Scientists from Brown University say they have demonstrated a novel method for stimulating autophagy. In a study (“Nuclear Export Inhibition Enhances HLH-30/TFEB Activity, Autophagy, and Lifespan”), published in Cell Reports, the team shows that the approach increased the lifespans of worms and flies, and experiments in human cells hint that the strategy could be useful in future treatments for Alzheimer's disease, amyotrophic lateral sclerosis (ALS), and other age-related neurodegenerative conditions.
“Transcriptional modulation of the process of autophagy involves the transcription factor HLH-30/TFEB. In order to systematically determine the regulatory network of HLH-30/TFEB, we performed a genome-wide RNAi [RNA interference] screen in C. elegans and found that silencing the nuclear export protein XPO-1/XPO1 enhances autophagy by significantly enriching HLH-30 in the nucleus, which is accompanied by proteostatic benefits and improved longevity. Lifespan extension via xpo-1 silencing requires HLH-30 and autophagy, overlapping mechanistically with several established longevity models. Selective XPO1 inhibitors recapitulated the effect on autophagy and lifespan observed by silencing xpo-1 and protected ALS-afflicted flies from neurodegeneration,” write the investigators.
“XPO1 inhibition in HeLa cells enhanced TFEB [transcription factor EB] nuclear localization, autophagy, and lysosome biogenesis without affecting mTOR [mechanistic target of rapamycin] activity, revealing a conserved regulatory mechanism for HLH-30/TFEB. Altogether, our study demonstrates that altering the nuclear export of HLH-30/TFEB can regulate autophagy and establishes the rationale of targeting XPO1 to stimulate autophagy in order to prevent neurodegeneration.”
“Autophagy dysfunction is present across a range of age-related diseases, including neurodegeneration,” said Louis Lapierre, Ph.D., an assistant professor of molecular biology, cell biology, and biochemistry at Brown who led the work. “We and others think that by learning how to influence this process pharmacologically, we might be able to affect the progression of these diseases. What we've shown here is a new and conserved entry point for stimulating autophagy.”
Autophagy involves the rounding up of misfolded proteins and obsolete organelles within a cell into autophagosomes which then fuse with a lysosome that breaks down those cellular macromolecules and converts it into components the cell can reuse.
Dr. Lapierre and his colleagues wanted to see if they could increase autophagy by manipulating a transcription factor that regulates autophagic activity. In order for the transcription factor to switch autophagic activity on, it needs to be localized in the nucleus of a cell. So Dr. Lapierre and his team screened for genes that enhance the level of the autophagy transcription factor, known as TFEB, within nuclei.
Using the nematode Caenorhabditis elegans, the screen found that reducing the expression of a protein called XPO1, which transports proteins out of the nucleus, leads to nuclear accumulation of the nematode version of TFEB. That accumulation was associated with an increase in markers of autophagy, including increased autophagosome and autolysosomes, as well as increased lysosome biogenesis. There was also a marked increase in lifespan among the treated nematodes of between about 15% and 45%.
“What we showed was that by blocking the escape of this transcription factor from the nucleus, we could not only influence autophagy but we could get an increase in lifespan as well,” Dr. Lapierre said.
The next step was to see if there were drugs that could mimic the effect of the gene inhibition used in the screening experiment. The researchers found that selective inhibitors of nuclear export (SINE), originally developed to inhibit XPO1 to treat cancers, had a similar effect, i.e., increasing markers of autophagy and significantly increasing lifespan in nematodes.
The researchers then tested SINE on a genetically modified fruit fly that serves as a model organism for the neurodegenerative disease ALS. Those experiments showed a small but significant increase in the lifespans of the treated flies. “Our data suggests that these compounds can alleviate some of the neurodegeneration in these flies,” noted Dr. Lapierre.
As a final step, the researchers set out to see if XPO1 inhibition had similar effects on autophagy in human cells as it had in the nematodes. After treating a culture of human HeLa cells with SINE, the researchers found that, indeed, TFEB concentrations in nuclei increased, as did markers of autophagic activity and lysosomal biogenesis.
“Our study tells us that the regulation of the intracellular partitioning of TFEB is conserved from nematodes to humans and that SINE could stimulate autophagy in humans,” continued Dr. Lapierre. “SINE have been recently shown in clinical trials for cancer to be tolerated, so the potential for using SINE to treat other age-related diseases is there.”
The scientists will now focus on testing these drugs in more clinically relevant models of neurodegenerative diseases. But this initial research is a proof of concept for this strategy as a means to increase autophagy and potentially treat age-related diseases, according to Dr. Lapierre