Selection of DNA-encoded chemical libraries against endogenous membrane proteins on live cells

Affinity Selections of DNA‐Encoded Chemical Libraries on Carbonic Anhydrase IX‐Expressing Tumor Cells Reveal a Dependence on Ligand Valence

“…[34] Li and co-workers developed an elegant strategy to introduce target-specific DNA tags, which facilitated the enrichment of protein ligands by increasing the local effective molarity of the DEL library in proximity to the target. [33] In analogy, Seitz and colleagues developed a peptide nucleic acid (PNA) labelling strategy for membrane proteins [47] 10 12 copies each in 100 μL) for selections performed with CAIX-expressing CT26.3E10 and CAIX-negative CT26.wt (selection duplicates). AAZ-DNA was annealed to short 3'-modified DNA, allowing to display one (monovalent, left) or two AAZ molecules (bivalent, right).…”

“…[14] Cell-based selections have been reported for certain types of encoded libraries (e. g. phage display antibody libraries) [26][27][28][29] and have recently been described also for PNA- [30] and DNAencoded chemical libraries. [31][32][33][34] In principle, cells would provide a natural environment to perform selections against membrane proteins near their physiological context. Cell-based selections have led to the discovery of new ligands to the NK3 tachykinin receptor, [31] to the δ opioid receptor and to intracellular proteins as CBX7 ChD [32] or p38α, [34] to the folate receptor and to the epidermal growth factor receptor.…”

“…Cell-based selections have led to the discovery of new ligands to the NK3 tachykinin receptor, [31] to the δ opioid receptor and to intracellular proteins as CBX7 ChD [32] or p38α, [34] to the folate receptor and to the epidermal growth factor receptor. [33] The reports highlight that target-specific DNA tagging, [33] covalent cross-linking, [32] cellular binder trap enrichment [34] and iterative selections at high library inputs with cells that strongly overexpress the target protein [31] may facilitate the recovery and discovery of target-binding DEL members in cell selections. A general investigation of parameters that may influence cell selections has been reported for phage display systems [35] but not for DELs.…”

Recent Advances on the Selection Methods of DNA‐Encoded Libraries

“…In our experience, membrane proteins typically only have low-or sub-nanomolar effective molarities. [80] Both target specificity and abundance may be addressed by protein overexpression, which not only increases target concentration, but also can be compared with the control selection without overexpression to identify target-specific ligands (Figure 5b). The Bradley group pioneered live-cell-based selection of encoded chemical libraries.…”

“…Recently, Li and co-workers developed a method that can select DELs against endogenous membrane proteins without overexpression or genetic tagging. [80] They addressed the target specificity and abundance issues by installing a DNA tag to the target prior to selection (Figure 5e). On the one hand, the DNA tag provides target specificity by guiding DEL hybridization; on the other hand, it boosts the ligand affinity through the avidity effect, thereby driving the binding equilibrium and retaining the binders on the cell surface.…”