Bioengineers close to brewing opioids without using poppies

For centuries, poppy plants have been grown to provide opium, the compound from which morphine and other important painkillers are derived.

Now, bioengineers at Stanford have hacked the DNA of yeast and reprogrammed these simple cells to make opioid-based medicines via a sophisticated extension of the basic brewing process that makes beer.

Led by Christina Smolke, PhD, associate professor of bioengineering, the team has already spent a decade genetically engineering yeast cells to reproduce the biochemistry of poppies, with the ultimate goal of producing opium-based medicines, from start to finish, in fermentation vats.

“We are now very close to replicating the entire opioid production process in a way that eliminates the need to grow poppies, allowing us to reliably manufacture essential medicines while mitigating the potential for diversion to illegal use,” Smolke said.

The research is described in a paper that was published Aug. 24 in Nature Chemical Biology. Smolke is the senior author, and the lead author is Kate Thodey, PhD, a postdoctoral scholar in bioengineering. Stephanie Galanie, a doctoral student in chemistry, is the other co-author.

In the paper, the researchers detail how they added five genes from two different organisms to yeast cells. Three of these genes came from the poppy itself, and the others from a bacterium that lives on poppy stalks.

This gene mashup was required to turn yeast into cellular factories that replicate two separate processes: how nature produces opium in poppies, and how pharmacologists use chemical processes to further refine opium derivatives into modern opioid drugs, such as hydrocodone.

From plants to pills

Morphine is one of three principal painkillers derived from opium. As a class, they are called opiates. The other two important opiates are codeine, which has been used as a cough remedy, and thebaine, which is further refined by chemical processes to create higher-value therapeutics such as oxycodone and hydrocodone, often sold under the brand names OxyContin and Vicodin, respectively.

Today, legal poppy farming is restricted to a few countries — including Australia, France, Hungary, India, Spain and Turkey — supervised by the International Narcotics Control Board, which seeks to prevent opiates like morphine from being refined into heroin.

The biggest market for legal opiates, and their opioid derivatives, is the United States, where pharmaceutical factories use chemical processes to create the refined products that are used as pain-killing pills. However, poppies are not grown in significant quantities in the United States, creating various international dependencies and vulnerabilities in the supply of these medicines.

Turning yeast into a factory

The thrust of Smolke’s work for a decade has been to pack the entire production chain, from the fields of poppies through all the subsequent steps of chemical refining, into yeast cells, using the tools of bioengineering.

What Smolke’s team has now done is to carefully reprogram the yeast genome —the master instruction set that tells every organism how to live — to behave like a poppy when it comes to making opiates.

The process involves more than simply adding new genes into yeast. Opioid molecules are complex, three-dimensional objects. In nature, they are made in specific regions of the poppy. Since yeast cells do not have these complex structures and tissues, the Stanford team had to recreate the equivalent of poppy-like “chemical neighborhoods” inside their bioengineered yeast cells.

It takes about 17 separate chemical steps to make the opioid compounds used in medicines. Some of these steps occur naturally in poppies, and the remaining ones occur by using synthetic chemical processes in factories. Smolke’s team wants all the steps to happen inside yeast cells within a single vat, including using yeast to carry out chemical processes that poppies never evolved to perform — such as refining opiates like thebaine into more valuable, semi-synthetic opioids like oxycodone.

So Smolke programmed her bioengineered yeast to perform these final industrial steps as well. To do this, she endowed the yeast with genes from a bacterium that feeds on dead poppy stalks. Since she wanted to produce several different opioids, her team hacked the yeast genome in slightly different ways to produce each of the slightly different opioid formulations, such as oxycodone or hydrocodone.

The missing link

All of this is described in the new research paper. But Smolke and her team must still clear one more hurdle in order to achieve the goal of pouring sugar into a stainless steel vat of bioengineered yeast and skimming off specific opioids at the end of the process. They must perform another set of bioengineering hacks to connect the two major advances they have made over the past decade.

This will allow us to create a reliable supply of these essential medicines in a way that doesn’t depend on years leading up to good or bad crop yields.

Remember that it takes about 17 chemical steps to go from poppy to pill. When she began the work in 2004, Smolke started early in the process and went about halfway through these chemical steps. In a 2008 paper, she reported success in that first phase of the project when, her bioengineered yeast produced a precursor to thebaine.

In the new paper, Smolke started with thebaine obtained from poppies, put this into her bioengineered yeast and got refined opioids at the end of the process.

Now her team must extend the 2008 process from sugar to thebaine. Once she forges this missing link in the chain of biochemical synthesis, she will have produced a bioengineered yeast that can perform all 17 steps from sugar to specific opioid drugs in a single vat.

“We are already working on this,” she said.

Smolke said it could take several more years to perfect these last steps in the lab and scale up the process to produce large-sized batches of bioengineered opioids that are pharmacologically identical to the opioids used today.

“This will allow us to create a reliable supply of these essential medicines in a way that doesn’t depend on years leading up to good or bad crop yields,” Smolke said. “We’ll have more sustainable, cost-effective and secure production methods for these important drugs.”

The research was supported by National Institutes of Health; the National Science Foundation; the Bill and Melinda Gates Foundation; the New Zealand Foundation for Research, Science and Technology; and a Stanford Graduate Fellowship.

The Department of Bioengineering, which is jointly operated by the School of Engineering and the School of Medicine, also supported the work.