Prothrombin prevents fatal T cell-dependent anemia during chronic virus infection of mice.

Thrombin promotes the proliferation and function of CD8+ T cells. To test if thrombin prevents exhaustion and sustains antiviral T cell activity during chronic viral infection, we depleted the thrombin-precursor prothrombin to 10% of normal levels in mice prior to infection with the clone 13 strain of lymphocytic choriomeningitis virus. Unexpectedly, prothrombin insufficiency resulted in 100% mortality after infection that was prevented by depletion of CD8+ T cells, suggesting that reduced availability of prothrombin enhances virus-induced immunopathology.

Targeting the contact system in a rabbit model of extracorporeal membrane oxygenation.

Previous studies suggested that contact pathway factors drive thrombosis in mechanical circulation. We used a rabbit model of veno-arterial extracorporeal circulation (VA-ECMO) to evaluate the role of factors XI and XII in ECMO-associated thrombosis and organ damage. Factors XI and XII (FXI, FXII) were depleted using established antisense oligonucleotides before placement on a blood-primed VA-ECMO circuit.

CFTR mRNAs with nonsense codons are degraded by the SMG6-mediated endonucleolytic decay pathway.

Approximately 10% of cystic fibrosis patients harbor nonsense mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene which can generate nonsense codons in the CFTR mRNA and subsequently activate the nonsense-mediated decay (NMD) pathway resulting in rapid mRNA degradation. However, it is not known which NMD branches govern the decay of CFTR mRNAs containing nonsense codons. Here we utilize antisense oligonucleotides targeting NMD factors to evaluate the regulation of nonsense codon-containing CFTR mRNAs by the NMD pathway.

Antisense oligonucleotide-mediated knockdown of Mpzl3 attenuates the negative metabolic effects of diet-induced obesity in mice.

Previously, we demonstrated that global knockout (KO) of the gene encoding myelin protein zero-like 3 (Mpzl3) results in reduced body weight and adiposity, increased energy expenditure, and reduced hepatic lipid synthesis in mice. These mice also exhibit cyclic and progressive alopecia which may contribute to the observed hypermetabolic phenotype. The goal of the current study was to determine if acute and peripherally restricted knockdown of Mpzl3 could ameliorate the negative metabolic effects of exposure to a high-fat and sucrose, energy-dense (HED) diet similar to what was observed in global Mpzl3 KO mice in the absence of a skin phenotype.

Pharmacological targeting of coagulation factor XI mitigates the development of experimental atherosclerosis in low-density lipoprotein receptor-deficient mice.

Background: Human coagulation factor (F) XI deficiency, a defect of the contact activation system, protects against venous thrombosis, stroke, and heart attack, whereas FXII, plasma prekallikrein, or kininogen deficiencies are asymptomatic. FXI deficiency, inhibition of FXI production, activated FXI (FXIa) inhibitors, and antibodies to FXI that interfere with FXI/FXII interactions reduce experimental thrombosis and inflammation. FXI inhibitors are antithrombotic in patients, and FXI and FXII deficiencies are atheroprotective in apolipoprotein E-deficient mice.

Ligand conjugated antisense oligonucleotide for the treatment of transthyretin amyloidosis: preclinical and phase 1 data.

Aims: Amyloidogenic transthyretin (ATTR) amyloidosis is a fatal disease characterized by progressive cardiomyopathy and/or polyneuropathy. AKCEA-TTR-L (ION-682884) is a ligand-conjugated antisense drug designed for receptor-mediated uptake by hepatocytes, the primary source of circulating transthyretin (TTR). Enhanced delivery of the antisense pharmacophore is expected to increase drug potency and support lower, less frequent dosing in treatment.

Immunogenicity Assessment of Inotersen, a 2'--(2-Methoxyethyl) Antisense Oligonucleotide in Animals and Humans: Effect on Pharmacokinetics, Pharmacodynamics, and Safety.

Inotersen (TEGSEDI™) is a 2'--(2-methoxyethyl)-modified antisense oligonucleotide, intended for treating hereditary transthyretin (TTR) amyloidosis with polyneuropathy. The potential immunogenicity (IM) response to inotersen was evaluated in chronic nonclinical safety studies and the pivotal phase 2/3 clinical study. The evaluation was designed to assess the characteristics of antidrug antibodies (ADAs) and their effects on the pharmacokinetics, pharmacodynamics, clinical efficacy, and safety in animals and humans.

SERPINA1 modulates expression of amyloidogenic transthyretin.

Hereditary transthyretin amyloidosis (ATTR) is caused by amyloid deposition of misfolded transthyretin (TTR) in various tissues. Recently, reduction of circulating serum TTR, achieved via silencing oligonucleotides, was introduced as therapy of ATTR amyloidosis. We explored the impact of Serpin Family A Member 1 (SERPINA1) on TTR mRNA and protein expression.

Evaluation of the Antihemostatic and Antithrombotic Effects of Lowering Coagulation Factor VII Levels in a Non-human Primate.

Introduction: Tissue factor (TF) and factor (F) VII, components of the extrinsic pathway of blood coagulation, are essential for hemostatic plug formation in response to injury; less clear are their roles in propagating thrombosis, as observational data in humans with congenital FVII deficiency suggests persistent thrombotic and bleeding risk even at significantly decreased FVII levels. We aimed to define the contribution of FVII to thrombus formation and hemostasis using a non-human primate model.

Methods: We treated baboons with a FVII antisense oligonucleotide (ASO) and measured platelet and fibrin deposition inside and distal to collagen- or TF-coated vascular grafts.

Population Pharmacokinetic-Pharmacodynamic Modeling of Inotersen, an Antisense Oligonucleotide for Treatment of Patients with Hereditary Transthyretin Amyloidosis.

A population pharmacokinetic (PK) and pharmacodynamic (PD) model was developed for inotersen to evaluate exposure-response relationships and to optimize therapeutic dosing regimen in patients with hereditary transthyretin (TTR) amyloidosis polyneuropathy (hATTR-PN). Inotersen PK and TTR level (PD) data were composed of one Phase 1 study in healthy subjects, one Phase 2/3 study in hATTR patients, and its one open-label extension study. Effects of intrinsic and extrinsic factors (covariates) on PK and PK/PD of inotersen were evaluated using a full model approach.

Antisense Oligonucleotides Targeting Jagged 1 Reduce House Dust Mite-induced Goblet Cell Metaplasia in the Adult Murine Lung.

Goblet cell metaplasia, excessive mucus production, and inadequate mucus clearance accompany and exacerbate multiple chronic respiratory disorders, such as asthma and chronic obstructive pulmonary disease. Notch signaling plays a central role in controlling the fate of multiple cell types in the lung, including goblet cells. In the present study, we explored the therapeutic potential of modulating the Notch pathway in the adult murine lung using chemically modified antisense oligonucleotides (ASOs).

Underlying Immune Disorder May Predispose Some Transthyretin Amyloidosis Subjects to Inotersen-Mediated Thrombocytopenia.

Inotersen, a 2'-O-methoxyethyl (2'-MOE) phosphorothioate antisense oligonucleotide, reduced disease progression and improved quality of life in patients with hereditary transthyretin amyloidosis with polyneuropathy (hATTR-PN) in the NEURO-TTR and NEURO-TTR open-label extension (OLE) trials. However, 300 mg/week inotersen treatment was associated with platelet count reductions in several patients. Mean platelet counts in patients in the NEURO-TTR-inotersen group remained ≥140 × 10/L in 50% and ≥100 × 10/L in 80% of the subjects.

Hepatic thrombopoietin gene silencing reduces platelet count and breast cancer progression in transgenic MMTV-PyMT mice.

In humans, platelet count within the normal range is required for physiological hemostasis, but, adversely, platelets also support pathological thrombosis. Moreover, by releasing growth factors, they may enhance neoplastic proliferation. We hypothesize that platelet count correlates with platelet-dependent pathologies, even within the range of hemostatic competence.

Steric Inhibition of 5' UTR Regulatory Elements Results in Upregulation of Human CFTR.

Cystic fibrosis (CF) is an autosomal recessive monogenic disease caused by mutations in the CFTR gene. Therapeutic approaches that are focused on correcting CFTR protein face the challenge of the heterogeneity in CFTR mutations and resulting defects. Thus, while several small molecules directed at CFTR show benefit in the clinic for subsets of CF patients, these drugs cannot treat all CF patients.

Antisense oligonucleotide treatment ameliorates IFN-γ-induced proteinuria in APOL1-transgenic mice.

African Americans develop end-stage renal disease at a higher rate compared with European Americans due to 2 polymorphisms (G1 and G2 risk variants) in the apolipoprotein L1 (APOL1) gene common in people of African ancestry. Although this compelling genetic evidence provides an exciting opportunity for personalized medicine in chronic kidney disease, drug discovery efforts have been greatly hindered by the fact that APOL1 expression is lacking in rodents. Here, we describe a potentially novel physiologically relevant genomic mouse model of APOL1-associated renal disease that expresses human APOL1 from the endogenous human promoter, resulting in expression in similar tissues and at similar relative levels as humans.

Targeting Translation Termination Machinery with Antisense Oligonucleotides for Diseases Caused by Nonsense Mutations.

Efforts to develop treatments for diseases caused by nonsense mutations have focused on identification of small molecules that promote translational read-through of messenger RNAs (mRNAs) harboring nonsense stop codons to produce full-length proteins. However, to date, no small molecule read-through drug has received FDA approval, probably because of a lack of balance between efficacy and safety. Depletion of translation termination factors eukaryotic release factor () 1 and in cells was shown to promote translational read-through of a luciferase reporter gene harboring a nonsense mutation.

Thrombin Signaling Promotes Pancreatic Adenocarcinoma through PAR-1-Dependent Immune Evasion.

Pancreatic ductal adenocarcinoma (PDAC) is associated with robust activity of the coagulation system. To determine mechanisms by which clotting factors influence PDAC tumor progression, we generated and characterized C57Bl/6-derived KPC ( ) cell lines. Tissue factor (TF) and protease-activated receptor-1 (PAR-1) were highly expressed in primary KPC pancreatic lesions and KPC cell lines similar to expression profiles observed in biopsies of patients with PDAC.

Sex-specific differences in emphysema using a murine antisense oligonucleotide model of α-1 antitrypsin deficiency.

α-1 Antitrypsin (AAT) deficiency is the leading genetic cause of emphysema; however, until recently, no genuine animal models of AAT deficiency existed, hampering the development of new therapies. This shortcoming is now addressed by both -null and antisense oligonucleotide mouse models. The goal of this study was to more fully characterize the antisense oligonucleotide model.

Nonsense-mediated RNA Decay Pathway Inhibition Restores Expression and Function of W1282X CFTR.

The recessive genetic disease cystic fibrosis (CF) is caused by loss-of-function mutations in the CFTR (CF transmembrane conductance regulator) gene. Approximately 10% of patients with CF have at least one allele with a nonsense mutation in CFTR. Nonsense mutations generate premature termination codons that can subject mRNA transcripts to rapid degradation through the nonsense-mediated mRNA decay (NMD) pathway.

IONIS-PKK a Novel Antisense Inhibitor of Prekallikrein and Bradykinin Production.

Kallikrein is the key contact system mediator responsible for the conversion of high-molecular-weight kininogen into the inflammatory vasodilator peptide bradykinin, a process regulated by C1-esterase inhibitor (C1-INH). In hereditary angioedema (HAE), genetic mutations result in deficient or dysfunctional C1-INH and dysregulation of the contact system leading to recurrent, sometimes fatal, angioedema attacks. IONIS-PKK is a second-generation 2'-O-(2-methoxyethyl)-modified chimeric antisense oligonucleotide, designed to bind and selectively reduce prekallikrein (PKK) mRNA in the liver.