More than 50,000 people die per year in the United States from pain relief medication. The misuse of opioids, which are very addictive, costs Americans many billions of dollars in health-care costs, combating crime and loss of working income. Researchers have now developed a method to test the side-effects of new drugs directly in our brain. The method can also be used to study sleep disturbances and metabolic diseases.
In 2017, the United States President announced a major crisis initiative in the health sector in relation to pain relief medicine. In Europe, 3 in 4 deaths caused by overdose among 15–39 year olds result from opioid misuse. This opioid epidemic stems from the development of medicinal drugs. How these drugs affect the brain is often tested in artificial conditions in the laboratory. Researchers can now take live snapshots of the effect on the mouse brain using a brand new technology.
“These drugs affect the G protein–coupled receptors in the brain. The problem is that, apart from relieving pain, they can have an unintended addictive effect. By measuring how a drug affects the proteins in the brain, we can now obtain a much more precise picture of both the positive and the negative effects. We can thereby discover the negative effects at an early stage in developing new candidate drugs,” explains Matthias Mann, Professor and Research Director, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen.
Measuring the effects of medication
The researchers have called this new method in vivo brain phosphoproteomics. It combines mass spectrometry, which measures the mass and structure of molecules, with advanced computer calculations. This method enables researchers to measure precisely which brain proteins are modified chemically at a given time on more than 50,000 sites.
The phosphorylation of cellular proteins can switch them on or off and hence influence cellular activity. The phosphorylation patterns therefore directly measure the cellular reaction to a specific drug. We can take the brain cells from a mouse and measure how the drug works at a given time. This can give us a much more precise picture of how a drug works directly in the brain.
To test the method, the researchers investigated the effect of a drug, U50,488H, which is often used in laboratory experiments because it is known to stimulate the kappa opioid receptors. These receptors react to drugs such as morphine, codeine and methadone. U50,488H can therefore give researchers an idea about whether the new method could be used to measure the effects of other opioid drugs.
“Soon after we had injected the drug into the brain, we measured a change in the phosphorylation level in 5% of all positions – and only in the regions of the brain in which we knew opioid receptors existed. We also detected that the phosphorylation level gradually declined. The method appears to be both precise and sensitive enough to measure how medication affects specific regions of the brain.”
Revealing the mysteries of sleep
This new method should therefore make testing how future drugs affect brain easier, cheaper and safer. In the long term, it may also strongly contribute to addressing the opioid epidemic, especially in the United States.
“The method should be able to reveal both whether the desired effect has been achieved and whether unwanted side-effects such as addiction and suicidal tendencies are avoided, which have only been addressed indirectly in pharmacological testing of existing drugs. The goal now is to try to help the pharmaceutical industry to use this new technically more difficult but safer method.”
However, phosphoproteomics has tremendous additional potential. One major goal of Matthias Mann and his colleagues is to discover new protein biomarkers in blood that can be used to diagnose and possibly prevent type 2 diabetes, obesity and other metabolic diseases. Ultimately, the researchers also see major opportunities for investigating one of the human body’s great mysteries: sleep.
Drugs affect the phosphorylation of proteins in our brain and so does lack of sleep. We will therefore use the method to investigate the human circadian rhythm and how this affects learning and health if we do not sleep enough. In the long term, this may help us to understand, diagnose and treat sleep disorders much more effectively.
”In vivo brain GPCR signaling elucidated by phosphoproteomics” has been published in Science. Matthias Mann is a Research Director, Novo Nordisk Foundation Center for Protein Research, Faculty of Health Sciences, University of Copenhagen and Director, Department of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, Munich, Germany.