High-throughput screening identifies new inhibitors of the multidrug transporter MRP4 — ASN Events
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  3. High-throughput screening identifies new inhibitors of the multidrug transporter MRP4

High-throughput screening identifies new inhibitors of the multidrug transporter MRP4 (#106)

Leanna Cheung 1 , Claudia L Flemming 1 , Fujiko Watt 1 , Denise Yu 1 , Zillan Neiron 1 , Tim W Failes 1 2 , Greg M Arndt 1 2 , Jamie I Fletcher 1 , Michelle Haber 1 , Murray D Norris 1
  1. Children's Cancer Institute Australia, Randwick, NSW, Australia
  2. ACRF Drug Discovery Centre for Childhood Cancer, Randwick, NSW, Australia

Neuroblastoma is a disease of the sympaticoadrenal lineage of the neural crest, which accounts for ~15% of all paediatric oncology deaths, and patients presenting with high risk disease face survival rates of only 50%. Improvements in survival rates are likely to require the development of new molecularly targeted therapies. The ABC transporter MRP4 (Multidrug Resistance Protein 4) is a potential therapeutic target in neuroblastoma. It is transcriptionally regulated by the MYCN oncogene, is highly expressed in poor-outcome neuroblastoma, and effluxes topotecan and irinotecan, two cytotoxic drugs increasingly used in neuroblastoma therapy. MRP4 also regulates the pharmacokinetics of its drug substrates and its inhibition can substantially increase their bioavailability. Moreover, MRP4 may have an important role in the growth and survival of neuroblastoma cells, independent of its role in chemoresistance.

Cell-based high-throughput screening was undertaken to identify novel inhibitors of MRP4 using two different approaches:

  1. A cell viability screen for compounds that sensitize MRP4-overexpressing cells to the MRP4 substrate 6-mercaptopurine, conducted using a 30,000 compound diversity library; and
  2. A bioluminescence screen identifying compounds inhibiting the efflux of the MRP4 substrate D-luciferin, conducted using a 3600 compound library containing FDA-approved drugs and known bioactive molecules.

MRP4 inhibitors were identified from both screens. Compounds were validated in MRP4-expressing cell lines using a range of diverse MRP4 substrates. From the diversity screen, two structurally unrelated compounds, Ceefourin 1 and Ceefourin 2, were shown to be highly selective for MRP4 over other ABC transporters, including P-glycoprotein, ABCG2 and MRP1, had low cellular toxicity, and displayed high microsomal and acid stability. We also identified the FDA-approved drug Dantrolene as an MRP4 inhibitor. These inhibitors may be useful as investigative tools, and as potential therapeutics that function by reversing MRP4-mediated chemoresistance, increasing the bioavailability of MRP4 substrate drugs, or targeting the cellular functions of MRP4.