Evaluation of antisense oligonucleotide therapy targeting Hsd17b13 in a fibrosis mice model.

Human genetic evidence suggests a protective role of loss-of-function variants in 17-beta hydroxysteroid dehydrogenase 13 (HSD17B13) for liver fibrotic diseases. Although there is limited preclinical experimental data on Hsd17b13 antisense oligonucleotide (ASO) or siRNA in a fibrosis model, several ASO and siRNA approaches are being tested clinically as potential therapies for nonalcoholic steatohepatitis (NASH). The aim of this study was to assess the therapeutic potential of Hsd17b13 ASO in a preclinical advanced NASH-like hepatic fibrosis in vivo model. In vitro testing on primary hepatocytes demonstrated that Hsd17b13 ASO exhibited strong efficacy and specificity for knockdown of the Hsd17b13 gene. In choline-deficient, L-amino acid-defined, HFD (CDAHFD)-induced steatotic and fibrotic mice, therapeutic administration of Hsd17b13 ASO resulted in a significant and dose-dependent reduction of hepatic Hsd17b13 gene expression. The CDAHFD group exhibited considerably elevated liver enzyme levels, hepatic steatosis score, hepatic fibrosis, and increased fibrotic and inflammatory gene expression, indicating an advanced NASH-like hepatic fibrosis phenotype. Although Hsd17b13 ASO therapy significantly affected hepatic steatosis, it had no effect on hepatic fibrosis. Our findings demonstrate, for the first time, that Hsd17b13 ASO effectively suppressed Hsd17b13 gene expression both in vitro and in vivo, and had a modulatory effect on hepatic steatosis in mice, but did not affect fibrosis in the CDAHFD mouse model of NASH.

MGAT2 inhibitor decreases liver fibrosis and inflammation in murine NASH models and reduces body weight in human adults with obesity.

Monoacylglycerol acyltransferase 2 (MGAT2) is an important enzyme highly expressed in the human small intestine and liver for the regulation of triglyceride absorption and homeostasis. We report that treatment with BMS-963272, a potent and selective MGAT2 inhibitor, decreased inflammation and fibrosis in CDAHFD and STAM, two murine nonalcoholic steatohepatitis (NASH) models. In high-fat-diet-treated cynomolgus monkeys, in contrast to a selective diacylglycerol acyltransferase 1 (DGAT1) inhibitor, BMS-963272 did not cause diarrhea. In a Phase 1 multiple-dose trial of healthy human adults with obesity (NCT04116632), BMS-963272 was safe and well tolerated with no treatment discontinuations due to adverse events. Consistent with the findings in rodent models, BMS-963272 elevated plasma long-chain dicarboxylic acid, indicating robust pharmacodynamic biomarker modulation; increased gut hormones GLP-1 and PYY; and decreased body weight in human subjects. These data suggest MGAT2 inhibition is a promising therapeutic opportunity for NASH, a disease with high unmet medical needs.

Combined Therapy with a CCR2/CCR5 Antagonist and FGF21 Analogue Synergizes in Ameliorating Steatohepatitis and Fibrosis.

(1) Background: With new potential drug targets emerging, combination therapies appear attractive to treat non-alcoholic steatohepatitis (NASH) and fibrosis. Chemokine receptor CCR2/5 antagonists can improve fibrosis by reducing monocyte infiltration and altering hepatic macrophage subsets. Fibroblast growth factor 21 (FGF21) may improve NASH by modulating lipid and glucose metabolism. We compared effects of single drug to combination treatment as therapeutic strategies against NASH. (2) Methods: We analyzed serum samples and liver biopsies from 85 nonalcoholic fatty liver disease (NAFLD) patients. A CCR2/5 inhibitor (BMS-687681-02-020) and a pegylated FGF21 agonist (BMS-986171) were tested in male C57BL/6J mice subjected to dietary models of NASH and fibrosis (choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) up to 12 weeks; short- (2w) or long-term (6w) treatment). (3) Results: In NAFLD patients, chemokine CCL2 and FGF21 serum levels correlated with inflammatory serum markers, only CCL2 was significantly associated with advanced liver fibrosis. In rodent NASH, CCR2/5 inhibition significantly reduced circulating Ly6C+ monocytes and hepatic monocyte-derived macrophages, alongside reduced hepatic inflammation and fibrosis. FGF21 agonism decreased body weight, liver triglycerides and histological NASH activity. Combination treatment reflected aspects of both compounds upon short- and long-term application, thereby amplifying beneficial effects on all aspects of steatohepatitis and fibrosis. (4) Conclusions: CCR2/5 inhibition blocks hepatic infiltration of inflammatory monocytes, FGF21 agonism improves obesity-related metabolic disorders. Combined therapy ameliorates steatohepatitis and fibrosis more potently than single drug treatment in rodent NASH, corroborating the therapeutic potential of combining these two approaches in NASH patients.

Evaluation of a PEGylated Fibroblast Growth Factor 21 Variant Using Novel Preclinical Magnetic Resonance Imaging and Magnetic Resonance Elastography in a Mouse Model of Nonalcoholic Steatohepatitis.

BACKGROUND: Treatments for nonalcoholic steatohepatitis (NASH) are urgently needed. Hepatic fat fraction and shear stiffness quantified by magnetic resonance imaging (MRI-HFF) and magnetic resonance elastography (MRE-SS), respectively, are biomarkers for hepatic steatosis and fibrosis. PURPOSE: This study assessed the longitudinal effects of fibroblast growth factor 21 variant (polyethylene glycol [PEG]-FGF21v) on MRI-HFF and MRE-SS in a NASH mouse model. STUDY TYPE: Preclinical. ANIMAL MODEL: This study included a choline-deficient, amino acid-defined, high-fat diet (CDAHFD) model and 6-week-old, male C57BL/6J mice (N = 78). FIELD STRENGTH/SEQUENCE: This study was performed using: 3T: gradient-echo two-point Dixon and spin-echo (SE) echo-planar imaging elastography (200 Hz) and 7T: SE two-point Dixon and SE elastography (200 Hz). ASSESSMENT: MRI and MRE were performed before control diet (CD) or CDAHFD (BD), before PEG-FGF21v dosing (baseline), and after PEG-FGF21v treatment (WK4/8). Regions of interest for MRI-HFF and MRE-SS were delineated by J.L. and H.T. (>5 years of experience each). Fibrosis and steatosis were measured histologically after picrosirius red and H&E staining. Alkaline phosphatase, alanine transaminase, bile acids, and triglycerides (TGs) were measured. STATISTICAL TESTS: Two-tailed Dunnett's tests were used for statistical analysis; untreated CDAHFD or baseline was used for comparisons. Imaging and histology/biochemistry data were determined using Spearman correlations. Bayesian posterior distributions for MRE-SS at WK8, posterior means, and 95% credible intervals were presented. RESULTS: CDAHFD significantly increased baseline MRI-HFF (3T: 21.97% ± 0.29%; 7T: 40.12% ± 0.35%) and MRE-SS (3T: 1.25 ± 0.02; 7T: 1.78 ± 0.06 kPa) vs. CD (3T: 3.45% ± 0.7%; 7T: 12.06% ± 1.4% and 3T: 1.01 ± 0.02; 7T: 0.89 ± 0.06 kPa). At 7T, PEG-FGF21v significantly decreased MRI-HFF (WK4: 28.97% ± 1.22%; WK8: 20.93% ± 1.15%) and MRE-SS (WK4: 1.57 ± 0.04; WK8: 1.36 ± 0.05 kPa) vs. untreated (WK4: 36.36% ± 0.62%; WK8: 30.58% ± 0.81% and WK4: 2.03 ± 0.06; WK8: 2.01 ± 0.04 kPa); 3T trends were similar. WK8 SS posterior mean percent attenuation ratios (RDI ) were -68% (-90%, -44%; 3T) and -64% (-78%, -52%; 7T). MRI-HFF was significantly correlated with H&E (3T, r = 0.93; 7T, r = 0.94) and TGs (both, r = 0.92). DATA CONCLUSIONS: MRI-HFF and MRE-SS showed PEG-FGF21v effects on hepatic steatosis and fibrosis across 3 and 7T, consistent with histological and biochemical data. LEVEL OF EVIDENCE: 1 TECHNICAL EFFICACY STAGE: 2.

Discovery of an Oxycyclohexyl Acid Lysophosphatidic Acid Receptor 1 (LPA1) Antagonist BMS-986278 for the Treatment of Pulmonary Fibrotic Diseases.

The oxycyclohexyl acid BMS-986278 (33) is a potent lysophosphatidic acid receptor 1 (LPA1) antagonist, with a human LPA1 Kb of 6.9 nM. The structure-activity relationship (SAR) studies starting from the LPA1 antagonist clinical compound BMS-986020 (1), which culminated in the discovery of 33, are discussed. The detailed in vitro and in vivo preclinical pharmacology profiles of 33, as well as its pharmacokinetics/metabolism profile, are described. On the basis of its in vivo efficacy in rodent chronic lung fibrosis models and excellent overall ADME (absorption, distribution, metabolism, excretion) properties in multiple preclinical species, 33 was advanced into clinical trials, including an ongoing Phase 2 clinical trial in patients with lung fibrosis (NCT04308681).

Discovery of 5-chloro-4-((1-(5-chloropyrimidin-2-yl)piperidin-4-yl)oxy)-1-(2-fluoro-4-(methylsulfonyl)phenyl)pyridin-2(1H)-one (BMS-903452), an antidiabetic clinical candidate targeting GPR119.

G-protein-coupled receptor 119 (GPR119) is expressed predominantly in pancreatic ß-cells and in enteroendocrine cells in the gastrointestinal tract. GPR119 agonists have been shown to stimulate glucose-dependent insulin release by direct action in the pancreas and to promote secretion of the incretin GLP-1 by action in the gastrointestinal tract. This dual mechanism of action has generated significant interest in the discovery of small molecule GPR119 agonists as a potential new treatment for type 2 diabetes. Herein, we describe the discovery and optimization of a new class of pyridone containing GPR119 agonists. The potent and selective BMS-903452 (42) was efficacious in both acute and chronic in vivo rodent models of diabetes. Dosing of 42 in a single ascending dose study in normal healthy humans showed a dose dependent increase in exposure and a trend toward increased total GLP-1 plasma levels.

Skeletal muscle contractile gene (TNNT3, MYH3, TPM2) mutations not found in vertical talus or clubfoot.

UNLABELLED: Arthrogryposis presents with lower limb contractures that resemble clubfoot and/or vertical talus. Recently, mutations in skeletal muscle contractile genes MYH3 (myosin heavy chain 3), TNNT3 (troponin T3), and TPM2 (tropomyosin 2) were identified in patients with distal arthrogryposis DA2A (Freeman-Sheldon syndrome) or DA2B (Sheldon-Hall syndrome). We asked whether the contractile genes responsible for distal arthrogryposis are also responsible for cases of familial clubfoot or vertical talus. We determined the frequency of MYH3, TNNT3, and TPM2 mutations in patients with idiopathic clubfoot, vertical talus, and distal arthrogryposis type 1 (DA1). We resequenced the coding exons of the MYH3, TNNT3, and TPM2 genes in 31 patients (five with familial vertical talus, 20 with familial clubfoot, and six with DA1). Variants were evaluated for segregation with disease in additional family members, and the frequency of identified variants was determined in a control population. In one individual with DA1, we identified a de novo TNNT3 mutation (R63H) previously identified in an individual with DA2B. No other causative mutations were identified, though we found several previously undescribed single-nucleotide polymorphisms of unknown importance. Although mutations in MYH3, TNNT3, and TPM2 are frequently associated with distal arthrogryposis syndromes, they were not present in patients with familial vertical talus or clubfoot. The TNNT3 R63H recurrent mutation identified in two unrelated individuals may be associated with either DA1 or DA2B. LEVEL OF EVIDENCE: Level II, prospective study. See the Guidelines for Authors for a complete description of levels of evidence.

Early results of the Ponseti method for the treatment of clubfoot in distal arthrogryposis.

BACKGROUND: Clubfoot occurs in approximately one in 1000 live births and is one of the most common congenital birth defects. Although there have been several reports of successful treatment of idiopathic clubfoot with the Ponseti method, the use of this method for the treatment of other forms of clubfoot has not been reported. The purpose of the present study was to evaluate the early results of the Ponseti method when used for the treatment of clubfoot associated with distal arthrogryposis. METHODS: Twelve consecutive infants (twenty-four feet) with clubfoot deformity associated with distal arthrogryposis were managed with the Ponseti method and were retrospectively reviewed at a minimum of two years. The severity of the foot deformity was classified according to the grading system of Diméglio et al. The number of casts required to achieve correction was compared with published data for the treatment of idiopathic clubfoot. Recurrent clubfoot deformities or complications during treatment were recorded. RESULTS: Twenty-two clubfeet in eleven patients were classified as Diméglio grade IV, and two clubfeet in one patient were classified as Diméglio grade II. Initial correction was achieved in all clubfeet with a mean of 6.9 +/- 2.1 casts (95% confidence interval, 5.6 to 8.3 casts), which was significantly greater than the mean of 4.5 +/- 1.2 casts (95% confidence interval, 4.3 to 4.7 casts) needed in a cohort of 219 idiopathic clubfeet that were treated during the same time period by the senior author with use of the Ponseti method (p = 0.002). Six feet in three patients had a relapse after initial successful treatment. All relapses were related to noncompliance with prescribed brace wear. Four relapsed clubfeet in two patients were successfully treated with repeat casting and/or tenotomy; the remaining two relapsed clubfeet in one patient were treated with extensive soft-tissue-release operations. CONCLUSIONS: Our early-term results support the use of the Ponseti method for the initial treatment of distal arthrogrypotic clubfoot deformity. Longer follow-up will be necessary to assess the risk of recurrence and the potential need for corrective clubfoot surgery in this patient population, which historically has been difficult to treat nonoperatively.

Impact of congenital talipes equinovarus etiology on treatment outcomes.

Although congenital talipes equinovarus (CTEV) is often idiopathic, additional birth defects occur in some patients that may have an impact on the treatment of this disorder. The purpose of this study was to determine the prevalence of associated malformations, chromosomal abnormalities, or known genetic syndromes, and to compare treatment outcomes of children with idiopathic CTEV with children with non-idiopathic CTEV. Of 357 children evaluated, 273 (76%) had idiopathic CTEV (179 males, 94 females; mean age 2 y 1 mo [SD 1 y 2 mo], range 0-18 y) and 84 (24%) had non-idiopathic CETV (51 males, 33 females; mean age 2 y 5 mo [SD 2 y], range 0-16 y). Disorders affecting the nervous system were found in 46 (54%) children with non-idiopathic CTEV. In a subgroup of patients treated entirely at our institution (n=196), children with non-idiopathic CTEV (n=47) required more casts for correction than those with idiopathic CTEV (n=149; 5.3 vs 4.6; p=0.016). There was also a greater risk of recurrence in non-idiopathic CTEV (14.9% vs 4%; p=0.009), but no significant difference in the need for extensive surgery (2.7% vs 8.5%; p=0.096). Treatment was initiated at a mean age of 13 weeks (range 1 wk to 2 y 6 mo) for both idiopathic and non-idiopathic patients, and treatment was assessed during a minimum 2-year follow-up. Non-idiopathic CTEV can be successfully treated with the Ponseti method of serial casting, with low recurrence rates or need for surgery.

Case report: Congenital knee dislocation in a patient with larsen syndrome and a novel filamin B mutation.

We treated a patient with multiple congenital joint dislocations and facial dysmorphisms consistent with Larsen syndrome. Sequencing of the FLNB gene resulted in identification of a novel, de novo 508G>C point mutation resulting in substitution of proline for a highly conserved alanine (A170P). This mutation has not been described previously but is likely causative because this alanine is highly conserved and is located in the calponin homology domain where other mutations have been described. We also report the successful use of a minimally invasive technique in achieving initial correction of bilateral congenital knee dislocations in this patient. The technique consists of serial manipulations and castings followed by an open quadriceps tenotomy. Longer followup is needed to ensure maintenance of correction and to avoid the need for more extensive surgery, which has been the traditional treatment for congenital knee dislocation associated with Larsen syndrome.

Foot abduction brace in the Ponseti method for idiopathic clubfoot deformity: torsional deformities and compliance.

The Ponseti method has shown to be effective in treating idiopathic clubfoot deformity. Application of a foot abduction brace is crucial to long term outcome. Compliance and possible effect on femoral anteversion and tibial torsion were assessed in 20 children with an average bracing period of 33 months. Nineteen patients were compliant with the bracing protocol and highly satisfied with the treatment protocol. Ultrasound measurement of femoral anteversion (40.5 degrees) and tibial external torsion (38.4 degrees) of the affected side were within the normal range. Range of motion of the ankle joint in all clubfeet resulted in an average dorsiflexion of 27.5 degrees. Among our patient group, the method proved to be highly accepted by almost all families, with excellent functional results. Application of the foot abduction brace did not result in pathological changes of femoral anteversion or tibial torsion.

A new approach to the treatment of congenital vertical talus.

Congenital vertical talus is an uncommon foot deformity that is present at birth and results in a rigid flatfoot deformity. Left untreated the deformity can result in pain and disability. Though the exact etiology of vertical talus is unknown, an increasing number of cases have been shown to have a genetic cause. Approximately 50% of all cases of vertical talus are associated with other neuromuscular abnormalities or known genetic syndromes. The remaining 50% of cases were once thought to be idiopathic in nature. However, there is increasing evidence that many of these cases are related to single gene defects. Most patients with vertical talus have been treated with major reconstructive surgeries that are fraught with complications such as wound necrosis, talar necrosis, undercorrection of the deformity, stiffness of the ankle and subtalar joint, and the eventual need for multiple operative procedures. Recently, a new approach to vertical talus that consists of serial casting and minimal surgery has resulted in excellent correction in the short-term. Longer follow-up will be necessary to ensure maintenance of correction with this new technique. A less invasive approach to the correction of vertical talus may provide more favorable long-term outcomes than more extensive surgery as has been shown to be true for clubfoot outcomes.

Synovial chondromatosis of the hip: a case report and clinicopathologic study.

Primary synovial chondromatosis (PSC) is a rare, usually monoarticular disorder of synovial joints. PSC is characterised by the formation of osteocartilaginous nodules in the synovial connective tissue. We report the case of a 32-year-old male with PSC of the left hip. At clinical examination abduction of the left hip was limited and rotation was painful. Ultrasound examination of the hip revealed joint effusion and multiple hyperechogenic foci due to distal acoustic shadowing. Plain radiographs showed a slight soft tissue swelling around the femoral neck and multiple round or ovoid calcifications of a uniform size. MRI revealed a large joint effusion with multiple small filling defects. Open total synovectomy was performed after dislocation of the femoral head. The diagnosis of PSC was confirmed by histological examination of the excised material. The majority of cells failed to exhibit any staining for cerb B-2 and ki-67. None of the sections showed more than 5% labelling for DNA-fragmentation proven by terminal deoxytransferase-mediated dUTD nick-end labeling (TUNEL), and all were completely non-reactive for p53 as well. In conclusion, immunohistochemical analysis suggests that in this case PSC originated from metaplasia and not from a proliferative process. After two years, the patient was free of symptoms and radiological control did not show evidence of recurrence or femoral head necrosis. Physical findings, diagnosis, histological features and management of PSC are discussed.