title: Ischemia Induces Quiescence and Autophagy Dependence in Hepatocellular Carcinoma.
abstract: Purpose To characterize hepatocellular carcinoma (HCC) cells surviving ischemia with respect to cell cycle kinetics, chemosensitivity, and molecular dependencies that may be exploited to potentiate treatment with transarterial embolization (TAE). Materials and Methods Animal studies were performed according to institutionally approved protocols. The growth kinetics of HCC cells were studied in standard and ischemic conditions. Viability and cell cycle kinetics were measured by using flow cytometry. Cytotoxicity profiling was performed by using a colorimetric cell proliferation assay. Analyses of the Cancer Genome Atlas HCC RNA-sequencing data were performed by using Ingenuity Pathway Analysis software. Activation of molecular mediators of autophagy was measured with Western blot analysis and fluorescence microscopy. In vivo TAE was performed in a rat model of HCC with (n = 5) and without (n = 5) the autophagy inhibitor Lys05. Statistical analyses were performed by using GraphPad software. Results HCC cells survived ischemia with an up to 43% increase in the fraction of quiescent cells as compared with cells grown in standard conditions (P < .004). Neither doxorubicin nor mitomycin C potentiated the cytotoxic effects of ischemia. Gene-set analysis revealed an increase in mRNA expression of the mediators of autophagy (eg, CDKN2A, PPP2R2C, and TRAF2) in HCC as compared with normal liver. Cells surviving ischemia were autophagy dependent. Combination therapy coupling autophagy inhibition and TAE in a rat model of HCC resulted in a 21% increase in tumor necrosis compared with TAE alone (P = .044). Conclusion Ischemia induces quiescence in surviving HCC cells, resulting in a dependence on autophagy, providing a potential therapeutic target for combination therapy with TAE.©RSNA, 2017 Online supplemental material is available for this article.
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Terence P F
authors.initials: TPF
authors.lastname: Gade
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Tucker
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Nakazawa
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Hunt
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Waihay
authors.initials: W
authors.lastname: Wong
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Bryan
authors.initials: B
authors.lastname: Krock
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Charles N
authors.identifier: 0000-0003-3792-2589
authors.initials: CN
authors.lastname: Weber
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Nadolski
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Timothy W I
authors.initials: TWI
authors.lastname: Clark
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Michael C
authors.initials: MC
authors.lastname: Soulen
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Emma E
authors.initials: EE
authors.lastname: Furth
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Winkler
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: Ravi K
authors.initials: RK
authors.lastname: Amaravadi
authors.affiliation: From the Penn Image-Guided Interventions Laboratory (T.P.F.G., S.J.H., C.N.W., G.J.N.), Department of Radiology (T.P.F.G., S.J.H., C.N.W., G.J.N., T.W.I.C., M.C. Soulen), and Department of Pathology (E.E.F.), Hospital of the University of Pennsylvania, Philadelphia, Pa; Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, 421 Curie Blvd, 456 BRB II/III, Philadelphia, PA 19104 (E.T., M.S.N., W.W., B.K., M.C. Simon); Abramson Family Cancer Center (B.K., R.K.A.) and Department of Chemistry (J.D.W.), University of Pennsylvania, Philadelphia, Pa.
authors.forename: M Celeste
authors.initials: MC
authors.lastname: Simon
mesh_terms: D000818:Animals; D001343:Autophagy; D006528:Carcinoma, Hepatocellular / Q000098:blood supply* / Q000473:pathology*; D059447:Cell Cycle Checkpoints; D045744:Cell Line, Tumor; D002470:Cell Survival; D004621:Embolization, Therapeutic; D008114:Liver Neoplasms, Experimental / Q000098:blood supply* / Q000473:pathology*; D051381:Rats; D017208:Rats, Wistar