Split-and-pool combinatorial

Directed Sorting Split-and-Pool Combinatorial Synthesis... 

Combinatorial chemistry has moved from specially centralized laboratories, often equipped with multimillion-dollar robots, onto the bench of individual medicinal chemists. This change in direction requires the availability of personal chemistry tools that are simple to operate, easy to arrange in the laboratory, and reasonably priced. Such instruments are now available for the effective synthesis of combinatorial libraries. The Encore synthesizer represents a simple and efficient personal chemistry tool that allows the execution of directed split-and-pool combinatorial synthesis. The current version of the Encore synthesizer is designed for solid-phase synthesis on SynPhase Lanterns however, it can be modified for synthesis on alternative solid supports such as resin plugs from Polymer Laboratories (e.g., StratoSpheres Plugs). 

Fig. 7.1. The strategy used to develop GGTI. A pilot library consisting of 171 compounds were screened for the ability to inhibit GGTase-I using RhoA as a substrate. This led to the identification of two groups of compounds, one with tetrahydropyridine scaffold and the other with dihydropyrrole scaffold. Solid-phase split-and-pool combinatorial synthesis of a large number of analogs of these initial hits led to the identification of P3-E5 and P5-H6. More than 700-fold increase in IC50 value for the inhibition of GGTase-I was obtained in the case of P3-E5 compared with the initial compound. Further derivatization of P5-H6 and P3-E5 led to cell active compounds P61-A6 and P61-E7, respectively.

Using a split-and-pool combinatorial synthesis strategy, Tamanoi and coworkers developed a library of approximately 4000 heterocycles generated from resin-bound allenoates by means of phosphine catalysis [36,37]. This library was initially screened to identify GGTase I inhibitors (Table 8.1), and some potent GGTase I inhibitors with novel... 

Figure 8.7 Schematic representation of split-and-pool combinatorial nucleotide synthesis. An example of split-and-pool (split-and-combine) synthesis of trideoxyribonucleotides complementary to codons by the sohd phase phosphoramidite nucleotide synthesis is illustrated. Each cycle in the synthesis includes spht (equal molar spUt), react (deprotection, coupling with 4 protected nucleotides and oxidation) and combine steps. After 3 cycles of 4 reactions (corresponding to couphng reactions with 4 nucleotides per cycle), the final combined library consists of a mixture all 64 trinucleotides...

V. Krchnak, V. Padera, The encore technique a novel approach to directed split-and-pool combinatorial synthesis, in B. A. Bunin, G. A. Morales, (Eds), Methods in Enzymology, Vol. 369, Elsevier, 2003, pp. 112-124. 

Combinatorial approaches have been applied to this chemistry. In a method amenable to split and pool, PAL, or Rink resin, 89 is modified with an acetoacetate to generate the solid supported aminocrotonate 90. Either a two- or three-component Hantzsch protocol is followed to produce 91. Treatment with TFA carries out the cleavage from the resin and the cyclization to dihydropyridine 92. 

The concept of reducing the number of reaction vessels and exponentially increasing the number of synthesized compounds was brought to a next level of simplicity by the split-and-pool method of Furka et al.5 The split-and-pool method was independently applied by Lam et al.6 in a one-bead-one-compound concept for the combinatorial synthesis of large compound arrays (libraries) and by Houghten et al.7 for the iterative libraries. Now several millions peptides could be synthesized in a few days. In Furka s method the resin beads receiving the same amino acid were contained in one reaction vessel—identical to Frank s method—however, the beads were pooled and then split randomly before each combinatorial step. Thus the method is referred to as the random split-and-pool method to differentiate it from Frank s method in which each solid-phase particle was directed into a particular reaction vessel (the directed split-and-pool method). 

The third challenge was to integrate and/or automate the handling of individual particles between combinatorial steps. During the early years of directed split-and-pool methodology the entire process was done... 

The algorithm described above is for a three-step combinatorial synthesis. However, the method is not limited to only three-step combinatorial libraries the solid-phase support can be derivatized before the directed split-and-pool synthesis on the Encore synthesizer. The necklace coding can also be a very useful tool during the chemistry development process. 

The split-and-pool synthesis not only simplifies the complexity of the combinatorial synthetic process, but also offers additional important benefits. To undertake a full range of solid-phase chemical reactions, elaborate reaction conditions are needed for some chemical transformations. These include, but are not limited to, low temperature and inert atmosphere conditions. Parallel synthesis of a thousand compounds requires handling of a thousand reaction vessels. The timely addition of sensitive reagents (e.g., butyl lithium) at low temperature (—78°) under inert atmosphere during parallel synthesis is not a trivial task. It can be done if sophisticated automated synthesizer equipment is designed to handle and tolerate such reaction conditions. Such a synthesis can alternatively be performed easily in a manual fashion using a split-and-pool method that requires only a limited number of reaction vessels. Examples from Nicolaou s17 and Schrei-ber s18,19 laboratories have shown that the split-and-pool method is the methodology of choice for the synthesis of complex and diversity-oriented combinatorial libraries. 

When performing a synthetic combinatorial chemistry experiment, several basically different strategies may be followed to create a library of compounds. The most commonly used are mixelsplU (or split and pool) synthesis [1] masking strategies [15, 16] and parallel synthesis. In this chapter, the attention is focussed on the application of parallel synthesis to catalysis in the liquid phase. 

Although our main research interest has been in the field of heterogeneous catalysis, we have performed several proof of concept studies showing that FTIR imaging is flexible enough to be applied to the study of resin-supported combinatorial libraries, of the type commonly employed in split-and-pool syntheses. It was shown that the... 

Nicolaou KC, Pfefferkom JA, Mitchell HJ, Roecker AJ, Barluenga S, Cao GQ, Affleck RL, Lillig JE, Natural product-like combinatorial libraries based on privileged structures, 2, Construction of a 10,000-membered bcnzopyran library by directed split-and-pool chemistry using Nanokans and optical encoding, J. Am. Chem. Soc., 122 9954—9967, 2000. 

Approximately 100 mg of resin was distributed to each of the reaetion bloek wells (of an ACT block or a Bohdan block) by pipetting a slurry of the resin in DMF/DCM (3 1) or as dry resin into each IRORI kan. The peptides were then assembled by the combinatorial chemistry apparatus suited for parallel or split-and pool-synthesis (34) using in situ neutralization/HBTU activation protocols for BOC chemistry. The resin was initially washed with DCM and the BOC protecting group removed by washing twice with a 40% solution of TFA in DCM. 

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