Abstract
The Plasmodium falciparum cysteine protease falcipain-2 (FP-2) is an attractive antimalarial target. Here, we discovered that the natural compound NP1024 is a nonpeptidic inhibitor of FP-2 with an IC50 value of 0.44 μmol L−1. The most exciting finding is that both in vitro and in vivo, NP1024 directly targets FP-2 in malaria parasite-infected erythrocytes as a natural fluorescent probe, thereby paving the way for an integration of malaria diagnosis and treatment.
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References
Aneja, B., Kumar, B., Jairajpuri, M.A., and Abid, M. (2016). A structure guided drug-discovery approach towards identification of Plasmodium inhibitors. RSC Adv 6, 18364–18406.
Bloor, S.J. (1995). An antimicrobial kaempferol-diacyl-rhamnoside from Pentachondra pumila. Phytochemistry 38, 1033–1035.
Conroy, T., Guo, J.T., Elias, N., Cergol, K.M., Gut, J., Legac, J., Khatoon, L., Liu, Y., McGowan, S., Rosenthal, P.J., et al. (2014). Synthesis of gallinamide A analogues as potent falcipain inhibitors and antimalarials. J Med Chem 57, 10557–10563.
Ettari, R., Bova, E., Zappala, M., Grasso, S., and Micale, N. (2010). Falcipain-2 inhibitors. Med Res Rev 30, 136–167.
Fujita, T., Sezik, E., Tabata, M., Yesilada, E., Honda, G., Takeda, Y., Tanaka, T., and Takaishi, Y., (1995). Traditional medicine in Turkey VII. Folk medicine in middle and west Black Sea regions. Econ Bot 49, 406–422.
Hansen, G., Heitmann, A., Witt, T., Li, H., Jiang, H., Shen, X., Heussler, V. T, Rennenberg, A., and Hilgenfeld, R. (2011). Structural basis for the regulation of cysteine-protease activity by a new class of protease inhibitors in Plasmodium. Structure 19, 919–929.
Hernández-González, J.E., Salas-Sarduy, E., Hernández Ramírez, L.F., Pascual, M.J., Álvarez, D.E., Pabón, A., Leite, V.B.P, Pascutti, P.G., and Valiente, P.A. (2018). Identification of (4-(9H-fluoren-9-yl) piperazin-1-yl) methanone derivatives as falcipain 2 inhibitors active against Plasmodium falciparum cultures. Biochim Biophys Acta Gen Subj 1862, 2911–2923.
Hogg, T., Nagarajan, K, Herzberg, S., Chen, L., Shen, X., Jiang, H., Wecke, M., Blohmke, C., Hilgenfeld, R., and Schmidt, C.L. (2006). Structural and functional characterization of falcipain-2, a hemoglobinase from the malarial parasite Plasmodium falciparum. J Biol Chem 281, 25425–25437.
Jain, M., Khan, S.I., Tekwani, B.L., Jacob, M.R., Singh, S., Singh, P.P., and Jain, R. (2005). Synthesis, antimalarial, antileishmanial, and antimicrobial activities of some 8-quinolinamine analogues. Bioorg Med Chem 13, 4458–4466.
Kerr, I.D., Lee, J.H., Pandey, K.C, Harrison, A., Sajid, M., Rosenthal, P.J., and Brinen, L.S. (2009). Structures of falcipain-2 and falcipain-3 bound to small molecule inhibitors: implications for substrate specificity. J Med Chem 52, 852–857.
Koehn, F.E., and Carter, G.T. (2005). The evolving role of natural products in drug discovery. Nat Rev Drug Discov 4, 206–220.
Landier, J., Parker, D.M., Thu, A.M., Carrara, V.I., Lwin, K.M., Bonnington, CA., Pukrittayakamee, S., Delmas, G., and Nosten, F.H. (2016). The role of early detection and treatment in malaria elimination. Malar J 15, 363.
Lew, V.L., Tiffert, T., and Ginsburg, H. (2003). Excess hemoglobin digestion and the osmotic stability of Plasmodium falciparum-infected red blood cells. Blood 101, 4189–4194.
Li, H., Huang, J., Chen, L., Liu, X., Chen, T, Zhu, J., Lu, W., Shen, X., Li, J., Hilgenfeld, R., et al. (2009). Identification of novel falcipain-2 inhibitors as potential antimalarial agents through structure-based virtual screening. J Med Chem 52, 4936–4940.
Liu, W., Li, Y., Learn, G.H., Rudicell, R.S., Robertson, J.D., Keele, B.F., Ndjango, J.B.N., Sanz, C.M., Morgan, D.B., Locatelli, S., et al. (2010). Origin of the human malaria parasite Plasmodium falciparum in gorillas. Nature 467, 420–425.
Hinrichs, D.J., and Makler, M.T. (1993). Measurement of the lactate dehydrogenase activity of Plasmodium falciparum as an assessment of parasitemia. Am J Tropical Med Hygiene 48, 205–210.
Marco, M., and Miguel Coteron, J. (2012). Falcipain inhibition as a promising antimalarial target. Curr Top Med Chem 12, 408–444.
Mojab, F. (2012). Antimalarial natural products: a review. Avicenna J Phytomed 2, 52–62.
Muganga, R., Angenot, L., Tits, M., and Frederich, M. (2010). Anti-plasmodial and cytotoxic activities of Rwandan medicinal plants used in the treatment of malaria. J Ethnopharmacol 128, 52–57.
Mustafa, J., Khan, S.I., Ma, G., Walker, L.A., and Khan, I.A. (2004). Synthesis and anticancer activities of fatty acid analogs of podophyllotoxin. Lipids 39, 167–172.
Mustofa, Sholikhah, E.N., and Wahyuono, S. (2007). In vitro and in vivo antiplasmodial activity and cytotoxicity of extracts of Phyllanthus nir-uri L. herbs traditionally used to treat malaria in Indonesia. Southeast Asian J Trop Med Public Health 38, 609–615.
Musyoka, T.M., Kanzi, A.M., Lobb, K.A., and Tastan Bishop, Ö. (2016a). Analysis of non-peptidic compounds as potential malarial inhibitors against Plasmodial cysteine proteases via integrated virtual screening workflow. J Biomol Struct Dyn 34, 2084–2101.
Musyoka, T.M., Kanzi, A.M., Lobb, K.A., and Tastan Bishop, Ö. (2016b). Structure based docking and molecular dynamic studies of plasmodial cysteine proteases against a South African natural compound and its analogs. Sci Rep 6, 23690.
Pandey, K.C., Barkan, D.T., Sali, A., and Rosenthal, P.J. (2009). Regulatory elements within the prodomain of falcipain-2, a cysteine protease of the malaria parasite Plasmodium falciparum. PLoS ONE 4, e5694.
Pandey, K.C., Wang, S.X., Sijwali, P.S., Lau, A.L., McKerrow, J.H., and Rosenthal, P.J. (2005). The Plasmodium falciparum cysteine protease falcipain-2 captures its substrate, hemoglobin, via a unique motif. Proc Natl Acad Sci USA 102, 9138–9143.
Repetto, G., del Peso, A., and Zurita, J.L. (2008). Neutral red uptake assay for the estimation of cell viability/cytotoxicity Nat Protoc 3, 1125–1131.
Schulz, F., Gelhaus, C., Degel, B., Vicik, R., Heppner, S., Breuning, A., Leippe, M., Gut, J., Rosenthal, P.J., and Schirmeister, T. (2007). Screening of protease inhibitors as antiplasmodial agents. Part I: aziridines and epoxides. ChemMedChem 2, 1214–1224.
Shenai, B.R., Sijwali, P.S., Singh, A., and Rosenthal, P.J. (2000). Characterization of native and recombinant falcipain-2, a principal trophozoite cysteine protease and essential hemoglobinase of Plasmodium falciparum. J Biol Chem 275, 29000–29010.
Stoye, A., Juillard, A., Tang, A.H., Legac, J., Gut, J., White, K.L., Charman, S.A., Rosenthal, P.J., Grau, G.E.R., Hunt, N.H., et al. (2019). Falcipain inhibitors based on the natural product gallinamide A are potent in vitro and in vivo antimalarials. J Med Chem 62, 5562–5578.
Thu, A.M., Phyo, A.P, Landier, J., Parker, D.M., and Nosten, F.H. (2017). Combating multidrug-resistant Plasmodium falciparum malaria. FEBS J 284, 2569–2578.
Trager, W., and Jensen, J.B. (2005). Human malaria parasites in continuous culture. J Parasitai 91, 484–486.
Tu, Y. (2011). The discovery of artemisinin (qinghaosu) and gifts from Chinese medicine. Nat Med 17, 1217–1220.
Wang, L., Zhang, S., Zhu, J., Zhu, L., Liu, X., Shan, L., Huang, J., Zhang, W., and Li, H. (2014). Identification of diverse natural products as falcipain-2 inhibitors through structure-based virtual screening. Bioorg Med Chem Lett 24, 1261–1264.
WHO. (2018). World Malaria Report 2018 (Switzerland: World Health Organization).
Zhu, J., Chen, T, Chen, L., Lu, W., Che, P., Huang, J., Li, H., Li, J., and Jiang, H. (2009). 2-Amido-3-(lH-indol-3-yl)-N-substitued-propanamides as a new class of falcipain-2 inhibitors. 1. design, synthesis, biological evaluation and binding model studies. Molecules 14, 494–508.
Acknowledgements
We thank Prof. Hua **e (Shanghai Insitute of Materia Medica, Chinese Academy of Sciences) for expert technical assistance with the target location experiments using confocal microscopy; Prof. ** Huang (East China University of Science and Technology) for the enzyme inhibition assays and the effect evaluation of NP1024 on degradation of hemoglobin by FP-2; Profs. Huaimin Zhu, Weiqing Pan, Heng Peng, and Weibin Guan (The Secondary Military Medical University) for sharing Plasmodium parasite strains and providing technical assistance with the in vitro culture of Plasmodium parasites; **anwen Yang (Third Institute of Oceanography, State Oceanic Administration) for providing natural compounds used in this study; Drs. Troy J. Smillie and Ikhlas A. khan (National Center for Natural Products Research, the University of Mississippi) for in vitro anti-malarial assay; We also thankProf Hualiang Jiang and Eric Xu (Shanghai Insitute of Materia Medica) for their technological assistance and helpful suggestions. This work was supported by the National Key Research and Development Program (2016YFA0502304 to H.L), the National Natural Science Foundation of China (81825020), the National Science & Technology Major Project “Key New Drug Creation and Manufacturing Program”, China (20187X09711002), the Fundamental Research Funds for the Central Universities, Special Program for Applied Research on Super Computation of the NSFC-Guangdong Joint Fund (the second phase) (Ul 501501), Professor of Chang Jiang Scholars Program (to W.Z.), and the Natural Science Foundation of Zhejiang Province (LY15H190007). Honglin Li is also sponsored by the National Program for Special Supports of Eminent Professionals and National Program for Support of Top-notch Young Professionals.
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Zhu, L., Shan, L., Zhu, J. et al. Discovery of a natural fluorescent probe targeting the Plasmodium falciparum cysteine protease falcipain-2. Sci. China Life Sci. 63, 1016–1025 (2020). https://doi.org/10.1007/s11427-019-1581-2
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DOI: https://doi.org/10.1007/s11427-019-1581-2