Possible New Target for Treating Brain Inflammation

March 2015

A team of TSRI scientists led by Benjamin F. Cravatt, chair of TSRI's Department of Chemical Physiology and member of TSRI's Skaggs Institute for Chemical Biology, set out to investigate PHARC, a rare and mysterious inherited disorder that was first described by Norwegian researchers in 2009.

“This finding is a good example of what can be gained from studying enzymes linked to rare human genetic disorders,” says Benjamin Cravatt, chair of TSRI's Department of Chemical Physiology.

Their investigation led to the identification of an enzyme that produces a class of inflammatory lipid molecules in the brain that may cause PHARC when present at abnormally high levels; the discovery may prove useful in treating PHARC and more common conditions that involve brain inflammation – including multiple sclerosis, Alzheimer's, Parkinson's, ALS, and secondary damage after stroke and head injuries. Such inflammation often fails to respond to standard anti-inflammatory drugs.

“This finding is a good example of what can be gained from studying enzymes linked to rare human genetic disorders,” said Dr. Cravatt.

Named for its unique set of typical symptoms (polyneuropathy, hearing loss, ataxia, retinitis pigmentosa, and cataract), PHARC usually manifests in early adolescence and progressively worsens with age. In 2010, PHARC was linked to gene mutations that inactivate the enzyme ABHD12.

The finding prompted Dr. Cravatt and his laboratory to develop a mouse model of the disease in which the ABHD12 gene is similarly inactivated. Studies of those “PHARC mice” revealed that the ABHD12 enzyme normally breaks down immune-signaling lipid molecules called lysophosphatidylserines (lyso-PSs) in the brain. ABHD12's absence in the PHARC mice leads to an abnormal buildup of lyso-PSs and subsequent neuroinflammation.

Having identified an enzyme that normally breaks down lyso-PS molecules, Dr. Cravatt and his laboratory then set out to find an enzyme that makes lyso-PSs. In principle, such an enzyme could be targeted with compounds that inhibit its activity to dial down lyso-PS levels in the nervous system and thus treat PHARC.

To find a lyso-PS-making enzyme, Cravatt lab Research Associate Siddhesh S. Kamat led the effort to develop a sophisticated test for detecting the enzyme-mediated conversion of precursor phosphatidylserine (PS) molecules to lyso-PSs. “Using this test we discovered a cryptic yet distinct lyso-PS-making enzyme activity in the mouse brain,” said Dr. Kamat.

The tests revealed especially high levels of this activity in the cerebellum, a brain region strongly impacted in PHARC. The team also found that this lyso-PS-making activity could be powerfully blocked in the lab dish by the weight-loss drug tetrahydrolipstatin (THL, also known as Orlistat and Xenical). THL is a known inhibitor of multiple enzymes, and the team was able to link the lyso-PS-making activity in mouse brains specifically to one of these enzymes: a previously uncharacterized enzyme called ABHD16A.

Dr. Cravatt's team then began a collaboration with the laboratory of chemist Amy R. Howell at the University of Connecticut to find a more potent and selective inhibitor of ABHD16A. The team eventually isolated a small-molecule compound called KC01 that disrupts ABHD16A activity in mammalian cells more selectively than THL does.

The team found that blocking ABHD16A activity with KC01 markedly reduced secreted lyso-PS levels in culture and brought elevated lyso-PS levels back down in cells derived from PHARC patients. The treatment also greatly reduced the secretion of inflammatory compounds by PHARC-mouse immune cells following exposure to a bacterial toxin.

Dr. Cravatt hopes to conduct future research in collaboration with the Howell laboratory to develop a better ABHD16A inhibitor that, unlike THL and KC01, can reach the brain from the bloodstream and thus be tested in live mice – and perhaps someday in PHARC patients. “We also think there is a potential for applying the lyso-PS-lowering strategy more broadly against neurological and immunological disorders,” said Dr. Cravatt.

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