Timothy Weeden, Senior Director, Head of Platform Development, Dyne Therapeutics
Timothy Weeden from Dyne Therapeutics presented their work on FORCE platform for the targeted delivery of oligonucleotides and the efficiency of their candidate DYNE-101 in the treatment of muscle diseases. In the first part of the talk, he introduced the FORCE platform and its main components. In the second part, he explained the applications and modularity of FORCE in the treatment of Duchenne Muscular Dystrophy (DMD) and Myotonic Dystrophy type 1 (DM1). Weeden stated that the FORCE platform had three essential components: fragment antibody, linker, and payload. The fragment antibody (Fab) was used to target transferrin I receptor (TfR1). The clinically-validated linker region was required for precise conjugation of the payload to the Fab. The payload was developed to target the genetic basis of the disease. He shared the advantages of FORCE platform over the use of naked oligo and AAV-based methods such as targeted delivery in muscle, durability, safety profile, the use of a titratable and redosable strategy, including scalable and well-established manufacturing processes. Their strategy was to target transferrin Receptor 1 which is a feasible target thanks to its ideal expression and recycling rate for muscle delivery. TfR1 is a transmembrane protein which is essential for iron mediated uptake in cells. They target TfR1 by developing an antibody fragment rather than monoclonal antibodies. Because chronic dosing strategy is applied, monoclonal antibodies that have been shown to downregulate TfR1 would not be an ideal candidate. Other advantages of the Fab developed by their team are the selectivity for TfR1 over TfR2, cross reactivity across species, efficient delivery, high stability and manufacturability, and absence of an interference with iron homeostasis.
The second component is the linker which confers the stability in serum and endolysosomal release of the payload and the proper conjugation of the payload to the Fab. Weeden shared a relevant data showing the efficacy of Val-Cit dipeptide linker in four different commercial antibody-drug conjugates (ADCs) across four different species indicated that more than 90% of parental drug measured at 72 hrs. The final component is the payload which are oligonucleotides in this study. They focused on antisense oligonucleotides (ASO) and siRNAs acting through different mechanisms of action. siRNAs show their activity in the cytoplasm through RISC loading and binding its target which triggers mRNA cleavage while ASOs lead to downregulation either by causing RNase1 cleavage of target mRNA or via splice modulation resulting exon skipping and the translation of a truncated yet still functional form of the protein.
Next, Weeden discussed the application of FORCE platform in the treatment of DM1 and DMD and shared preclinical efficacy results in disease models and muscle delivery studies. DM1 is a triplet repeat disease associated with mutation in DMPK (DM1 protein kinase) gene causing the expansion of CTG repeat in the 3’UTR region of the gene. FORCE has been designed to target toxic nuclear DMPK RNA to correct spliceopathy by degrading toxic RNA within the foci. They have developed two in vitro and an in vivo model with different size of expansion representing the patient population with DM1 to test their drug candidates. They assessed knockdown and splicing correction by quantifying mRNA expression via qPCR and foci reduction via FISH and showed that DMPK mRNA expression and DMPK foci were reduced while mis-splicing was corrected upon DYNE-101 administration.
As an in vivo model, they utilized a mouse model developed by crossing a mouse expressing human TfR1 with another expressing toxic human DMPK including 1000 CTG repeats. They demonstrated a significant reduction of DMPK expression as well as splicing correction in different skeletal muscles. They also showed that DYNE-101 administration resulted in efficient knockdown and well-tolerated in non-human primates (NHPs).
Weeden continued his talk by focusing on splice modulation for the treatment of Duchenne muscular dystrophy (DMD) which is associated with the mutations in the gene encoding dystrophin. Missing exons in DMD pre-mRNA induce frameshift and early stop codon formation resulting in the absence of functional dystrophin expression. Phosphorodiamidate morpholino oligomer (PMO) treatment via FORCE platform for exon skipping resulted in truncation of pre-mRNA allowing the expression of truncated but functional dystrophin protein.
Pharmacokinetic profiling studies on mouse model lacking dystrophin expression indicated a rapid uptake of ASOs into muscle tissue, a slightly delayed exon skipping, and a further delay in protein production. They also demonstrated the robustness and the durability of the strategy by western blot and dystrophin-positive fibers via fluorescent microscopy analysis. They compared the delivery efficiency of naked ASOs and their candidate DYNE-251 in NHPs and indicated that they achieved over 40 times greater amount of PMO in the muscle tissue of the animals. They also demonstrated safety profile of their candidate in NHPs and promising results reaching up to 52% exon skipping efficiency in diaphragm. Weeden explained that the FORCE platform enables targeted delivery of therapeutic oligonucleotides to muscle with considerable efficiencies demonstrated in in vivo and in vitro studies. They achieved efficient target modification across preclinical disease models for DM1 and DMD, and were able to target and correct toxic and dysfunctional proteins produced as a result of mutations. Additionally, enhanced muscle delivery, target engagement and a favorable safety profile have been shown in various studies.
