Solid-Phase Synthesis and Combinatorial Technologies

9.1 PHARMACEUTICAL APPLICATIONS 433

screenings plus any other available data. These data come from the library quality control, performed after the synthesis, from the HTS of a high-quality pharmaceuticalfocused library, and from further profiling of some of its members. When the library is tested on many biological targets, the stored data become invaluable in judging the library usefulness and maybe also in performing related combinatorial efforts.

A search on the integrated structural/relational database system downloads each stored piece of information. Browsing the library, extracting SAR data related to structural subclasses, and pulling out library subsets to be tested in future screenings are the main goals of such a search. Proper data management of combinatorial and screening results is, and will become ever more, crucial to maximize the return of synthetic and biological efforts toward the discovery of novel drugs. A few excellent reviews have covered the issues related to data management of combinatorial libraries and HTS (40–46), and the reader should consult them for a more detailed picture of this relevant topic.

9.1.7 An Example: Synthesis of Chalcone-Based, Druglike Libraries

Powers (47) reported the synthesis in solution of a set of modular, discrete libraries containing >74,000 members based on the combinatorialization of a common intermediate scaffold (see also Sections 4.2.2 and 8.2.6). A core chalcone library L1 was modified (libraries from libraries) (48), producing libraries L2–L10 and expanding both the number and diversity of the compounds due to the smaller sets of different scaffold-derived libraries (vide infra).

Aldol condensation of acetophenones (M1, 32 representatives) with aldehydes (M2, 40 representatives) produced a high-quality core chalcone array L1 (Fig. 9.8), with an excellent 96% average yield. The monomer structures are reported in Fig. 9.9. Combination of aromatic or heteroaromatic mono-, di-, or trisubstituted methyl ketones M1 with aromatic or heteroaromatic monoor disubstituted aldehydes M2 produced 1280 chalcones with suitable, druglike MWs (452 > MWs > 186) and lipophilicity.

Condensation of L1 with various reagents to produce disubstituted five-member heterocycles was envisaged (Fig. 9.10). Reaction with hydroxylamine 9.11 provided

a: NaOH, EtOH/H2O 4/1, rt, 16 hrs.

Figure 9.8 Synthesis of the solution-phase discrete chalcone library L1.

R3 = H, 4-OMe, 4-F, 4-iPr,

3-CF3, 3,4-(CH2)3

a: NaOH, EtOH, 80°C, 12 hrs; b: various reaction conditions; c: NaOH, EtOH, 70°C, 8 hrs.

Figure 9.10 Solution-phase discrete five-membered heterocyclic libraries L2–L3 obtained from the solution-phase discrete chalcone library L1.

Trisubstituted six-member carbocycles were also prepared. Condensation of L1 with commercial acetoacetanilides 9.14a–f produced, by one-pot Michael addition and Robinson annulation, the cyclohexenone library L4 (7680 members, Fig. 9.11). Noncommercial acetoacetamides M1, prepared by condensation of diketene 9.15 with 40 primary and secondary amines (structures not shown in the paper), were used as monomers and condensed with a 320-member L1 subset to give the expanded cyclohexenone library L5 (12,800 members). Difficulties in preparing extremely large discrete libraries, even using fully automated robotic workstations, and the need to diversify the screening set obliged the authors to limit the number of monomers and chalcones for any specific library/scaffold (see also L10 below).

Polysubstituted dihydropyridines were also considered as targets. Condensation of L1 with enamino ester 9.16 and enamino nitrile 9.17 under standard Hantsch conditions only proved successful for the latter, furnishing the pyridine library L6 (1280 members). Oxidation of the initially formed dihydropyridines is presumably favored due to the highly conjugated system formed (Fig. 9.12). Hantsch condensation with cyclic enamino esters 9.18 and 9.19a–f was also successful, providing respectively the bicyclic libraries L7 (1280 members) and L8 (7680 members, Fig. 9.12).

Finally, two polycyclic systems were built. First, the cyclic guanidine embedded into the 2-aminobenzimidazole nucleus 9.20a–f was used to build the tricyclic library L9 (7680 members, Fig. 9.13); then a complex spiro-polyheterocycle library L10

436 APPLICATIONS OF SYNTHETIC LIBRARIES

Figure 9.11 Solution-phase discrete six-membered heterocyclic libraries L4–L5 obtained from the solution-phase discrete chalcone library L1.

(25,600 members) was prepared by one-pot condensation of isatins (M1, 16 representatives), α-amino acids (M2, 20 representatives), and a chalcone subset (M3, 80 representatives from L1, Fig. 9.13). The monomers for the synthesis of L10 were again selected to maximize the library diversity and to produce a druglike profile for the library individuals to be considered as drug candidates.

The screening set of roughly 75,000 compounds was characterized analytically. Twenty-five percent of the compounds were submitted to HPLC/MS using ELSD, producing average purities of >85% for each sample. The generic libraries L1–L10 and their assessed synthetic schemes could easily be expanded using larger sets of diversifying monomers in a more focused attempt, if one or more hits were found from a specific library, without significant efforts.

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R3

and other 5-, 6- and 7-member cycles

WO9815532 (1998)

Novartis

Generic solid-phase synthesis of heterocycles was claimed

A common precursor to all libraries was described and claimed

Several relevant documents found by the International Search Report

Figure 9.14 Patents regarding scaffold-based combinatorial libraries: main features and claims.

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A few additional comments can be made. Many combinatorial patents have been filed by small biotechnology companies to let the scientific world appraise their libraries (or technologies), rather than to really protect them. To date, major pharmaceutical companies have not patented to a large extent in the field. The more conservative approach of patenting well-characterized, more downstream compounds is still prevalent. The time lag between first filing and patent publication may disprove such precepts, though, in the near future. Most accessible patent applications have prior art,

Figure 9.15 Patents regarding technology-based combinatorial libraries: main features and claims.

442 APPLICATIONS OF SYNTHETIC LIBRARIES

which was discovered during the International Examiners’ Search Report, and their relevance should be strongly affected. No major litigations concerning claimed libraries or technologies have appeared yet, but such events are likely to appear in the near future (see, e.g., the structures and claims of Figs. 9.14 and 9.15).

Some recent articles (49–55) have reviewed the field of combinatorial technologies patenting. Any combinatorial scientist must know that current and future intellectual property is going to be influenced by technological breakthroughs and by the increase in popularity of combinatorial technologies in many other fields of application.

9.1.10 An Example: Exploration of Structure–Activity Relationship of a Peptidomimetic Hit via Combinatorial/Medicinal Chemistry

Bhandari et al. (56) modified a hit structure derived from the primary screening of various libraries (>300,000 compounds) on the zinc metalloenzyme phosphomannose isomerase from the yeast Candida albicans (CaPMI) to find enzyme inhibitors as potential antifungal agents. During primary screening only a 1296-member SP dipeptide pool library (L11, Fig. 9.16) showed activity on the enzyme. Its deconvolution and analytical characterization led to the discovery of a by-product, derived from incomplete coupling, that showed activity on the enzyme. This compound (9.21, Fig. 9.16) showed a weak inhibitory activity on CaPMI (IC50 = 40 M) and was selected for further chemical profiling.

At first, substituted phenoxybutyric, phenoxyacetic, and phenylacetic acids (M1, 66 representatives, Fig. 9.17) were used to cap the aminoindane carboxamide 9.22 in solution to expand the capping residue diversity, producing the discrete array L12. Only three chlorosubstituted phenoxybutyric acids produced active compounds 9.23– 9.25, with the 3,4-diclorophenoxy substitution pattern as in 9.25 being found to be optimal (Fig. 9.17). This initial exploration was enhanced by using the SP synthetic route reported in Fig. 9.18. Rink-supported aminoindane carboxamide 9.26 was coupled with bromobutyric acid; then the resulting resin-bound bromide 9.27 was

SP pool library

36 pools containing 36 compounds

Figure 9.16 Structure of the primary SP peptidomimetic library L11 and of the deconvoluted hit 9.21 active on C. albicans phosphomannose isomerase (CaPMI).