Next Generation SICLOPPS Screening for the Identification of Inhibitors of the HIF-1α/HIF-1ß Protein-Protein Interaction.

Split-intein circular ligation of proteins and peptides (SICLOPPS) is a method for generating intracellular libraries of cyclic peptides that has yielded several first-in-class inhibitors. Here, we detail a revised high-content, high-throughput SICLOPPS screening protocol that utilizes next-generation sequencing, biopanning, and computational tools to identify hits against a given protein-protein interaction. We used this platform for the identification of inhibitors of the HIF-1α/HIF-1ß protein-protein interaction. The revised platform resulted in a significantly higher positive hit rate than that previously reported for SICLOPPS screens, and the identified cyclic peptides were more active in vitro and in cells than our previously reported inhibitors. The platform detailed here may be used for the identification of inhibitors of a wide range of other targets.

Genetically Encoded Cyclic Peptide Libraries: From Hit to Lead and Beyond.

With the increasing utilization of high-throughput screening for lead identification in drug discovery, the need for easily constructed and diverse libraries which cover significant chemical space is greater than ever. Cyclic peptides address this need; they combine the advantageous properties of peptides (ease of production, high diversity, high potential specificity) with increased resistance to proteolysis and often increased biological activity (due to conformational locking). There are a number of techniques for the generation and screening of cyclic peptide libraries. As drug discovery moves toward tackling challenging targets, such as protein-protein interactions, cyclic peptide libraries are expected to continue producing hits where small molecule libraries may be stymied. However, it is important to design robust systems for the generation and screening of these large libraries, and to be able to make sense of structure-activity relationships in these highly variable scaffolds. There are a plethora of possible modifications that can be made to cyclic peptides, which is both a weakness and a strength of these scaffolds; high variability will allow more precise tuning of leads to targets, but exploring the whole range of modifications may become an overwhelming challenge.