Mixed active metabolites of the SNP-6 series of novel compounds mitigate metabolic dysfunction-associated steatohepatitis and fibrosis: promising results from pre-clinical and clinical trials

Abstract

Background: Metabolic dysfunction-associated steatohepatitis (MASH) is a growing global health concern with no effective pharmacological treatments. SNP-630, a newly developed synthetic molecule with multiple mechanisms of action, and a mixture of two of its active metabolites (SNP-630-MS) inhibit CYP2E1 expression to prevent reactive oxygen species generation, thereby reducing the accumulation of hepatic triglycerides and lowering chemokine levels. This study investigated the SNP-630's potential to alleviate the liver injury in MASH and its efficacy in both a mouse model and patients with MASH to identify a drug candidate that targets multiple pathways implicated in MASH.

Methods: SNP-630 and SNP-630-MS were separately administered to the MASH mouse model. The tolerability, safety, and efficacy of SNP-630-MS were also evaluated in 35 patients with MASH. The primary endpoint of the study was assessment of the changes in serum alanine aminotransferase (ALT) levels from baseline to week 12, while the secondary endpoints included the evaluation of liver inflammation, steatosis, and fibrosis parameters and markers.

Results: SNP-630 treatment in mice improved inflammation, liver steatosis, and fibrosis compared with that in the MASH control group. Both SNP-630 and SNP-630-MS treatments markedly reduced ALT levels, hepatic triglyceride content, and the expression of inflammatory cytokines monocyte chemoattractant protein 1 and fibrotic collagen (i.e., Col1a1, Col3a1, and Timp1) in mice. In the clinical trial, patients treated with SNP-630-MS exhibited significant improvement in ALT levels at week 12 compared with baseline levels, with no reports of severe adverse events. This improvement in ALT levels surpassed that achieved with most other MASH candidates. SNP-630-MS demonstrated potential antifibrotic effects, as evidenced by a significant decrease in the levels of fibrogenesis-related biomarkers such as CCL4, CCL5, and caspase 3. Subgroup analysis using FibroScan measurements further indicated the efficacy of SNP-630-MS in ameliorating liver fibrosis.

Conclusions: SNP-630 and SNP-630-MS demonstrated favorable results in mice. SNP-630-MS showed excellent tolerability in mice and patients with MASH. Efficacy analyses indicated that SNP-630-MS improved liver steatosis and injury in patients with MASH, suggesting that SNP-630 and 630-MS are promising therapeutic options for MASH. Larger scale clinical trials remain warranted to assess the efficacy and safety of SNP-630 in MASH.

Trial registration: ClinicalTrials.gov NCT03868566. Registered 06 March 2019-Retrospectively registered, https://clinicaltrials.gov/study/NCT03868566.

Keywords: Alanine aminotransferase; Metabolic dysfunction-associated steatohepatitis; SNP-630.

Fig. 1

Metabolic scheme and profile of SNP-630. a Schematic illustration of SNP-630 metabolism. The structures of new chemical entities, SNP-630 and SNP-630-MS, are shown. b Metabolic profiles of SNP-630 (% remaining) and its metabolites (% formation) in human blood

Fig. 2

SNP-630 treatment alleviates HFD-induced hepatic steatosis, inflammation, and fibrosis in the MASH mouse model. a SNP-630 inhibits CYP2E1 catalytic activity. The metabolic ratios of chlorzoxazone (CZX) and its CYP2E1-drived metabolite, 6-hydroxychlorzoxazone (6-OH-CZX), in mice are plotted versus the SNP-630 treatment dose. Data are presented as mean ± SE (n = 8 for each group). b Representative immunoblots of SNP-630 treatment significantly reduced CYP2E1 protein expression. Western blotting of CYP2E1 and β-actin was performed in mouse liver lysates. c Quantification of CYP2E1 protein expression level. d Liver/body weight ratio (e) Liver TG (f) Liver total cholesterol. g ALT h Histopathological analysis of liver tissues. Representative images of Sirius red (left) and H&E (right) staining of livers from the blank group, HFD group, and SNP-630 250 mg/kg group (scale bar: 50 μm). i NAFLD Activity Score (NAS) (j) Hepatic collagen deposition. Fields from Sirius red-stained sections were scanned using a slide scanner (Axio Scan.Z1), and the fibrotic area was measured with digital image analysis using the ImageJ software (k) Representative results of expression of hepatic Col3a1 mRNA. l Representative results of expression of hepatic Timp1 mRNA. Data are expressed as mean ± SEM. Statistical analyses were carried out using a one-way analysis of variance (n = 8/each group). MASH: Metabolic dysfunction-associated steatohepatitis; HFD: high-fat diet

Fig. 3

Hepaprotective effects of SNP-630-MS on the HFD-induced mouse model of MASH. MASH was induced in male C57BL/6 mice with HFD for 21 weeks. HFD-induced MASH mice were divided into three groups and received an HFD along with vehicle, 187.5/187.5 (assigned as SNP-630-MS-High), or 62.5/62.5 mg/kg SNP-630-MS (assigned as SNP-630-MS-Low) by oral gavage once daily. Mice on a normal chow diet received the vehicle as the normal control (blank group). a Body weight, b liver/body weight ratio, c liver TG, d liver TCHO, e ALT, f NAS, g fibrosis stage, h expression of hepatic Col3a1 mRNA, and (i) expression of hepatic Timp1 mRNA. Data are expressed as mean ± SEM. Statistical analyses were carried out using a one-way analysis of variance (n = 12/each group). MASH: Metabolic dysfunction-associated steatohepatitis; HFD: high-fat diet

Fig. 4

SNP-630-MS clinical trial profile. a The disposition of one participant who withdrew after 3 days of dosing cannot be reported. b Analysis of primary efficacy for the modified intention-to-treat (mITT) population. ALT: alanine aminotransferase

Fig. 5

SNP-630-MS met the primary endpoint of improvement in ALT after 12-week treatment in mITT population. a Participants meet the primary endpoint (improvement in ALT). Mean changes from baseline during treatment with two SNP-630-MS tablets (n = 17) and one SNP-630-MS tablet (n = 18) daily for up to 12 weeks. Error bars show standard errors. *P < 0·05, ***P < 0.005 compared with baseline. b Comparison of the FibroScan results at different stages. Mean ± standard deviation (n = 5–12). c Subgroup analyses for the primary endpoint of reduction in ALT levels. Response by baseline fibrosis stage (FibroScan), PNPLA3 genotype, sex, age, BMI, MRI-PDFF, type 2 diabetes, and dose. mITT modified intention-to-treat, PNPLA3 patatin-like phospholipase domain-containing protein 3, BMI body mass index, MRI-PDFF magnetic resonance imaging-proton density fat fraction, ALT alanine aminotransferase