New Research: Rapamycin prevents dopaminergic neuron loss in PD models

Dr. Alex Whitworth describes his latest findings, demonstrating rapamycin can prevent dopaminergic loss and PD pathology using Drosophila and mammalian PD models through activation of the translation inhibitor 4E-BP. -Holli Kawadler

Can you summarize your findings and your next steps?

AW: Here we have used unbiased genetic screening techniques to identify a modifiers of manifestations of Parkinson disease associated pathology in a simple model animal system. We have identified a potent protective pathway that can be activated to provide a remarkable degree of protection in a number of model system. Furthermore, we have used a known chemical modulator of that pathway to achieve a significant degree of protection in both animal model systems and cells from PD patients. Importantly, we saw complete suppression of dopaminergic neurodegeneration two genetic animal models.

There are a number of obvious extensions to this work. For instance, it will be important to demonstrate the efficacy of rapamycin in a more complex model system such as a mouse model, in particular a model that reflects a greater proportion of all PD cases such as the newly described LRRK2 BAC transgenic mouse model. It would also be nice to test the effects of rapamycin in an representative model of sporadic PD but arguably none such exists to date. When a suitable pre-clinical model has been tested, it would also be essential to optimise the administration of rapamycin, not least to find the optimal effective dose but also to avoid unwanted side-effects known to occur with rapamycin.

Another major avenue that this work highlights is the opportunity to use these model systems to screen for novel compounds that can provide the same level of neuroprotection. Alternatively, one could specifically target the 4E-BP pathway for chemical modulators that may provide better efficacy than rapamycin and eliminate the detrimental side-effects.

The Thor (4EBP) allele came from a screen of parkin modifiers and you show that increasing 4EBP expression is protective in this system, as you might predict.  But you also show that loss of the fly LRRK2 homologue also has the same effect, of increasing 4EBP expression and can rescue the parkin or PINK1 deficient flies.  Does this mean that LRRK2 and PINK1/parkin are in the same pathway?  Where would you place parkin/PINK1 and LRRK2 relative to each other?

AW: Although genetic interactions in Drosophila have very nicely shown that Parkin and PINK1 functionally interact and that Parkin acts downstream of PINK1, I don't think the same conclusion can be drawn in this case for LRRK2; that is, I do not think it is likely that LRRK2 acts in a common pathway with PINK1/Parkin. For instance, the mutant phenotypes of parkin and PINK1 are remarkably similar in Drosophila while LRRK2  mutants do not resemble these.  Presently the full range of genetic interaction studies that would support LRRK2 functioning in the same pathway as PINK1/Parkin has not been completed. As we argue in the paper, I think these results reflect the potential effect of LRRK2 on 4E-BP function in a parallel pathway from Parkin and PINK1, but which can converge downstream of the defects seen in parkin/PINK1 mutants.

The rescue in mammalian cells of mitochondrial function by rapamycin (figure 5c) is fairly modest and the effects in flies are incomplete for functional measures (eg 4b).  Does this have implications for how effective we think treatment of PD patients might be?  Is there another way to approach the problem that might be more impactful?

AW: Taking this data set alone, the effect of rapamycin has shown some variable or incomplete rescue, although always statistically significant, and remarkably rapamycin was able to completely suppress the key feature of PD pathology - dopaminergic neurodegeneration. We could try to rationalise why these variabilities occur; for example, the widespread destruction of flight muscle tissue or the rather non-specific depolarisation of mitochondria cannot be rescued whereas the more subtle phenotypes of partial dopaminergic neuron loss or mitochondrial morphology defects that can be fully restored, may simply reflect the potency of the protective effect of rapamycin in our assay system. It is important to remember that these experiments have not ben conducted to try to optimise the action of rapamycin in vitro or in vivo. We conducted our experiments guided by previously reported use of rapamycin in other systems. Also, no quantification has yet been made of the pharmacokinetics of rapamycin administered in our assay system. One obvious area for improvement would be to determine the optimum efficiency of rapamycin in vitro and in vivo in long term neuroprotection.

However, it is important to realise that what we have shown here is the identification of a potent protective cellular mechanism, regulated protein translation, and the successful and beneficial effect of a drug already known to modulate that pathway. This link paves the way to use these and other models systems to test new and potentially better, more effective drugs with the same overall mechanism of action. 

I think this work is important in highlighting the enormous potential that the use of relatively simple model organisms such as Drosophila can offer for the drug discovery process. Not only was the identification of 4E-BP and protein translation mechanisms made through genetic screening in flies, but we have shown that potentially protective pathways in flies are also extremely relevant to human pathology. 

Luke S Tain, Heather Mortiboys, Ran N Tao, Elena Ziviani, Oliver Bandmann & Alexander J Whitworth. Rapamycin activation of 4E-BP prevents parkinsonian dopaminergic neuron loss. Nature Neuroscience Published online: 16 August 2009 | doi:10.1038/nn.2372

Dietary restriction has been associated with a number of beneficial effects, most notably extending lifespan, slowing cancer progression, and neuroprotection. A recent publication in Cell delves into the mechanism of dietary restriction, identifying 4E-BP, a translation inhibitor and substrate of mTOR, as an essential player in extending lifespan (Zid et al., 2009). However, given the recent association between 4E-BP and neuroprotection - preventing dopaminergic loss in Parkinson's disease models (Tain et al., 2009) - this work rekindles an interest in dietary restriction and PD, identifying a potential mechanism for neuroprotection.

Zid et al. used a genome-wide screen identifying translational changes in Drosophila upon dietary restriction. Their results indicate 4E-BP is upregulated upon dietary restriction and mediates transcriptional upregulation of mitochondrial genes (including Complex I and Complex IV) to play an important role in extending lifespan. Previous work by Dr. Alex Whitworth and colleagues demonstrated that treatment with rapamycin (the mTOR inhibitor) resulted in 4E-BP activation and subsequent protection of dopaminergic neurons in a Drosophila  PD model. Both act via the mTOR signaling pathway (responsive to energy levels or via direct inhibition), begging the question of whether the mechanism identified in response to dietary restriction (regulation of mitochondrial genes) is also the mechanism leading to neuroprotection.

Stepping back, there are a number of reports demonstrating dietary restriction-induced neuroprotection, specifically in Parkinson's disease models. In 1999, Duan and Mattson showed dietary restriction is protective in the mouse MPTP model. Notably, in 2004 Maswood et al. demonstrated caloric restriction can decrease neurochemical and behavioral deficits and increase GDNF and BDNF in a nonhuman primate model of Parkinson's disease. Since then, there has been little progress further investigating protective factors of dietary restriction for Parkinson's disease. In fact, recent work showed dietary restriction was not protective in the 6-OHDA model (Armentero et al., 2008).

Given these conflicting reports, have these results been replicated either way? Is anyone currently working on investigating caloric restriction and Parkinson's disease? Are there other negative data out there disproving neuroprotection from dietary restriction? From a clinical perspective, what about an association between eating habits and PD risk? Is there a relationship between BMI and PD?

Brian M. Zid, Aric N. Rogers, Subhash D. Katewa, Misha A. Vargas, Marysia C. Kolipinski, Tony Au Lu, Seymour Benzer, Pankaj Kapahi. 4E-BP Extends Lifespan upon Dietary Restriction by Enhancing Mitochondrial Activity in Drosophila. Cell. 139, 149-160 (2009)