^40,41 blockade of CS-E signalling may also suppress the activity of inflammatory neutrophils. Regarding the NHI, a >1 point decline was only achieved by GUT-1 250 nM but not by placebo, again indicating the potential of GUT-1 for induction of histological improvement.

Induction of clinical remission by a locally administered drug was an intriguing finding. Rapid reductions not only in the MES but also in the SFS at week 2 are considered to contribute to achieving clinical remission. As GUT-1 was shown to quickly inhibit submucosal oedema associated with acute inflammation in mice,⁹ the rapid improvement of stool frequency may be attributable to reduced submucosal oedema as well as reduced neutrophils. A rapid healing response at week 1 after single dosing of GUT-1 was previously observed in our Phase I trial in Crohn’s disease.^16,17 Since renewal of the entire intestinal epithelium is considered to occur every 3–5 days due to a high cellular turnover rate,⁴² the possibility anti-remodelling efficacy of GUT-1 may contribute to support adequate epithelial renewal within 2 weeks, leading to reduced bleeding mediated by epithelial injury or ulceration.

We previously reported that submucosally injected CHST15 siRNA was able to spread rapidly to at least 10 cm from the site of injection both circumferentially and longitudinally and to remain in the colon tissue for at least 6 days in mice.^9,17 Based on the non-clinical findings, we set the present injection procedure in an attempt to cover left-sided lesions and set week 2 and 4 as time points for evaluation in this first-in-patient study for UC. Inhibition of CHST15 by GUT-1 250 nM was observed as early as week 2 and the grade of inhibitions was still evident at week 4, although complete inhibition was not achieved by this dose and regimen. Meanwhile, GUT-1 25 nM inhibited CHST15 expression at week 2, but the inhibition seemed temporal. The kinetics of the target CHST15 protein expression after single dosing of GUT-1 25 nM suggest that the effect of GUT-1 at a lower concentration was insufficient and repeated dosing may be needed to increase efficacy. In addition, given that SFS and RBS remained stable from week 2 to 4, repeated injections, i.e. injections at weeks 0 and 2, may contribute to further suppress the clinical symptom scores during induction therapy.

There are several limitations to the present study. First, the sample size of 24 treated patients was not sufficient to allow a full clinical conclusion, especially regarding the rates of endoscopic improvement and clinical remission. The obtained results must be regarded as preliminary and need to be confirmed in subsequent larger clinical studies. In addition, the MES-based evaluation does not always adequately reflect endoscopic response, and therefore central evaluation of endoscopic improvement by the MES and another methodology using videorecording must be considered in future studies. Second, the observation period was limited to 2 or 4 weeks. Although the study period was reasonable to investigate signs of rapid onset of action of the applied oligonucleotide drug, most study periods for induction therapies are 8 weeks long. Third, the application was a single submucosal dosing. It would be of interest to see whether repeated injections during the induction period [i.e. 8 weeks] could further diminish mucosal CHST15 expression and enhance GUT-1’s effects on endoscopic improvement and clinical remission. Although GUT-1 250 nM was the highest dose tested, it remains unclear if even higher efficacy would be attainable using an increased dose. Fourth, only patients with left-sided colitis were included in our study. Fifth, qualitative histopathological examination revealed that GUT-1 250 nM diminished the area of established collagen fibrils in the inflamed mucosa, but pre-specified quantitative assessment for fibrosis was not performed. For assessment of fibrosis, the definitions of histological endpoints, grading scales, and scoring technique should be adequately defined in future clinical studies.

Several potential clinical advantages of GUT-1 are suggested by the presented study results. First, plasma concentrations of GUT-1 were below the limit of detection, suggesting a low probability of inducing systemic side effects. This would also comprise the absence of immunogenic reactions and subsequent loss of response. Second, a rapid onset of action of GUT-1 was observed. This is especially beneficial in refractory patients, as a slow acting drug might be insufficient in these cases, necessitating alternative therapeutic options including surgery. Third, GUT-1 was effective in anti-TNF refractory patients, in whom new therapeutic options are highly warranted. Fourth, local submucosal administration did not cause any local side effects and the total time spent on the eight injections was a few minutes.

In conclusion, the present study shows that local application of GUT-1 could be a novel option for induction therapy in active UC. The effects may be due to a reduced sulphated matrix and CS-E generation, by targeting CHST15, which simultaneously acts to repress neutrophil-based mucosal inflammation and accelerate degradation of established fibrosis, leading to tissue remodelling in UC.

Funding

This work was sponsored by Stelic Institute & Co., Inc.

Conflict of Interest

H.Y. was a founder of Stelic Institute & Co., and is an unpaid scientific advisor of GUT Inc. GUT Inc. has patents related to GUT-1 for the treatment of IBD. K.S. is an inventor of a patent related to submucosal injection. Other authors have no conflicts of interest. MN reports research grants and/or personal fees from Abbvie, MSD, Takeda, Boehringer, Roche, Pfizer, Janssen, Pentax, and PPD. RA has served as a speaker, consultant, or received research grants from AbbVie, Amgen, Arena Pharmaceuticals, Astra Zeneca, Biogen, Boehringer Ingelheim, Bristol-Myers Squibb, Cellgene, Celltrion Healthcare, DrFalk Pharma, Galapagos, Gilead, InDex Pharmaceuticals, Janssen-Cilag, Lilly, MSD Sharp & Dohme, Novartis, Pandion Therapeutics, Pfizer, Roche Pharma, Samsung Bioepsis, Stelic Institute, Takeda Pharma, and Viatris.

Acknowledgments

We are grateful to Professor Yoichi Ajioka at Division of Molecular and Diagnostic Pathology, Graduate School of Medical and Dental Sciences, Niigata University, Niigata, Japan, for his histological assessment.

Author Contributions

R.A., H.A., T.H., and M.N. oversaw the clinical trial, analysed and interpreted the data, and contributed to the writing and editing of the manuscript. R.A., H.Y., H.A., and M.N. planned the clinical study protocol. R.A., T.K., M.W., S.H., O.D., R.H., C.M., T.K., J.D., Je.M., C.S., B.H., Jo.M., and M.N. provided clinical and scientific input, recruited patients, and conducted the trial and reviewed the manuscript. K.S., J.Y., and S.T. performed molecular analyses and reviewed the manuscript. K.S. and J.Y. performed histological assessment.

Data Availability

Individual participant data collected during the trials after de-identification are available for the present study. The study protocol, statistical analysis plan, and informed consent form are also available for the period between the initial 3 months and last 5 years after article publication. These requests are reviewed and approved by an independent review panel based on scientific merit. All data provided are anonymized to respect the privacy of patients who have participated in the trial, in line with applicable laws and regulations. Proposals should be directed to corresponding author at [email protected]; to gain access, data requestors will need to sign a data access agreement.

References