title: Hepatocellular carcinoma-targeted drug discovery through image-based phenotypic screening in co-cultures of HCC cells with hepatocytes.
abstract: Hepatocellular carcinoma (HCC) is one of the most common malignant cancers worldwide and is associated with substantial mortality. Because HCCs have strong resistance to conventional chemotherapeutic agents, novel therapeutic strategies are needed to improve survival in HCC patients.
  
  
  
  Here, we developed a fluorescence image-based phenotypic screening system in vitro to identify HCC-specific drugs in co-cultures of HCC cells with hepatocytes. To this end, we identified two distinctive markers of HCC, CHALV1 and AFP, which are highly expressed in HCC cell lines and liver cancer patient-derived materials. We applied these markers to an HCC-specific drug screening system.
  
  
  
  Through pilot screening, we identified three anti-folate compounds that had HCC-specific cytotoxicity. Among them, pyrimethamine exhibited the greatest HCC-specific cytotoxicity. Interestingly, pyrimethamine significantly increased the size and number of lysosomes and subsequently induced the release of cathepsin B from the lysosome to the cytosol, which triggered caspase-3-dependent apoptosis in Huh7 (HCC) but not Fa2N-4 cells (immortalized hepatocytes). Importantly, Fa2N-4 cells had strong resistance to pyrimethamine relative to Huh7 cells in 2D and 3D culture systems.
  
  
  
  These results demonstrate that this in vitro image-based phenotypic screening platform has the potential to be widely adopted in drug discovery research, since we promptly estimated anticancer activity and hepatotoxicity and elucidated functional roles of pyrimethamine during the apoptosis process in HCC.
authors.affiliation: Cancer Biology Research Laboratory, Institut Pasteur Korea, 16, Daewangpangyo-ro 712 beon-gil, Bundang-gu, Seongnam-si, Gyeonggi-do, 13488, Korea.|Laboratory of Biochemistry, Division of Life Sciences, Korea University, 145, Anam-ro, Seongbuk-gu, Seoul, 02841, Korea.
authors.forename: Jae-Woo
authors.initials: JW
authors.lastname: Jang
authors.affiliation: Cancer Biology Research Laboratory, Institut Pasteur Korea, 16, Daewangpangyo-ro 712 beon-gil, Bundang-gu, Seongnam-si, Gyeonggi-do, 13488, Korea.|Laboratory of Biochemistry, Division of Life Sciences, Korea University, 145, Anam-ro, Seongbuk-gu, Seoul, 02841, Korea.
authors.forename: Yeonhwa
authors.initials: Y
authors.lastname: Song
authors.affiliation: Division of Gastroenterology and Hepatology, ASAN Medical center, Olympic-ro 43-gil, Songpagu, Seoul, 05505, Korea.
authors.forename: Kang Mo
authors.initials: KM
authors.lastname: Kim
authors.affiliation: Division of Gastroenterology and Hepatology, ASAN Medical center, Olympic-ro 43-gil, Songpagu, Seoul, 05505, Korea.
authors.forename: Jin-Sun
authors.initials: JS
authors.lastname: Kim
authors.affiliation: Division of Radiation Oncology, ASAN Medical center, Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, Korea.
authors.forename: Eun Kyung
authors.initials: EK
authors.lastname: Choi
authors.affiliation: Laboratory of Biochemistry, Division of Life Sciences, Korea University, 145, Anam-ro, Seongbuk-gu, Seoul, 02841, Korea. joonkim@korea.ac.kr.
authors.forename: Joon
authors.initials: J
authors.lastname: Kim
authors.affiliation: Cancer Biology Research Laboratory, Institut Pasteur Korea, 16, Daewangpangyo-ro 712 beon-gil, Bundang-gu, Seongnam-si, Gyeonggi-do, 13488, Korea. shr1261@ip-korea.org.
authors.forename: Haengran
authors.initials: H
authors.lastname: Seo
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use*; D014408:Biomarkers, Tumor / Q000378:metabolism; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000473:pathology; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D055808:Drug Discovery / Q000379:methods*; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D005493:Folic Acid Antagonists / Q000494:pharmacology; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000473:pathology; D018613:Microscopy, Confocal; D058990:Molecular Targeted Therapy / Q000379:methods*; D011739:Pyrimethamine / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D000970:Antineoplastic Agents; D014408:Biomarkers, Tumor; D005493:Folic Acid Antagonists; D011739:Pyrimethamine
keywords: 3D culture systems; Co-cultures; Hepatocellular carcinoma; Phenotypic screening; Pyrimethamine