Positional scanning synthesis

Figure 7.22 Positional scanning synthesis of the four h)q)othetical subUbraries 1-4.

Because of their ease of synthesis and their structural similarity to peptides, many laboratories have used peptoids as the basis for combinatorial drug discovery. Peptoids were among the first non-natural compounds used to establish the basic principles and practical methods of combinatorial discovery [17]. Typically, diverse libraries of relatively short peptoids (< 10 residues) are synthesized by the mix-and-split method and then screened for biological activity. Individual active compounds can then be identified by iterative re-synthesis, sequencing of compounds on individual beads, or indirect deduction by the preparation of positional scanning libraries. 

Backes BJ, Harris JL, Leonetti F, et al. Synthesis of positional-scanning hbraries of fluorogenic peptide substrates to define the extended substrate specificity of plasmin and thrombin. Nat Biotechnol 2000 18 187-93. 

Experimental Procedure for the Parallel Synthesis of Heterocyclic Positional Scanning Libraries 47 to 51... 

Although not identical, both the orthogonal and positional scan formatted libraries share the features that all mixtures are made at the start of the library process and only individual compound synthesis is required after the first screening of mixtures. This is an extra initial effort with regard to the synthesis of mixtures when compared to an iterative method. The advantage is that no intermediate mixture syntheses will be required. If prepared in sufficient quantity, the library can be screened over a large number of assays, and the added effort of initial mixture syntheses will be translated into an efficiency in deconvolution relative to the continual resynthesis of mixtures with iterative deconvolution. 

These techniques can be broadly split into two groups, the first of which can be represented by pooling methods, where deconvolution is obtained via various chemical steps, run in parallel or after the library synthesis. Pooling methods normally require multiple synthesis of many library members, including inactive individuals, in different pool formats. They are not single bead methods, so they are independent from analytical methods for structure determination. This group includes iterative deconvolution, recursive deconvolution, subtractive deconvolution, positional scanning and mutational... 

Positional scanning, or 1-D indexing, was reported as a deconvolutive technique by Houghten and coworkers [35] and subsequently applied extensively to solid-phase peptide library synthesis. An example of solid-phase small organic molecule 2-D indexed library synthesis and deconvolution was reported by Berk et al. [36]. Description of this strategy, called spatially arrayed mixture (SpAM), and a comparison with classical positional scanning, are shown in Figure 8.9. 

Positional scanning was repeatedly used by Houghten and coworkers [38] in solid-phase peptide synthesis, but few examples of its application to small molecule libraries have been reported. Smith et al. [39] deconvoluted a solid-phase library of 1600 esters/amides, Andrus et al. [40] deconvoluted a solid-phase library of non-natural polyenes, Leone-Bay et al. [41] deconvoluted the activity of an indexed library in an in vivo assay, while Pirrung et al. reported both a solution phase 54-member carbamate library [42] and a solution-phase 72-member tetrahydroacridine library [43]. 

Boger et al. [47] introduced the so-called deletion synthesis deconvolution as a process typical for convergent dimerization libraries, based on the methodical elimination of one element from the variable library units and on the evaluation of loss vs. gain of activity. This method was compared with positional scanning in a following paper by the same group [48] and proved to be effective and useful. 

Boger DL, Lee JK, Goldberg J, Jin Q, Two comparisons of the performance of positional scanning and deletion synthesis for the identification of active constituents in mixture combinatorial libraries, J. Org. Chem., 65 1467-1474, 2000. 

Synthesis of a Scaffolded Positional-Scanning Peptide Library Bnilt on a Bicyclic Peptide Template )... 

A positional scanning combinatorial library (Fig. 3) can be used in place of iterative synthesis and screening [9]. In this approach, all library mixtures are synthesized and assayed for activity. The most active mixture from each positional library reveals the preferred residue at each position. Because all possible combinations of residues are represented in one of the sublibraries, the approach should be insensitive to synergistic effects associated with the structure. The orthogonal combinatorial library consists of two different libraries made up of the same compounds and synthesized in a way that mixtures from each library will only share one peptide or subunit [10]. In this way, these libraries are nonoverlapping, or orthogonal. 

A process called deconvolution is commonly employed to determine biological activity.The final library pools are not combined but are tested as either on-bead or detached compound mixtures. The most active pool defines which synthon is preferred in the last step. The synthesis is repeated to the penultimate set of pools and these are then allowed to react with the best last step synthon. Alternatively, pools of conserved resin from the penultimate step held back during the original synthesis may be used. The most active pool found on retesting defines the best last two synthons. This process is repeated until the most active member is identified. A somewhat similar method termed position scanning also has found successful application, especially in the analysis of peptides. ... 

However, if in one position there is no clear preference for a definitive substituent, all combinations of the preferred substituent must be synthesized in order to find the most active compound. Although active compounds are identified without iterative synthesis using positional scanning, in comparison with iterative deconvolution there is an increased likelihood that the most potent compound(s) will not be identified [110,111]. 

The advantage of positional scanning over iterative deconvolution is that the sublibrary syntheses are carried out at once. But the number of split and pool operations that would be required for library synthesis is rather high so that building block mixtures are used in several of the library synthesis steps in order to reduce the number of operations. A thorough examination of the theoretical and experimental aspects of iterative and positional scanning deconvolution has been published [110],... 

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