Yeast produce low-cost, high-quality cannabinoids

February 27, 2019

Science Daily/University of California - Berkeley

Synthetic biologists have created an enzymatic network in yeast that turns sugar into cannabinoids, including tetrahydrocannabinol and cannabidiol, but also novel cannabinoids not found in the marijuana plant itself. The yeast factories would be more environmentally friendly and less energy intensive than growing the plant and separating out the psychoactive and non-psychoactive ingredients. They may also yield cannabinoid derivatives with unexpected medical uses.

University of California, Berkeley, synthetic biologists have engineered brewer's yeast to produce marijuana's main ingredients -- mind-altering THC and non-psychoactive CBD -- as well as novel cannabinoids not found in the plant itself.

Feeding only on sugar, the yeast are an easy and cheap way to produce pure cannabinoids that today are costly to extract from the buds of the marijuana plant, Cannabis sativa.

"For the consumer, the benefits are high-quality, low-cost CBD and THC: you get exactly what you want from yeast," said Jay Keasling, a UC Berkeley professor of chemical and biomolecular engineering and of bioengineering and a faculty scientist at Lawrence Berkeley National Laboratory. "It is a safer, more environmentally friendly way to produce cannabinoids."

Cannabis and its extracts, including the high-inducing THC, or tetrahydrocannabinol, are now legal in 10 states and the District of Columbia, and recreational marijuana -- smoked, vaped or consumed as edibles -- is a multibillion-dollar business nationwide. Medications containing THC have been approved by the Food and Drug Administration to reduce nausea after chemotherapy and to improve appetite in AIDS patients.

CBD, or cannabidiol, is used increasingly in cosmetics -- so-called cosmeceuticals -- and has been approved as a treatment for childhood epileptic seizures. It is being investigated as a therapy for numerous conditions, including anxiety, Parkinson's disease and chronic pain.

But medical research on the more than 100 other chemicals in marijuana has been difficult, because the chemicals occur in tiny quantities, making them hard to extract from the plant. Inexpensive, purer sources -- like yeast -- could make such studies easier.

Plus, he added, there is "the possibility of new therapies based on novel cannabinoids: the rare ones that are nearly impossible to get from the plant, or the unnatural ones, which are impossible to get from the plant."

Keasling, the Philomathia Foundation Chair in Alternative Energy at Berkeley, and his colleagues will report their results online Feb. 27 in advance of publication in the journal Nature.

Plugging chemical pathways into yeast

Cannabinoids join many other chemicals and drugs now being produced in yeast, including human growth hormone, insulin, blood clotting factors and recently, but not yet on the market, morphine and other opiates.

One of the pioneers of synthetic biology, Keasling has long sought to exploit yeast and bacteria as "green" drug factories, eliminating the expensive synthetic or extractive processes common in the chemical industry and the often toxic or environmentally- damaging chemical byproducts.

Cannabis cultivation is a prime example of an energy-intensive and environmentally-destructive industry. Farms in northwest California have polluted streams with pesticide and fertilizer runoff and helped drain watersheds because marijuana plants are water-hungry. Illegal grows have resulted in clear-cutting and erosion.

Indoor cultivation under grow lights with ventilation fans uses a lot of energy, accounting for a growing percentage of annual power consumption. One study estimated that California's cannabis industry accounted for 3 percent of the state's electricity usage. Indoor grows have caused blackouts in some cities, and energy consumption can add more than $1,000 to the price of a pound of weed.

Hence Keasling's interest in finding a "green" way to produce the active chemicals in marijuana.

"It was an interesting scientific challenge," he admitted, that was akin to other challenges he and his team have successfully overcome in yeast: producing an antimalarial drug, artemisinin; turning plant waste into biofuels; synthesizing flavors and scents for the food and cosmetics industries and chemical intermediates for making new materials. "But when you read about cases of patients who have seizures and are helped by CBD, especially children, you realize there is some value in these molecules, and that producing cannabinoids in yeast could really be great."

With approval and oversight by the U.S. Drug Enforcement Agency -- cannabis is still illegal under federal law -- Berkeley postdoc Xiaozhou Luo and visiting graduate student Michael Reiter, who led the project, started assembling in yeast a series of chemical steps to produce, initially, the mother of all cannabinoids, CBGA (cannabigerolic acid). In both marijuana and yeast, the chemical reactions involve the acid form of the compounds: CBGA and its derivatives, THCA and CBDA. They readily convert to CBG, THC and CBD when exposed to light and heat.

Turning yeast into chemical factories involves co-opting their metabolism so that, instead of turning sugar into alcohol, for example, yeast convert sugar into other chemicals that are then modified by added enzymes to produce a new product, such as THC, that the yeast secrete into the liquid surrounding them. The researchers ended up inserting more than a dozen genes into yeast, many of them copies of genes used by the marijuana plant to synthesize cannabinoids.

One step, however, proved to be a roadblock for Keasling's group and competing groups: an enzyme that performs a key chemical step in making CBGA in the marijuana plant didn't work in yeast.

Rather than engineer a different synthetic pathway, Berkeley postdoc Leo d'Espaux and graduate student Jeff Wong went back to the plant itself and isolated a second enzyme, prenyl transferase, that does the same thing, and stuck it in the yeast.

"It worked like gangbusters," Keasling said.

Once they had yeast-producing CBGA, they added another enzyme to convert CBGA to THCA and a different enzyme to create a pathway to CBDA. Though the products the yeast produce are predominantly THC or CBD, Keasling said, each must still be separated from other chemicals present in tiny quantities.

They also added enzymes that made the yeast produce two other natural cannabinoids, CBDV (cannabidivarin) and THCV (tetrahydrocannabivarin), whose effects are not well understood.

Surprisingly, Xiaozhou and Michael discovered that the enzymatic steps involved in making CBGA in yeast are flexible enough to accept a variety of starter chemicals -- different fatty acids in place of the one used by the marijuana plant, hexanoic acid -- that generate cannabinoids that do not exist in the plant itself. They also got the yeast to incorporate chemicals into cannabinoids that could later be chemically altered in the lab, creating another avenue for producing never-before-seen, but potentially medically useful, cannabinoids.

Keasling subsequently founded an Emeryville, California, company, Demetrix Inc., which d'Espaux and Wong later joined, that licensed the technology from Berkeley to use yeast fermentation to make cannabinoids.

"The economics look really good," Keasling said. "The cost is competitive or better than that for the plant-derived cannabinoids. And manufacturers don't have to worry about contamination -- for example, THC in CBD -- that would make you high."

https://www.sciencedaily.com/releases/2019/02/190227131838.htm

Cannabis: A new frontier in therapeutics

February 15, 2015

Science Daily/McGill University Health Centre

While debate about recreational marijuana use continues, researchers are investigating the effectiveness of cannabis for treating pain, spasticity, and a host of other medical problems. In a symposium organized by the McGill University Health Centre (MUHC) as part of the 2015 American Association for the Advancement of Science Annual Meeting held this week in San Jose, California, experts from North America and the U.K. share their perspectives on the therapeutic potential of medical cannabis and explore the emerging science behind it.

"We need to advance our understanding of the role of cannabinoids in health and disease through research and education for patients, physicians and policy-makers," says Dr. Mark Ware, director of clinical research at the Alan Edwards Pain Management Unit at the MUHC, in Canada.

As a pain specialist Dr. Ware regularly sees patients with severe chronic pain at his clinic in Montreal, and for some of them, marijuana appears to be a credible option. "I don't think that every physician should prescribe medical cannabis, or that every patient can benefit but it's time to enhance our scientific knowledge base and have informed discussions with patients."

Increasing numbers of jurisdictions worldwide are allowing access to herbal cannabis, and a range of policy initiatives are emerging to regulate its production, distribution, and authorization. It is widely believed that there is little evidence to support the consideration of cannabis as a therapeutic agent. However, several medicines based on tetrahydrocannabinol (THC), the psychoactive ingredient of cannabis, have been approved as pharmaceutical drugs.

Leading British cannabis researcher Professor Roger Pertwee, who co-discovered the presence of tetrahydrocannabivarin (THCV) in cannabis in the 70's, recently published with collaborators some findings of potential therapeutic relevance in the British Journal of Pharmacology. "We observed that THCV, the non-psychoactive component of cannabis, produces anti-schizophrenic effects in a preclinical model of schizophrenia," says Pertwee, professor of Neuropharmacology at Aberdeen University. "This finding has revealed a new potential therapeutic use for this compound."

Neuropsychiatrist and Director of the Center for Medicinal Cannabis Research (CMCR) at the University of California, San Diego Dr. Igor Grant is interested in the short and long-term neuropsychiatric effects of marijuana use. The CMCR has overseen some of the most extensive research on the therapeutic effects of medical marijuana in the U.S. "Despite a commonly held view that cannabis use results in brain damage, meta analyses of extensive neurocognitive studies fail to demonstrate meaningful cognitive declines among recreational users," says Dr. Grant. "Bain imaging has produced variable results, with the best designed studies showing null findings."

Dr. Grant adds that while it is plausible to hypothesize that cannabis exposure in children and adolescents could impair brain development or predispose to mental illness, data from properly designed prospective studies is lacking.

https://www.sciencedaily.com/releases/2015/02/150215070209.htm