Everolimus Not Neuroprotective in the R6/2 mice

Dr. Steven Hersch and colleagues have investigated the effect of the rapamycin derivative, everolimus, on the R6/2 mice.  While there was improvement in rotarod performance and the drug did penetrate the brain, it failed to protect neurons.  Everolimus reduced the HD protein in skeletal muscle tissue but not in the brain where it did not activate autophagy.

Here's another major article that adds to our information about autophagy.

Dr. Steven Hersch and colleagues have investigated the effect of the rapamycin derivative, everolimus, on the R6/2 mice.  While there was improvement in rotarod performance and the drug did penetrate the brain, it failed to protect neurons.  Everolimus reduced the HD protein in skeletal muscle tissue but not in the brain where it did not activate autophagy.

Everolimus is an inhibitor of mTOR (mammalian target of rapamycin).  Inhibiting mTOR has been previously found to lead to an increase in macroautophagy (commonly referred to as autophagy) which is of interest to HD researchers as a possible way to clear away the HD protein which accumulates in cells.  Autophagy is a very old cellular house cleaning process found in organisms from yeast to mammals. Damaged parts of the cell, pathogens, and large proteins are surrounded by autophagosomes. The autophagosomes deliver their cargo to the lysosomes by fusing with them. The lysosomes then consume the material.

Everolimus is approved for advanced kidney disease and as an immunosuppressant for transplant patients.  The researchers were interested in finding out if this drug would be an effective treatment for Huntington’s disease since it is already available for these other purposes.

Everolimus was found to have penetrated the brain as measured by decreased phosphorylation of the mTOR target protein S6 kinase.  However, autophagy was not upregulated in the brain as it was in the muscles as measured by normalized levels of the cystolic protein LC3BII which is associated with autophagosomes.  Levels of soluble HD protein were reduced in muscle tissue but not in the brain.  No signs of neuroprotection were found as measured by brain weight, striatal volume, or striatal neuronal cell body volume. Rotarod performance, a standard measure of motor ability was improved on everolimus.  The authors suggest that the positive results in rotarod performance were due to increased autophagy in the muscle tissues.  Survival data was not reported.

These results point to the need to closely examine the potential of autophagy as a treatment which has looked promising since a 2004 study by Rubinsztein and colleagues.  In that study, aggregate load was reduced and survival time increased in a cell model and in a drosophila model.   Improvement was found in rotarod performance, grip strength, the wire maneuver test, and tremors in an N171-82Q HD mouse model.  Aggregates were also reduced.  However, because the study was conducted in the UK where regulations require the mice to be euthanized before end stage disease, there were no survival data.

In 2010, Rubinsztein and colleagues investigated the effects of rilmenidine on the N171-82Q mice.  Rilmenidine, a drug which is FDA approved for hypertension, gets into the brain, has been safe for long term use, and was previously shown in a screening to induce autophagy through an M-tor independent pathway. The researchers found that rilmenidine reduced levels of the solutble HD protein but not aggregates in the brain.  Mice treated with the drug had improved grip strength, did better on the wire maneuver test and had less severe tremors but did not improve on the rotarod test.  Again, survival data could not be obtained.

The mouse modes are different in the Rubinsztein studies and in the Hersch study. Rubinsztein’s N171-82Q HD mice are slower to develop symptoms and were treated before symptom onset while the R6/2 mice which have rapid onset of symptoms were treated after symptoms began in the Hersch study.  The lack of survival time data and the discrepancies between the studies suggest that more questions need to be answered before autophagy inducers go into clinical trials. 

Additional research is needed to determine whether autophagy inducers can increase survival time.  Also raising concerns is the recent research by Dr. Ana Maria Cuervo and colleagues who found that that cargo recognition was impaired in Huntington's disease and cellular garbage wasn't getting to the lysosomes.  Does autophagy clear away the HD protein or is the process itself impaired in Huntington’s Disease?  If so, at what point in the disease process does the impairment occur? Is there a window of time in which autophagy inducement would be an effective treatment?

We will continue to report on preclinical studies of drugs which induce autophagy as they are published.

Other references:

G Oroz, Francesco Scaravilli, Douglas F Easton, Rainer Duden, Cahir J O'Kane, and David C Rubinsztein.  “Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease.” Nature Genetics 2004 Jun;36(6):585-95.

Claudia Rose,  Fiona M. Menzies, Maurizio Renna, Abraham Acevedo-Arozena, Silvia Corrochano, Oana Sadiq, Steve M. Brown and David C. Rubinsztein. “Rilmenidine attenuates toxicity of polyglutamine expansions in a mouse model of Huntington's disease.” Human Molecular Genetics 2010 Junw 1;19 (11):2144-53.

Marta Martinez-Vicente, Zsolt Talloczy, Esther Wong, Guomei Tang, Hiroshi Koga, Susmita Kaushik, Rosa de Vries, Esperanza Arias, Spike Harris, David Sulzer & AnaMaria Cuervo. “ Cargo recognition failure is responsible for inefficient autophagy in Huntington’s disease.” Nature Neuroscience 2010 Apr 11. [Epub ahead of print]

Steven Hersch, M.D., Ph.D.

The mTOR kinase inhibitor Everolimus decreases S6 kinase phosphorylation but fails to reduce mutant huntingtin levels in brain and is not neuroprotective in the R6/2 mouse model of Huntington's diseas

Jonathan H Fox, Teal Connor, Vanita Chopra, Kate Dorsey, Jibrin A Kama, Dorothee Bleckmann, Claudia Betschart, Daniel Hoyer, Stefan Frentzel, Marian DiFiglia, Paolo Paganetti, and Steven M. Hersch

abstract

Background:

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a CAG repeat expansion within the huntingtin gene. Mutant huntingtin protein misfolds and accumulates within neurons where it mediates its toxic effects. Promoting mutant huntingtin clearance by activating macroautophagy is one approach for treating Huntington's disease (HD). In this study, we evaluated the mTOR kinase inhibitor and macroautophagy promoting drug everolimus in the R6/2 mouse model of HD.

Results:

Everolimus decreased phosphorylation of the mTOR target protein S6 kinase indicating brain penetration. However, everolimus did not activate brain macroautophagy as measured by LC3B Western blot analysis. Everolimus protected against early declines in motor performance; however, we found no evidence for neuroprotection as determined by brain pathology. In muscle but not brain, everolimus significantly decreased soluble mutant huntingtin levels.

Conclusions:

Our data suggests that beneficial behavioral effects of everolimus in R6/2 mice result primarily from effects on muscle. Even though everolimus significantly modulated its target brain S6 kinase, this did not decrease mutant huntingtin levels or provide neuroprotection.