The development of a new selection system for phage display and other combinatorial libraries is proposed. Conventional affinity selection techniques contain an intrinsic logical contradiction. They are intended to select the best binding ligands to a given ligate, but the retrieval of the best binders is based upon breaking their affinity interaction with ligate. Thus, selection is actually performed by two criteria: (1) ligands should have affinity levels that are high enough to stay bound during washing cycles but (2) low enough to enable their elution. Such selection system favors species with moderate affinity rather than the ones with the highest affinities that a library possibly has to offer. Carbene-generating cleavable photocrosslinkers can serve as detachable anchors capable of covalent insertion into non-reactive plastics. We propose to apply these reagents for the reversible immobilization of ligates on non-adsorptive, low-background supports. The resulting selection system will include conventional washing and elution steps, but only as a means to eliminate non-specific and moderately specific ligands. The retrieval of the highest specificity ligands will not be based on breaking of their affinity interaction with ligate. Instead, cleavage in mild conditions of a specific bond incorporated in the anchor will """"""""de-immobilize"""""""" the ligate together with the most specific ligands. The proposed approach is expected to deliver ligands with the affinity levels unmatched by currently used combinatorial selection techniques. This will bring new developments in many medical applications, particularly cancer diagnostics and treatment, by providing enhanced sensitivity and reliability of early detection assays, better drugs for chemotherapy and better targeting systems for drug delivery.
Screening of combinatorial libraries is a powerful method widely used for drug discovery, molecular markers research and molecular targeting (gene therapy, drug delivery, etc.). The proposed approach is expected to dramatically simplify and accelerate the selection process and to bring the affinity and specificity of resulting ligands to a new level. We anticipate a profound commercial interest to the proposed system (reagent kits, custom- modified plastic supports and complete library enrichment services) from major pharmaceutical companies and research centers.
