New Tamoxifen Analog Reduces Amphetamine Neurochemical and Behavioral Effects

Colleen Carpenter
Colleen Carpenter

Amphetamines are the second most abused drug worldwide, but no effective treatments are available for amphetamine abuse. At this year’s Experimental Biology meeting, graduate student Colleen Carpenter from the University of Michigan presented data revealing that the chemotherapy agent tamoxifen can serve as a platform for the creation of a pharmacological treatment against amphetamine abuse.

Currently, the mainstay of amphetamine abuse treatment is psychotherapy. “The problem with behavioral treatment methods is that they are not very effective,” Carpenter said. “We’re looking at a pharmacological way to block amphetamine abuse.” In the lab of Dr. Margaret Gnegy at the University of Michigan Department of Pharmacology, Carpenter is looking for pharmacological treatments to inhibit the rewarding and reinforcing effects of amphetamine.

The effects of amphetamine are caused by the release of the neurotransmitter dopamine into the synapse via the dopamine transporter in the brain. “People have actually tried to target the dopamine transporter directly,” says Carpenter. “The problem is, many ligands that bind to the dopamine transporter act as dopamine uptake blockers. Blocking uptake of dopamine is bad, because that’s how cocaine works. We still have the increase in extracellular dopamine at the synapse which has its own reward functions.” The Gnegy lab is taking an alternative approach. Rather than targeting the dopamine transporter, they are now looking at the upstream regulator of the dopamine transporter, protein kinase C.

It is known that amphetamine activates protein kinase C, or PKC, which ultimately enhances the release of dopamine through the dopamine transporter. By modulating the activity of PKC, perhaps dopamine release could be reduced, decreasing the rewarding and reinforcing effects of amphetamine.

There are numerous available inhibitors of PKC but the selective estrogen receptor modulator tamoxifen is the only PKC inhibitor known to cross the blood-brain barrier.   Tamoxifen acts at a regulatory subunit of PKC, so it causes less problems than one that acts at the active site.

Courtesy of Gnegy Lab
Courtesy of Gnegy Lab

At this year’s ASPET meeting, Carpenter reveals recent data suggesting that a compound modeled after tamoxifen called C091, designed and synthesized by the Vahlteich Medicinal Chemistry Core at the University of Michigan, is able to reduce PKC activity and dopamine release without affecting the estrogen receptor at relevant concentrations. In addition, behavioral studies with the new compound showed significant decreases in amphetamine-related behavior in mice.

First, Carpenter determined that C091 is a better inhibitor of PKC activity than tamoxifen, as indicated by changes in the ability of PKC to phosphorylate specific protein targets when treated with C091 versus tamoxifen.

She then investigated the potential for C091 to bind estrogen receptors, as this nonspecific binding could cause unwanted side effects if C091 is purposed as a therapeutic against amphetamine abuse. To determine this, she performed an estrogen receptor competitive binding assay. When compared with estradiol and tamoxifen, C091 did not bind to the estrogen receptor at concentrations where it effectively inhibited PKC.

To determine changes in actual dopamine release at the synapse, Carpenter measured the levels of dopamine release induced by amphetamine with and without treatment with various doses of C091. In order to measure dopamine release in vitro, the ends of dopamine transporter-expressing nerve terminals from the striatum are pinched off to form small spheres called synaptosomes. These small spheres are able to respond to amphetamine exposure and secrete dopamine via the dopamine transporter. A suprafusion system is used to collect the dopamine released from the synaptosomes. In short, this system allows the controlled perfusion of drug solutions over the synaptosomes and the dopamine-containing eluent is collected in fractions for quantification. Treatment with amphetamine stimulated dopamine release from striatal synaptosomes. However, dopamine efflux was reduced with increasing doses of C091.

After proving that C091 inhibits PKC activity and reduces dopamine release, Carpenter moved on to a critical experiment: would C091 have actual effects on amphetamine- induced behavior in an animal? For this experiment, mice were pretreated with C091 followed by amphetamine treatment, and changes in locomotion were observed. Normally, amphetamine exposure significantly increases mouse locomotion due to its action in the brain. When mice were treated with C091 systemically prior to amphetamine, their locomotion was greatly reduced.

Future studies with C091 will include a model in which the effects of C091 on amphetamine self-administration can be determined, as well as microdialysis for measurement of dopamine levels in vivo.

To read more about the research performed in the Gnegy lab, click here.

For more information about the creation of tamoxifen analogs, check out

de Medina, P., Favre, G. & Poirot, M. Multiple targeting by the antitumor drug tamoxifen: a structure-activity study. Current medicinal chemistry. Anti-cancer agents 4, 491-508 (2004).


Author: ilovebraaains

I am a neuroscientist using zebrafish to study mechanisms of neuroregeneration.

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