PARP inhibitors and radiation potentiate liver cell death in vitro. Do hepatocellular carcinomas have an achilles’ heel? - 10/11/20

Doi : 10.1016/j.clinre.2020.09.014

Laetitia Gerossier ^a,^{^{1
These authors contributed equally to this work.}}, Anaëlle Dubois ^a,^{^{1
These authors contributed equally to this work.}}, Alexia Paturel ^a, Nadim Fares ^a,^{^{2
Present address: Department of Digestive Oncology Rangueil Universitary Hospital, Hopitaux de Toulouse, Toulouse France.}}, Damien Cohen ^a, Phillippe Merle ^a,^b, Joel Lachuer ^a,^c, Anne Wierinckx ^a,^c, Pierre Saintigny ^a,^d, Brigitte Bancel ^e, Janick Selves ^f, Anne Schnitzler ^g, Bérengère Ouine ^h, Aurélie Cartier ^h, Leanne de Koning ^h, Vincent Puard ^h, Ivan Bieche ^g, Hector Hernandez-Vargas ^a, Janet Hall ^a, Isabelle Chemin ^a,⁎
^a Univ Lyon, Université Claude Bernard Lyon 1, INSERM, CNRS, Centre Léon Bérard, Centre De Recherche En Cancérologie De Lyon, Lyon, 69008, France
^b Department of Hepatology, Groupement Hospitalier Nord, Hospices Civils De Lyon, Lyon, 69000 France
^c ProfileXpert, SFR-Est, CNRS UMR-S3453, INSERM US7, Lyon Cedex 08, F-69373, France
^d Department of Medical Oncology, Centre Léon Bérard, Lyon, France
^e Service d’Anatomopathologie, Groupement Hospitalier Est, Hospices Civils De Lyon, Lyon, 69000, France
^f Anatomie Et Cytologie Pathologiques Pôle IUC Oncopole CHU Institut Universitaire Du Cancer De Toulouse - Oncopole, Toulouse, F- 31059, France
^g Department of Genetics, Institut Curie, PSL Research University, Paris, F-75005, France
^h Department of Translational Research, Institut Curie, PSL Research University, Paris, F-75005, France

^⁎Corresponding author.

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Highlights

•	PARPi cytotoxicity is significantly higher when combined with X-rays in liver cells
•	HBx expression significantly lowered this survival, probably driven by SMC5/6 loss
•	HCC Tumor tissues had higher PARP mRNA levels than Peri-Tumor and control livers
•	Increased DNA damage levels are found during liver disease progression

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Abstract

Background

A promising avenue for cancer treatment is exacerbating the deregulation of the DNA repair machinery that would normally protect the genome. To address the applicability of poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) combined with radiotherapy for the treatment of hepatocellular carcinoma (HCC) two approaches were used: firstly, the in vitro sensitivity to the PARPi Veliparib and Talazoparib +/- radiation exposure was determined in liver cell lines and the impact of the HBV X protein (HBx) that deregulates cellular DNA damage repair via SMC5/6 degradation was investigated. Secondly, PARP expression profiles and DNA damage levels using the surrogate marker gammaH2AX were assessed in a panel of control liver vs HCC tissues.

Methods

Cell cytotoxicity was measured by clonogenic survival or relative cell growth and the DNA damage response using immunological-based techniques in Hep3B, PLC/PRF/5, HepG2- and HepaRG-derived models. Transcriptome changes due to HBx expression vs SMC6 loss were assessed by RNA sequencing in HepaRG-derived models. PARP and PARG transcripts (qPCR) and PARP1, H2AX and gammaH2AX protein levels (RPPA) were compared in control liver vs HBV-, HCV-, alcohol- and non-alcoholic steatohepatitis-associated HCC (tumor/peritumor) tissues.

Results

PARPi cytotoxicity was significantly enhanced when combined with X-rays (2Gy) with Talazoparib having a greater impact than Veliparib in most in vitro models. HBx expression significantly lowered survival, probably driven by SMC5/6 loss based on the transcriptome analysis and higher DNA damage levels. PARP1 and PARP2 transcript levels were significantly higher in tumor than peritumor and control tissues. The HBV/HCV/alcohol-associated tumor tissues studied had reduced H2AX but higher gammaH2AX protein levels compared to peritumor and control tissues providing evidence of increased DNA damage during liver disease progression.

Conclusions

These proof-of-concept experiments support PARPi alone or combined with radiotherapy for HCC treatment, particularly for HBV-associated tumors, that warrant further investigation.

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Abbreviations : HCC, RT, HBV, SBRT, PARP, PARPi, IR, SMC5/6, HR, DSBs, PARG, gammaH2AX, PT, T, HepaRG TR, HepaRG TRX, HepaRG TRX sh-scramble, HepaRG TRX sh-SMC6, HU, DAPI, AU, D₃₇, RPPA, FPKM, DDR