fibrolamellar carcinoma drug repurposing pediatric rare tumors patient derived xenografts drug screening fusion gene Abbreviations: ATR -Ataxia telangiectasia and Rad3-related protein AURKA -Aurora kinase A AURKB -Aurora kinase B Bcl-xL -B-cell lymphoma-extra large encoded by the BCL2-like 1 gene Bcl2 -B-cell lymphoma 2 ITS -Insulin, (human) Transferrin, Selenium BID -BH3 Interacting Domain Death Agonist BIM -Bcl-2-like protein 11 CA12 -Carbonic anhydrase 12 CDc7 -Cell division cycle 7-related protein kinase CDK -Cyclin depedent protein kinase Cyp19A1 -Also know as aromatase or estrogen synthase DAB -3,3'-Diaminobenzidine DMSO -Dimethyl sulfoxide DNA-PK -DNA protein kinase EGFR -Epidermal growth factor receptor eIF4F -Eukaryotic initiation factor 4F ErbB2 -erythroblastic oncogene B, also known as Her-2 protooncogene Neu and as epidermal growth factor receptor-2 FLC -fibrolamellar hepatocellular carcinoma H&E -hematoxylin and eosin HSA -Highest Single Agent HCC -hepatocellular carcinoma HDAC -Histone deacetylase HSP70 -Heat Shock Protein 70

Conflict of interest: GZ and DZ are inventors of patent no. 10807977 for use of BCL-XL PROTACs as antitumor agents. GZ and DZ are cofounders of and have equity in Dialectic Therapeutics, which develops BCL-XL/2 PROTACs to treat cancer.

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Fibrolamellar hepatocellular carcinoma (FLC) is a usually lethal primary liver cancer driven by a somatic dysregulation of protein kinase A. We show that the proteome of FLC tumors is distinct from that of adjacent nontransformed tissue. These changes can account for some of the cell biological and pathological alterations in FLC cells, including their drug sensitivity and glycolysis. Hyperammonemic encephalopathy is a recurrent problem in these patients, and established treatments based on the assumption of liver failure are unsuccessful. We show that many of the enzymes that produce ammonia are increased and those that consume ammonia are decreased. We also demonstrate that the metabolites of these enzymes change as expected. Thus, hyperammonemic encephalopathy in FLC may require alternative therapeutics.

<div>Abstract<p>To repurpose therapeutics for fibrolamellar carcinoma (FLC), we developed and validated patient-derived xenografts (PDX) from surgical resections. Most agents used clinically and inhibitors of oncogenes overexpressed in FLC showed little efficacy on PDX. A high-throughput functional drug screen found primary and metastatic FLC were vulnerable to clinically available inhibitors of TOPO1 and HDAC and to napabucasin. Napabucasin's efficacy was mediated through reactive oxygen species and inhibition of translation initiation, and specific inhibition of eIF4A was effective. The sensitivity of each PDX line inversely correlated with expression of the antiapoptotic protein Bcl-xL, and inhibition of Bcl-xL synergized with other drugs. Screening directly on cells dissociated from patient resections validated these results. This demonstrates that a direct functional screen on patient tumors provides therapeutically informative data within a clinically useful time frame. Identifying these novel therapeutic targets and combination therapies is an urgent need, as effective therapeutics for FLC are currently unavailable.</p>Significance:<p>Therapeutics informed by genomics have not yielded effective therapies for FLC. A functional screen identified TOPO1, HDAC inhibitors, and napabucasin as efficacious and synergistic with inhibition of Bcl-xL. Validation on cells dissociated directly from patient tumors demonstrates the ability for functional precision medicine in a solid tumor.</p><p><i>This article is highlighted in the In This Issue feature, p. 2355</i></p></div>