By Olivia Trani

A research team led by Virginia Commonwealth University has gained new insights into the molecular mechanisms that cause cocaine use disorder, identifying a potential therapeutic strategy to inhibit the drug’s addictive effect.

By tweaking one specific component that helps control the brain’s dopamine levels, the researchers were able to block the biological process that reinforces cocaine-seeking behaviors. Their preclinical findings, published in the Journal of Neuroscience, could open new avenues for developing a medication to help people overcome cocaine abuse.

Stimulant drugs, such as cocaine and methamphetamine, are involved in roughly half of all overdose deaths in the United States, either taken on their own or in combination with opioids. While a number of therapeutics exist for other forms of substance use disorders, such as opioid and alcohol abuse, there are currently no medications approved by the Food and Drug Administration to treat stimulant drug abuse.

“We still have gaps in our understanding the neurobiology of cocaine use disorder, and there is nothing in the pipeline in terms of having a therapeutic for this disorder,” said Lankupalle Jayanthi, Ph.D., an associate professor in the VCU School of Medicine’s Department of Pharmacology and Toxicology. She led the new study with Sammanda Ramamoorthy, Ph.D., a professor in the Department of Pharmacology and Toxicology.

While this work is still in its early stages, researchers are encouraged by the advancements their team has made in understanding the molecular mechanisms that drive cocaine misuse.

“It’s more important than ever that we keep researching why people become addicted to cocaine and how we may be able to help them through medication,” Ramamoorthy said.

Tackling dopamine disruption 

Jayanthi and Ramamoorthy have been studying cocaine’s effects on the brain for over 25 years in hopes of identifying pharmaceutical strategies to help people overcome substance use. They specifically analyze how cocaine impacts the brain’s dopamine levels. 

Dopamine is a neurotransmitter that plays a critical role in the brain’s reward system, the neural network responsible for pleasure and reinforcing behaviors. Whenever a person engages in a satisfying activity, dopamine is released from nerve cells. Subsequently, these neurotransmitters trigger a cascade of molecular reactions in the brain that improve mood and motivation. 

Stimulants like cocaine hijack the brain’s reward system. At first, the drug prompts a spike in dopamine levels and causes short-term feelings of euphoria and excitement. However, chronic use depletes extracellular dopamine levels over time, leading to dissatisfaction, unease and lack of motivation. Consequently, users often seek out more cocaine or other drugs of abuse to make up this dopamine deficiency, which is why this drug is highly addictive.

Cocaine drains dopamine levels by targeting two proteins that work together. Kappa opioid receptors regulate how much dopamine is released by the brain’s nerve cells. If the amount is high enough, the receptors activate dopamine transporters, which act as molecular vacuums and suck dopamine back into the nerve cells, priming the system for the next release.

Cocaine disrupts this balance. Cocaine use increases levels of a peptide called dynorphin, which binds to kappa opioid receptors and makes them more sensitive to dopamine levels. This process sets off a chain reaction that causes dopamine transporters to work in overdrive. Because dopamine is being sucked up at a higher rate, the kinds of everyday activities that typically trigger a dopamine boost no longer have the same effect. Instead, they promote negative feelings, including aversive and dysphoric experiences, and can lead to increased drug intake and relapse.

Researchers have attempted to develop medications that prevent cocaine use from interfering with this system, but there has yet to be success in getting drug candidates past clinical trials due to adverse side effects.

“The challenge is we need to develop a therapeutic that inhibits the downstream effects of cocaine use without interfering with the proteins’ normal functions,” Ramamoorthy said.

A new path for breaking the cycle  

By gaining molecular insights into how kappa opioid receptors and dopamine transporters interact, Jayanthi and Ramamoorthy’s team have discovered a possible therapeutic strategy to reduce cocaine’s impact on dopamine levels, which could minimize the stimulant’s addictive effects. 

Their earlier studies made two key discoveries. One was that kappa opioid receptors activate dopamine transporters – and trigger their vacuum function – through a process called phosphorylation, in which a phosphate group is added to the transporter’s molecular structure. The second was the specific spot where the dopamine transporters become phosphorylated, which revealed that the phosphate group attaches to an amino acid called threonine-53. 

Jayanthi and Ramamoorthy’s team hypothesized that if they could control the level of threonine-53 phosphorylation occurring to the dopamine transporters, they may be able to prevent the proteins from working in overdrive, and consequently avoid some of the adverse and addictive effects of cocaine. 

To test this theory, the research team developed a mouse model with slightly modified dopamine transporters. The key alteration was swapping out threonine-53 with another amino acid called alanine, which cannot bind to phosphate groups.

“By making this small change, the modified dopamine transporter cannot be phosphorylated at threonine-53,” Jayanthi said.

The researchers then ran various tests to assess whether blocking threonine-53 phosphorylation in dopamine transporters would ultimately disrupt the molecular mechanisms that lead to the adverse effects of cocaine use. 

When injected with a drug that activates kappa opioid receptors, mice with normal dopamine transporters showed increased phosphorylation at the threonine-53 site and increased transporter activity. The mice also showed increased aversiveness and reward-seeking behavior, which are typical side effects from using drugs that activate kappa opioid receptors. On the other hand, mice with the modified dopamine transporters did not show enhanced transporter activity and experienced no behavioral side effects. The researchers believe that therapeutics targeting this site-specific phosphorylation could reduce cocaine’s addictive effects. 

Developing a drug

Jayanthi, Ramamoorthy and their colleagues are now developing an mRNA-based drug, also known as a minigene, that produces peptides whose structure matches the threonine-53 phosphorylation site on dopamine transporters. Since the peptides have the same phosphate-bonding site, kappa opioid receptors can target these peptides and relieve some pressure off the dopamine transporters. This in turn would prevent dopamine transporters from working in overdrive and depleting extracellular dopamine levels.

“If you truly want to understand something, you need to know its root cause,” Jayanthi said. “Binding cocaine to dopamine transporters is the root cause of cocaine’s behavioral effects.”

Ramamoorthy added, “we hope that our progress in understanding the root causes of cocaine use disorder ultimately brings us closer to making therapeutics a reality for those in recovery.”

He and his colleagues are applying their molecular insights into other dopamine-related conditions, like ADHD, schizophrenia, bipolar disorder, and autism spectrum disorder. In another preclinical study, published in Molecular Psychiatry, the VCU research team and collaborators from Florida Atlantic University explored how targeting kappa opioid receptors and dopamine transporters could potentially correct dopamine levels for those with these disorders. 

The work by Jayanthi and Ramamoorthy’s team in both of these projects was supported by the National Institutes of Health, the Brain & Behavior Research Foundation, and the Max Kade Fellowship of the Austrian Academy of Sciences.