title: Hepatic stellate cells induce hepatocellular carcinoma cell resistance to sorafenib through the laminin-332/α3 integrin axis recovery of focal adhesion kinase ubiquitination.
abstract: In patients with hepatocellular carcinoma (HCC) receiving sorafenib, drug resistance is common. HCC develops in a microenvironment enriched with extracellular matrix proteins including laminin (Ln)-332, produced by hepatic stellate cells (HSCs). Ln-332 is the ligand of α3β1 and α6β4 integrins, differently expressed on the HCC cell surface, that deliver intracellular pathways. The aim of this study was to investigate the effect of Ln-332 on sorafenib's effectiveness. HCC cells were challenged with sorafenib in the presence of Ln-332 and of HSC conditioned medium (CM). Sorafenib impaired HCC cell proliferation and induced apoptosis. HSC-CM or Ln-332 inhibited sorafenib's effectiveness in HCC cells expressing both α3β1 and α6β4. Inhibiting α3 but not α6 integrin subunit using blocking antibodies or small interfering RNA abrogated the protection induced by Ln-332 and HSC-CM. Hep3B cells expressing α6β4 but lacking the α3 integrin were insensitive to Ln-332 and HSC-CM protective effects. Hep3B α3-positive, but not wild-type and scramble transfected, cells acquired protection by sorafenib when plated on Ln-332-CM or HSCs. Sorafenib dephosphorylated focal adhesion kinase (FAK) and extracellular signal-regulated kinases 1/2, whereas Ln-332 and HSC-CM partially restored the pathways. Silencing FAK, but not extracellular signal-regulated kinases 1/2, abrogated the protection induced by Ln-332 and HSC-CM, suggesting a specific role for FAK. Sorafenib down-regulated total FAK, inducing its proteasomal degradation, while Ln-332 and HSC-CM promoted the escape of FAK from ubiquitination, probably inducing a preferential membrane localization.
  
  
  
  This study unveils a novel mechanism of sorafenib resistance depending on the α3β1/Ln-332 axis and requiring FAK ubiquitination, providing new insights into personalizing therapy for patients with HCC. (Hepatology 2016;64:2103-2117).
authors.affiliation: National Cancer Institute, Istituto Tumori G. Paolo II, Bari, Italy.
authors.forename: Amalia
authors.initials: A
authors.lastname: Azzariti
authors.affiliation: National Institute of Gastroenterology, IRCCS "S. De Bellis", Castellana Grotte Bari, Italy.
authors.forename: Serena
authors.initials: S
authors.lastname: Mancarella
authors.affiliation: National Cancer Institute, Istituto Tumori G. Paolo II, Bari, Italy.
authors.forename: Letizia
authors.initials: L
authors.lastname: Porcelli
authors.affiliation: National Cancer Institute, Istituto Tumori G. Paolo II, Bari, Italy.
authors.forename: Anna Elisa
authors.initials: AE
authors.lastname: Quatrale
authors.affiliation: University of Florence Medical School, Florence, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Caligiuri
authors.affiliation: University of Bari Medical School, Bari, Italy.
authors.forename: Luigi
authors.initials: L
authors.lastname: Lupo
authors.affiliation: National Institute of Gastroenterology, IRCCS "S. De Bellis", Castellana Grotte Bari, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Dituri
authors.affiliation: National Institute of Gastroenterology, IRCCS "S. De Bellis", Castellana Grotte Bari, Italy.
authors.forename: Gianluigi
authors.initials: G
authors.lastname: Giannelli
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy*; D019008:Drug Resistance, Neoplasm; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000502:physiology*; D055166:Hepatic Stellate Cells / Q000502:physiology*; D006801:Humans; D039422:Integrin alpha3 / Q000502:physiology*; D007797:Laminin / Q000502:physiology*; D008113:Liver Neoplasms / Q000188:drug therapy*; D009536:Niacinamide / Q000031:analogs & derivatives* / Q000627:therapeutic use; D010671:Phenylurea Compounds / Q000627:therapeutic use*; D000077157:Sorafenib; D014407:Tumor Cells, Cultured; D054875:Ubiquitination
chemical_list: D000970:Antineoplastic Agents; D039422:Integrin alpha3; D007797:Laminin; D010671:Phenylurea Compounds; D009536:Niacinamide; D000077157:Sorafenib; D051416:Focal Adhesion Protein-Tyrosine Kinases