title: Antiproliferative activity and phenotypic modification induced by selected Peruvian medicinal plants on human hepatocellular carcinoma Hep3B cells.
abstract: BACKGROUND
  The high incidence of human hepatocellular carcinoma (HCC) in Peru and the wide use of medicinal plants in this country led us to study the activity against HCC cells in vitro of somes species used locally against liver and digestive disorders.
  
  
  METHODS
  Ethnopharmacological survey: Medicinal plant species with a strong convergence of use for liver and digestive diseases were collected fresh in the wild or on markets, in two places of Peru: Chiclayo (Lambayeque department, Chiclayo province) and Huaraz (Ancash department, Huaraz province). Altogether 51 species were collected and 61 ethanol extracts were prepared to be tested. Biological assessment: All extracts were first assessed against the HCC cell line Hep3B according a 3-step multi-parametric phenotypic assay. It included 1) the evaluation of phenotypic changes on cells by light microscopy, 2) the measurement of the antiproliferative activity and 3) the analysis of the cytoskeleton and mitosis by immunofluorescence. Best extracts were further assessed against other HCC cell lines HepG2, PLC/PRF/5 and SNU-182 and their toxicity measured in vitro on primary human hepatocytes.
  
  
  RESULTS
  Ethnopharmacological survey: Some of the species collected had a high reputation spreading over the surveyed locations for treating liver problems, i.e. Baccharis genistelloides, Bejaria aestuans, Centaurium pulchellum, Desmodium molliculum, Dipsacus fullonum, Equisetum bogotense, Gentianella spp., Krameria lapacea, Otholobium spp., Schkuhria pinnata, Taraxacum officinale. Hep3B evaluation: Fourteen extracts from 13 species (Achyrocline alata, Ambrosia arborescens, Baccharis latifolia, Hypericum laricifolium, Krameria lappacea, Niphidium crassifolium, Ophryosporus chilca, Orthrosanthus chimboracensis, Otholobium pubescens, Passiflora ligularis, Perezia coerulescens, Perezia multiflora and Schkuhria pinnata) showed a significant antiproliferative activity against Hep3B cells (IC50≤ 50µg/mL). This was associated with a lack of toxicity on primary human hepatocytes in vitro. Immunofluorescence experiments on Hep3B cells showed that crude extracts of Schkuhria pinnata and Orthrosanthus chimboracensis could block Hep3B cells in mitosis with an original phenotype. Crude extracts of Perezia coerulescens, Perezia multiflora, Achyrocline alata, Ophryosporus chilca, Otholobium pubescens and Hypericum laricifolium could modify the overall microtubule cytoskeletal dynamics of Hep3B cells in interphase by an original mechanism.
  
  
  CONCLUSIONS
  Our method allowed us to select 9 extracts which displayed antiproliferative activities associated with original cellular phenotypes on Hep3B cells, regarding known microtubule-targeting drugs. Both chemical and cellular studies are ongoing in order to elucidate natural compounds and cellular mechanisms responsible of the activities described.
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France; Institut de Recherche pour le Développement; IRD; UMR 152 Pharma-DEV, F-31062 Toulouse cedex 9, France. Electronic address: maelle.carraz@ird.fr.
authors.forename: Maëlle
authors.initials: M
authors.lastname: Carraz
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France.
authors.forename: Cédric
authors.initials: C
authors.lastname: Lavergne
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France; Institut de Recherche pour le Développement; IRD; UMR 152 Pharma-DEV, Mission IRD, Casilla 18-1209 Lima, Peru.
authors.forename: Valérie
authors.initials: V
authors.lastname: Jullian
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France; Centre Nationalde la Recherche Scientifique; CNRS; UMR 152 Pharma-DEV; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France.
authors.forename: Michel
authors.initials: M
authors.lastname: Wright
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France.
authors.forename: Jean Edouard
authors.initials: JE
authors.lastname: Gairin
authors.affiliation: Universidad Ricardo Palma, Av. Benavides 5440 - Santiago de Surco, Lima 33, Peru.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Gonzales de la Cruz
authors.affiliation: Université de Toulouse; UPS; UMR 152 Pharma-DEV; Université Toulouse 3; Faculté, des Sciences Pharmaceutiques; 35 Chemin des Maraîchers, F-31062 Toulouse cedex 9, France; Institut de Recherche pour le Développement; IRD; UMR 152 Pharma-DEV, F-31062 Toulouse cedex 9, France.
authors.forename: Geneviève
authors.initials: G
authors.lastname: Bourdy
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D020834:Ethnopharmacology / Q000379:methods; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy*; D010568:Peru; D010936:Plant Extracts / Q000494:pharmacology*; D010946:Plants, Medicinal / Q000737:chemistry*
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D010936:Plant Extracts
keywords: Assay; Hep 3B cell line; Hepatocellular carcinoma; Liver; Medicinal plants; Peru; Phenotypic cell based