Researchers may have discovered how to prevent the cellular-cleaning protein ABCG2 from removing chemotherapeutics from tumor cells, according to a recent study published by Gose et al in Nature Communications. These research findings may improve future cancer therapeutics.
Background
Most new cancer cases are treated with chemotherapy, which involves a course of one or several drugs that usually take the form of small molecules. However, the body naturally employs a family of proteins called adenosine triphosphate (ATP)-binding cassettes that function as cellular cleaners and indiscriminately collect and transport excess chemicals from the cells. This process may encourage the protein ABCG2 to remove chemotherapy drugs from the tumor cells before they have had any effect—which could reduce the effectiveness of treatment. Because ABCG2 excels at its job, the researchers stressed that despite the design of the chemotherapy, the protein may have the tools to remove it from the cells.
"ABCG2 plays a role in clearing toxins and protecting healthy stem cells from chemotherapy and toxic chemotherapeutic agents," explained senior study author John Schuetz, PhD, of the Department of Pharmacy and Pharmaceutical Sciences at the St. Jude Children’s Research Hospital. "We showed a few years ago that ABCG2 levels were quite high in certain subtypes of medulloblastoma, which impacted how much of the drug could get in to kill the cancer cells. In this study, we found why the protein is able to clear out such a broad spectrum of chemo[therapy] drugs, which can now inform efforts to improve anticancer strategies,” he emphasized.
Study Methods and Results
In this study, Dr. Schuetz and his colleagues took an interrogative approach to answer how ABCG2 removes chemotherapies from the cells by probing the substrate binding site. "We were completely puzzled by ABCG2's promiscuity until we did these studies," Dr. Schuetz stressed. "We found that unlike some of its genetic family members, which have a very hydrophobic binding pocket, the ABCG2 pocket is dotted with hydrophilic residues as well as hydrophobic ones,” he highlighted.
ATP-binding cassettes are usually responsible for removing hydrophobic molecules such as lipids, whereas other transporters "take care" of hydrophilic molecules. The researchers noted that ABCG2, however, is involved in both processes—making it capable of removing many types of anticancer drugs from the cells and limiting therapeutic efficacy.
The researchers found that two amino acids—a threonine and an asparagine—stood out within the substrate binding site of ABCG2. The distinguishing feature of these amino acids was that they were polar and therefore hydrophilic. To test whether these two residues were involved in ABCG2's promiscuity, the researchers turned each amino acid in the substrate binding site into a featureless alanine and observed how each mutation affected the transport of small molecules into cells.
The researchers discovered that although most of the mutations resulted in a moderately decreased ability of ABCG2 to bind substrates and transport them across the cell membrane, some capabilities were increased. For hydrophobic molecules, the ABCG2 with an alanine—instead of an asparagine—was twofold more effective at transporting these compounds. And, its ability to transport hydrophilic compounds was almost completely removed.
"The protein toggles like a one-way door to be open on one side of the membrane or the other to move things out of the cell. This movement excludes any water," Dr. Schuetz detailed. "With this asparagine to alanine mutation, it's allowing some water to move in. This moves the hydrophilic substrates and locks them in tight, so they stick around much longer and are not transported very well,” he emphasized.
Conclusions
The results of the study may indicate a way forward to combat chemotherapy resistance. ABCG2 inhibitors are often combined with chemotherapies, but preventing ABCG2 function can result in off-target detrimental effects. "The goal is to design ABCG2 inhibitors that have minimal effect on normal tissues but target the tumors," Dr. Schuetz underscored.
The researchers hope their new findings may lead to the development of more effective, less detrimental inhibitors designed to target the binding site threonine in ABCG2 when paired with hydrophilic molecules used in chemotherapy.
Disclosure: For full disclosures of the study authors, visit nature.com.
