Illuminating Receptor Behaviors in Acidic Environments

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Groundbreaking approach could have profound impacts on research and drug discovery

Researchers who are members of Sylvester Comprehensive Cancer Center and colleagues in the Isom lab in the Department of Molecular and Cellular Pharmacology at the University of Miami Miller School of Medicine have used a novel yeast platform to reveal how a large set of G protein-coupled receptors (GPCRs) behave in acidic environments. These findings could have a major impact on drug discovery for cancer, inflammatory diseases, pain and other conditions. The study was published in PNAS.

From left, Corin O’Shea, Geoff Taghon, Daniel Isom, Ph.D., and Nick Kapolka.

“We discovered that yeast is the perfect system for studying how the pH changes regulate human GPCR signaling because it tolerates acidity much better than mammalian cells,” said Daniel Isom, Ph.D., assistant professor of molecular and cellular pharmacology and senior author on the study. “Using this approach to profile many GPCRs, we found that most are active at higher (less acidic) pH values and act like binary switches that turn off abruptly when pH drops.”

GPCRs, and other receptor types, act like cellular USB ports that bring in news from the outside world. When molecules (called ligands) plug into GPCRs, the receptors send instructions into the cell, helping it respond to environmental changes. Ligands could be proteins, peptides (pieces of a protein) or small molecule drugs designed to illicit a specific cellular response.

The study found GPCRs are often less likely to allow ligand binding when the environmental pH becomes more acidic, which could have an enormous impact on cancer studies and drug development, as tumor microenvironments are generally acidic.

Highly selective drugs

“We’re trying to develop pH-intelligent therapeutics, looking for drugs that bind these receptors only at low pH or only at high pH,” Dr. Isom said. “Drugs could be designed to be highly selective, only targeting receptors in acidified microenvironments but avoiding them in normal tissue.”

These findings are built on a sophisticated yeast model, developed by Dr. Isom’s lab, which can rapidly identify molecules when they bind to GPCRs.

Saccharomyces cerevisiae (baker’s yeast) is an ideal model because it has only one GPCR pathway. Using CRISPR gene editing, the researchers introduce human GPCR genes into the yeast, along with a fluorescent reporter gene that glows when the receptor is activated. This model, called dynamic cyan induction by functional integrated receptors (DCyFIR), has great potential to rapidly identify therapeutic molecules.

 Changing the pH

“When we change pH outside a mammalian cell, the pH also changes inside the cell, but that doesn’t happen in yeast,” Dr. Isom said. “That allows us to interrogate the isolated ligand-GPCR interaction as a function of pH without affecting anything inside the cell.”

Front to back, Nicholas Kapolka and Geoffrey Taghon screen GPCR ligand interactions.

This breakthrough finding is a game-changer, because until now almost all drug discovery and testing in cells has been done at the body’s normal pH, around 7.4 (7 is neutral). By combining the acid tolerance of yeast with DCyFIR, the Isom lab is establishing a new era of drug discovery in which large libraries of lead compounds and approved drugs can be screened against receptors over a wide range of physiologically relevant pH values. The potential ramifications abound.

There are more than 800 GPCRs, making them the most common cellular receptors, as well as the most therapeutically targeted family of receptors. While this work has obvious applications in oncology, Dr. Isom notes that pH studies in yeast could also help optimize therapies for inflammatory environments, which are also acidic, and even lead to more targeted pain killers.

“If we could develop opioids that work better at low pH and really don’t work at normal pH, we could affect the opioid receptors in the peripheral versus central nervous system,” Dr. Isom said. “If we can do that, we may be able to avoid addiction and other unwanted side effects.”

Miller School co-authors were Nick Kapolka, Jacob Rowe, Geoffrey Taghon, William Morgan, and Corin O’Shea.