Spatially addressable parallel libraries

Geysen et al. [1] introduced this method for generating peptides in a reusable form. In this method, peptides are synthesized on polyaciylate-grafted polyethylene pins arranged in standard 96-well microtiter plate format, and screening is performed by an enzyme- 

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In essence only two approaches exist to generate combinatorial peptide libraries biologic and synthetic library approaches. According to the different techniques used, the synthetic library approach can be divided additionally into five methods 1) the spatially addressable parallel library method (1, 4, 8),... 

An 8000-member library of trisamino- and aminooxy-l,3,5-triazines has been prepared by use of highly effective, microwave-assisted nucleophilic substitution of polypropylene (PP) or cellulose membrane-bound monochlorotriazines. The key step relied on the microwave-promoted substitution of the chlorine atom in monochlorotriazines (Scheme 12.7) [35]. Whereas the conventional procedure required relatively harsh conditions such as 80 °C for 5 h or very long reaction times (4 days), all substitution reactions were found to proceed within 6 min, with both amines and solutions of cesium salts of phenols, and use of microwave irradiation in a domestic oven under atmospheric reaction conditions. The reactions were conducted by applying a SPOT-synthesis technique [36] on 18 x 26 cm cellulose membranes leading to a spatially addressed parallel assembly of the desired triazines after cleavage with TFA vapor. This concept was later also extended to other halogenated heterocycles, such as 2,4,6-trichloropyrimidine, 4,6-dichloro-5-nitropyrimidine, and 2,6,8-trichloro-7-methylpurine, and applied to the synthesis of macrocyclic peptidomimetics [37]. 

Combinatorial libraries are prepared by the (1) parallel synthesis of arrays, (2) split-pool method, (3) biological method, or (4) spatially addressable parallel synthesis [74,78-80]. Parallel synthesis is carried out by the simultaneous synthesis of an array of different compounds. Several methods are available. In the multipin method, the peptide synthesis is carried out on polyethylene rods that have attached protected amino acids [81]. The amino acid sequence of a synthesized peptide on a particular pin depends on the order in which the amino acids are added. The number of products synthesized is the same as the number of pins. Another version of parallel synthesis, known as the teabag method, uses resin-filled bags in place of pins [74]. By pooling the resin portions from the appropriate bags, followed by redistribution and further coupling with a specific amino acid, a peptide library can be synthesized. The SPOT method uses a cellulose paper membrane as a solid support, which acts as an open reactor. Respective reagent solutions are pipetted onto several spots to synthesize as many peptides as the spots chosen [74,82]. 

In this chapter we will discuss current approaches for analytical characterization of combinatorial libraries in a pharmaceutical industry environment. Recently, several analytical groups have presented very similar strategies for analysis of libraries [7-9]. As will be shown later, the key to successful analytical characterization of a combinatorial library is to perform analytical and chemical work in parallel with the library development. The accumulation of data and analytical experience during this process results in an assessment of library quality with a high level of confidence, even if as little as 5-10% of the library components are analyzed. Utilization of the strategy will be demonstrated using two examples analysis of a library synthesized on a robotic station in spatially addressed format and analysis of a library synthesized in accordance with split-and-mix technology. 

A special case of parallel synthesis is the spatially addressable synthesis pioneered by Fodor et al. [17,18] in 1991. Here, each member of the library is synthesized at a specific location on a functionalized silica wafer rather than on resin beads in separate reaction vessels. This approach, based initially upon solid-phase peptide synthesis and semiconductor photolithographic techniques by using photolabile amino protecting groups, allows the synthesis of combinatorial libraries containing about 50000 compounds localized to a 50 pm square site on a silica wafer ( library on a chip ). 

According to Blackwell [103] the application of microwave irradiation to expedite solid-phase reactions could be the tool that allows combinatorial chemistry to deliver on its promise - providing rapid access to large collections of diverse small molecules. Several different approaches to microwave-assisted solid-phase reactions and library synthesis are now available. These include the use of solid-supported reagents, multi-component coupling reactions, solvent-free parallel library synthesis, and spatially addressable library synthesis on planar solid support. 

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