Article Figures & Data
Figures
- Fig. 1.
Screening cascade for chemical cocktails, SPECS compounds, and microbial broths. Schematically shown are the procedures and selection criteria leading from primary screening to in vitro testing and the start of a medicinal chemistry program. First, the inhibitory activity should be above a defined value (e.g., ≥40% inhibition at 50 μM) and must be confirmed; second, the structure must be of interest (acceptable molecular weight; lipophilicity) and have a potential for chemical variations; third, the identified hit would, preferentially, have in vitro activity.
- Fig. 2.
(A) The monoquinoline-derived hypothesis superimposed on chloroquine. (B) Fit of the monoquinoline-derived hypothesis for Ro 06-8463, which was found by 3-D database screening (activity for the resistant strain K1, 0.3 μM; activity for the sensitive strain NF54, 0.026 μM). Feature definitions: blue, hydrophobic; green, hydrogen bond acceptor; cyan, hydrogen bond donor; red, positive ionizable.
Tables
- Table 1.
Effects of substances possibly interfering with the hematin polymerization reaction
Class and compound Concn % Inhibition IC50 Concn tolerated Cations and anions NaCl 100 mM 10 KCl 100 mM 10 NH4Cl 100 mM 14 MgCl2 100 mM 16 CaCl2 100 mM −15 Na2HPO4 100 mM 26 NaH2PO4 100 mM 14 Detergents Tween 80 1% 88 0.14% 0.003% Triton X-100 1% 37 SDS 1% 46 Chelating agents EDTA 10 mM −19 EGTA 10 mM −10 Tannins Catechin 100 μg/ml 11 Gallic acid 100 μg/ml −43 Tannic acid 100 μg/ml 100 34 μg/ml 3 μg/ml Pigments Chlorophyll a 100 μg/ml 33 Chlorophyll b 100 μg/ml 28 Melanin 100 μg/ml 40 Saponin (saikosaponin A) 100 μg/ml −9 Antibiotics Daunomycin 100 μg/ml −27 Amphotericin B 100 μg/ml 4 Penicillic acid 100 μg/ml −19 Actinomycin D 100 μg/ml 37 SH-group reacting agents DTT 100 μg/ml 89 5.1 μg/ml 1 μg/ml Cysteine 100 μg/ml −13 GSH 100 μg/ml 41 Fatty acids Stearic acid 100 μg/ml 11 Oleic acid 100 μg/ml 30 Linoleic acid 100 μg/ml −15 Proteases Actinase E 10 μg/ml −13 S peptidase 10 μg/ml −29 Proteinase K 10 μg/ml −6 Organic solvents MeOH 10% −29 EtOH 10% −28 Acetonitrile 10% −40 DMSO 10% −37 - Table 2.
Results of tests for inhibition of hematin polymerization
Chemical source of compounds No. of compounds in primary screening No. (%) of hits in: No. (%) of active single compounds Primary screening Reassay Roche cocktails 11,887 268 (2.3) 69 (0.6) 38 (0.3) NCC 97 16 (16) 16 (16) 5 (5.2) SPECS 15,760 36 (0.2) 13 (0.1) Combinatorial chemistry libraries 1,380 0 Microbial broths Actinomycetes 6,000 23 (0.4) 3 (0.1) 0 Fungi 6,320 42 (0.7) 17 (0.3) 10 (0.2) Bacteria 2,320 13 (0.6) 4 (0.2) 1 - Table 3.
Classes of compounds identified as inhibitors of the hematin polymerization reaction in HTS
Compound class Structure IC50 (μM) in hematin polymerization assay IC50 (μM) against P. falciparum growth in culture % Inhibition ofP. berghei growth in vivoa IC50 (μM) in cytotoxicity assayb K1 NF54 Chloroquine (Ro 01-6014) 
80 0.45 0.0125 100 NT Triarylcarbinol (Ro 06-9075) 30 0.97 1.16 98.4 4.66 Piperazine (Ro 10-3428) <100 0.21 0.27 Inactive 4.45 Miscellaneous (Ro 14-3955) 24 1.93 2.7 Inactive 63.3 Benzophenone (Ro 22-8014) 24 1.32 1.78 88 NT Imide (Ro 20-1120) 13 16 16 NT NT Hydrazide (Ro 04-4410) 19 15.4 15.4 Inactive NT Indole (Ro 90-2341) 14 28.5 28.5 NT NT Isoxazole (Ro 90-0987) 12 28.5 28.5 NT NT - Table 4.
Selection of the most active compounds found by 3-D database screening
Compound structure Roche no. IC50 (μM) in hematin polymerization assay IC50 (μM) against P. falciparum growth in culture NF54 K1 Ro 06-8463 28 0.026 0.3 Ro 06-9075 30 1.16 0.97 Ro 10-3428 <100 0.27 0.21 Ro 10-8995 100 0.14 0.29 Ro 47-3191 NTa 0.46 0.3 ↵a NT, not tested.
