RNA Interference Therapeutics

Design of siRNA therapeutics involves in silico generation of sequences against the open reading frame of the gene of interest for all available isoforms, using published sequence information. Ideal siRNA length, chemistry, and delivery mechanism should be considered at this stage, and are all dependent on the target cell type. Attention should also be paid to sequence specificity and unspecific binding, thermodynamics, and cross-species homology, especially if animal models are to be utilized in efficacy and safety testing stages of discovery.

Selecting lead candidates for further development requires a combination of bioinformatics and in vitro screening. High-throughput screening for RNAi therapeutics is conducted in 384-well assay formats to identify lead candidates. At this stage, the target gene of interest and therapeutic indication will dictate the specific cell model used, for example, siRNA therapeutics for liver disorders can be screened in iPSC-derived hepatocytes. Our experts design screening cascades to identify the best lead candidates to take forward, examining the effect of sequence length, chemical modifications, and binding location, for example.

Once lead candidates have been selected, disease-relevant cell and animal models can be employed to demonstrate efficacy, investigate toxicity potential, explore delivery mechanisms, assess tolerability, and identify translational biomarkers.

Our expert teams can advise on the most suitable models and readouts for your program, considering disease, model phenotype, and translation into the clinical setting. We have a wide range of models for the most common areas for which siRNA therapeutics are being developed, including cardiovascular and metabolic disorders, cancer, and neurological disorders.

Early assessment of the toxicity potential of any lead candidate is vital to ensure that you proceed through drug discovery with the most suitable candidate. siRNA therapeutics are known to trigger innate immune responses as they may be recognized as viral byproducts. While this can be mitigated through chemical modification, it’s critical to investigate this in a human cell model, such as human peripheral blood mononuclear cells (PBMCs).

Your strategy for toxicity and safety studies should be designed to support clinical development and therapeutic use, using translational delivery methods and readouts where possible. For acute, sub-chronic, and chronic toxicity studies, our RNA experts can advise on the most appropriate species selection as well as selection of the most appropriate, translational route of administration.

Filing an investigational new drug (IND) application is a vital regulatory step in RNAi therapeutic development. Our RNA experts and Scientific Advisory Services (SAS) team can support you in planning, launching, and completing IND-enabling studies, with nonclinical studies designed to reflect the objectives of your clinical program.

RNA interference (RNAi) is an endogenous intracellular process where RNA molecules suppress gene expression in a sequence-specific manner. In this process, double-stranded RNA is cleaved into smaller fragments known as short-interfering RNA (siRNA), which then bind the RNA-induced silencing complex (RISC). Following integration into this protein complex siRNA is unwound into single strands. The guide strand binds to complementary mRNA and then is degraded by the RISC complex, preventing translation to a protein and silencing the target gene.

siRNA therapeutics are short sequences of double-stranded, non-coding RNA that are specifically designed to bind and induce degradation of mRNA sequences via endogenous RNAi pathways. This prevents translation of the target mRNA and effectively silences a disease-associated gene. They are under investigation as disease-modifying therapies in a wide range of disorders with a genetic cause or where excess protein production causes symptoms.

There are a few ways in which siRNA therapeutics differ from ASOs (antisense oligonucleotides). Firstly, siRNA therapeutics are a sequence of double-stranded, non-coding RNA whereas ASOs are single-stranded oligonucleotides. siRNA therapies and ASOs differ in their effect duration and potency, immunogenic and toxicity potential, as well as targeting and delivery mechanisms. In general, siRNA therapeutics are recommended for transient knockdown of cytoplasmic mRNA targets and are best suited for delivery to the liver or lungs. ASOs are recommended where targeting of nuclear, pre-mRNA is required, or where the target gene is expressed in the CNS or muscle, and a longer duration of action is required.

Several siRNA therapies have been approved by the FDA over the past decade, for various cardiovascular diseases and metabolic disorders affecting the liver:

• Patisiran and Vutrisiran have been approved for the treatment of polyneuropathy due to hereditary transthyretin amyloidosis (hATTR) and reduce production of the transthyretin protein in the liver.
• Lumasiran and Nedosiran have been approved for the treatment of primary hyperoxaluria and relieve symptoms by reducing urinary and plasma oxalate levels.
• Givosiran is a GalNAc conjugated siRNA-approved treatment for acute hepatic porphyria (AHP) that decreases production of heme and subsequent toxic buildup of porphyrin molecules.
• Inclisiran has been approved for the treatment of heterozygous familial hypercholesterolemia (HeFH) and atherosclerotic cardiovascular disease (ASCVD) and lowers low density lipoprotein (LDL) levels by inhibiting degradation of LDL receptors, and increases LDL removal from blood.