Liver‐Targeted Small‐Molecule Inhibitors of Proprotein Convertase Subtilisin/Kexin Type 9 Synthesis

Abstract

Targeting of the human ribosome is an unprecedented therapeutic modality with a genome‐wide selectivity challenge. A liver‐targeted drug candidate is described that inhibits ribosomal synthesis of PCSK9, a lipid regulator considered undruggable by small molecules. Key to the concept was the identification of pharmacologically active zwitterions designed to be retained in the liver. Oral delivery of the poorly permeable zwitterions was achieved by prodrugs susceptible to cleavage by carboxylesterase 1. The synthesis of select tetrazole prodrugs was crucial. A cell‐free in vitro translation assay containing human cell lysate and purified target mRNA fused to a reporter was used to identify active zwitterions. In vivo PCSK9 lowering by oral dosing of the candidate prodrug and quantification of the drug fraction delivered to the liver utilizing an oral positron emission tomography 18F‐isotopologue validated our liver‐targeting approach.

First Food-Derived Peptide Inhibitor of the Protein–Protein Interaction between Gain-of-Function PCSK9D374Y and the Low-Density Lipoprotein Receptor

“…There is therefore an increasing demand for small molecules as PCSK9 inhibitors. 12 After the demonstration that lupin protein hydrolysates are able to inhibit the PCSK9−LDLR binding, 7 the following research was aimed at elucidating the structures of at least some active peptides out of those complex hydrolysates. Using a multidisciplinary approach combining absorption experiments performed with differentiated Caco-2 cells as an in vitro intestinal model 13 and in silico molecular modeling, 7 it was possible to identify two active peptides, i.e., LILPKHSDAD (P5) and GQEQSHQDEGVIVR (T9), derived from the hydrolysis of β-conglutin with pepsin and trypsin, respectively.…”

Small Molecule Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9) Inhibitors: Hit to Lead Optimization of Systemic Agents

“…In consideration of these toxicology findings, which were not evident in the upfront CellTiter-Glo ATP viability assay, in vitro bone marrow cellular toxicity assays for rat (initial toxicology species, rat lineage (−) cells) and human (CD34 + cells) 19 were developed to screen for future compounds devoid of bone marrow effects. Indeed, when 7f was assessed in the rat lineage (−) assay, it reduced cell viability (IC 50 = 1.2 μM) within 3-fold margin of its functional activity (PCSK9 Pro-Luc IC 50 = 0.38 μM).…”

“…Therefore, we sought to understand the in vitro selectivity profile of 7l versus the previously profiled 7f and assess its in vivo activity. Of note, using the previously described ribosome binding assay that employs [ 3 H]-7f as the radioligand, 18,19 7l was found to bind to the 80S isolated ribosomes more weakly than 7f (Table 4) in spite of comparable activity for the inhibition of PCSK9.…”

“…18 In parallel, efforts to achieve a liver-selective PCSK9 inhibitor via a pro-drug strategy were also explored. 19 Herein, we disclose our medicinal chemistry efforts to transform 1 into molecules with high systemic bioavailability and an improved balance of safety and efficacy suitable for further exploration.…”

Overcoming the Challenges of Making a Single Enantiomer N-1 Substituted Tetrazole Prodrug Using a Tin-Mediated Alkylation and Enzymatic Resolution

“…2 However, discovery of a small molecule PCSK9 inhibitor with the possibility to deliver it by the more convenient oral route has lagged behind the mAbs. Previous disclosures from the Pfizer laboratories have described small molecule protein translation inhibitors of PCSK9 such as PF-06446846 (1), 3,4 PF-07556769 (2), 5,6 and PF-06815345 (3; Figure 1). 6 Since PCSK9 is synthesized primarily in the liver, the mode-of-action of the aforementioned inhibitors makes them well-suited for application of a tissue-targeted delivery approach.…”