Absorption, Distribution, Metabolism, and Excretion of US Food and Drug Administration-Approved Antisense Oligonucleotide Drugs

Abstract

Absorption, distribution, metabolism, and excretion (ADME) are the key biologic processes for determination of a drug's pharmacokinetic parameters, which have direct impacts on efficacy and adverse drug reactions (ADRs). The chemical structures, dosage forms, and sites and routes of administration are the principal determinants of ADME profiles and consequent impacts on their efficacy and ADRs. Newly developed large molecule biologic antisense oligonucleotide (ASO) drugs have completely unique ADME that is not fully defined. ASO-based drugs are single-stranded synthetic antisense nucleic acids with diverse modes of drug actions from induction of mRNA degradation, exon skipping and restoration, and interactions with proteins. ASO drugs have a great potential to treat certain human diseases that have remained untreatable with small molecule-based drugs. The ADME of ASO drugs contributes to their unique set of ADRs and toxicity. In this review, to better understand their ADME, the 10 US Food and Drug Administration (FDA)-approved ASO drugs were selected: fomivirsen, pegaptanib, mipomersen, nusinersen, inotersen, defibrotide, eteplirsen, golodirsen, viltolarsen, and casimersen. A meta-analysis was conducted on their formulation, dosage, sites of administration, local and systematic distribution, metabolism, degradation, and excretion. Membrane permeabilization through endocytosis and nucleolytic degradation by endonucleases and exonucleases are major ADME features of the ASO drugs that differ from small-molecule drugs. The information summarized here provides comprehensive ADME characteristics of FDA-approved ASO drugs, leading to a better understanding of their therapeutic efficacy and their potential ADRs and toxicity. Numerous knowledge gaps, particularly on cellular uptake and subcellular trafficking and distribution, are identified, and future perspectives and directions are discussed. SIGNIFICANCE STATEMENT: Through a systematic analysis of the existing information of absorption, distribution, metabolism, and excretion (ADME) parameters for 10 US Food and Drug Administration (FDA)-approved antisense oligonucleotide (ASO) drugs, this review provides an overall view of the unique ADME characteristics of ASO drugs, which are distinct from small chemical drug ADME. This knowledge is useful for discovery and development of new ASO drugs as well as clinical use of current FDA-approved ASO drugs.

Fig. 1.

Timeline of the 10 FDA-approved antisense oligonucleotide (ASO) drugs with FDA approval dates, companies that developed the drugs, and target diseases. FH, familial hypercholesterolemia; PNA, polyneuropathy of amyloidosis; SMA, spinal muscular atrophy; VOD, veno-occlusive disease.

Fig. 2.

The mode of drug action of the 10 FDA-approved ASO drugs. (A) ASOs with RNase H-mediated mRNA degradation. By binding to their target mRNAs, ASO drugs cause the targeted mRNAs to be broken down into smaller pieces within the cell. ASO drugs in this modality are fomivirsen targeting on CMV IE2 mRNA, mipomersen targeting on Apo B mRNA, and inotersen targeting on transthyretin (TTR) mRNA. (B) ASO-mediated exon skipping. By sterically blocking a splicing site in the exon with mutations, ASO drugs result in skipping of the mutated exon and increased production of protein with enhanced function due to the skipped exon. ASO drugs in this mode include eteplirsen for exon 51 skipping, golodirsen and viltolarsen for exon 53 skipping, and casimersen for exon 45 skipping in Dystrophin mRNA. (C) ASO-mediated exon inclusion. By sterically blocking the splicing site in intron 7 of the SMN2 pre-mRNA, nusinersen blocks key intrinsic splicing factors heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) and heterogeneous nuclear ribonucleoprotein A2 (hnRNP A2) and results in translation of full-length, functional SMN2 protein. (D) Aptamer ASO. Pegaptanib is a polynucleotide aptamer, which undergoes folding to bind to targeted vascular endothelial growth factor (VEGF). This prevents a cascade of kinase-mediated cell signaling, which then results in angiogenesis. (E) ASO with multiple modes. Defibrotide aims to bind to adenosine receptors and trigger a cascade of events. Plasmin is inhibited and can no longer hydrolyze clots. Key proclotting factors such as VEGF, tumor necrosis factor (TNF-α), intracellular adhesion molecule 1 (ICAM1), endothelium, plasminogen activator inhibitor 1 (PAI1), thromboxane A2 (TXA2), and thrombin are decreased. Anticlotting factors such as prostaglandins I2 and E2 and tissue factor pathway inhibitor (TFPI) all are increased. Fibrinopeptide A levels are increased due to chronic coagulation from disease. Clot-free blood vessels lead to general healing of damaged endothelial cells and tissue (Pescador et al., 2013).

Fig. 3.

Administration paths, formulation, and dosing information of the FDA-approved ASO drugs. The ASO drugs are divided based on their administration paths with (A) intravitreal injection, (B) intrathecal injection, (C) subcutaneous injection, and (D) intravenous infusion. The information for formulation and dosing is extracted from the FDA drug labels at Drugs@FDA: FDA-Approved Drugs (https://www.accessdata.fda.gov/scripts/cder/daf/index.cfm).

Fig. 4.

Comparison of absorption, distribution, metabolism, and excretion (ADME) of small chemical drugs and ASO drugs. (A) ADME of an orally administrated small chemical drug. After absorption, a small chemical drug is normally distributed to metabolic organs of intestine, liver, and kidney via the bloodstream. In the metabolic organs, such as the liver, the drug is taken up by hepatocytes through either passive or active transport against concentration gradients with the help of uptake transporters. Normally, the lipophilic drug is biotransformed by either Phase I or II metabolizing enzymes to more polar water-soluble metabolites in rough endoplasmic reticulum (ER) membrane. The parental drug and metabolites are excreted from hepatocyte cells via efflux transporters into the bloodstream and then become available as an active form to target organs. Therapeutic concentrations of the drug in its target organs are determined by uptake and efflux transporters and Phase I and II metabolizing enzymes, which are tightly regulated at transcriptional and translation levels by transcription factors and nuclear receptors. (B) ADME of an ASO drug. After an intravenous infusion, subcutaneous injection, or direct injection, ASOs travel through the bloodstream, where they can be broken down via endogenous exo- and endonucleases. Once ASOs reach their target organ/cell, such as the liver, they enter cells through various endocytic pathways, including clathrin-mediated, caveolin-mediated, and micropinocytosis endocytosis, into early endosomes (EEs). After being internalized into EEs, ASO drugs go through subcellular trafficking to the late endosomes. A proportion of ASO drugs (maybe very small proportion) is released from late endosomes to cytoplasm to target mRNAs or pre-mRNAs in either the cytoplasm or the nucleus, where they execute their therapeutic actions. Many small cellular proteins including COPII can assist subcellular trafficking to the nucleus. However, the mechanism of nuclear trafficking is not fully understood. Another proportion of ASO drugs undergo into lysosomes or further are released out of the cell via three proposed mechanisms: membrane leakage, back-fusion mediated release, and vesicle-mediated release. Back-fusion mediated release is believed to help endogenous exo- and endonucleases break the ASO down further. ASO drugs may also leave the cell completely via the exosome release from the cell.