. A Small Molecule Screen in Stem-Cell-Derived Motor Neurons Identifies a Kinase Inhibitor as a Candidate Therapeutic for ALS. Cell Stem Cell. 2013 Jun 6;12(6):713-26. PubMed.


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  1. This is an exciting paper showing the ability to use a specific class of stem cell-derived neurons—motor neurons—to screen for potential neuroprotective small molecules. While the mouse ESC-derived neural cultures appear suitable for systematic screening, the authors have been able to then evaluate selected hits on human motor neurons derived from ESC- as well as control- and patient-derived iPSC cultures. I was delighted to see that olesoxime was used as a comparator and clearly promoted survival of trophic factor-deprived human motor neurons with similar potency as we found using trophic factor-deprived primary rodent motor neurons. Even though it appears less active than the selected hit, kenpaullone, the large variability in the response to trophic factors, the gold standard, makes it hard to define a maximum response or calibrate the relative response of compounds in this assay. Having human-derived neurons to use as a way of selecting compounds is an important advance for CNS drug discovery, which suffers exceptionally from the high translational risk associated with current animal models. It may still be difficult to define how activity in this model will translate in terms of clinical significance, particularly for such a rapidly progressing neurodegenerative disease as ALS. Despite its activity in these cell assays, olesoxime did not have a statistically significant benefit over riluzole in an 18-month trial in ALS patients. One wonders what effect riluzole, the only approved drug for ALS, might have in this model?

    In terms of technical achievement, this paper is a real tour de force. A screening assay was possible using mouse-derived GFP-expressing motor neurons, which allowed the researchers to overcome a number of challenges. Motor neurons represent typically only 30-50 percent of the cells in the mixed cultures (GFP expression is driven by a motor neuron-specific promoter), and new progenitor-derived motor neurons can appear during the assay, giving rise to false positives. Fluorescent-activated cell sorting showed that the compounds acted directly on motor neurons.

    The authors selected trophic factor deprivation to trigger robust and rapid motor neuron death and stringently selected hits from a 5,000-compound library based on activity in two separate assays derived from “control” or mutant SOD1 mouse motor neurons. Testing in a number of secondary assays (PI3K/Akt inhibition, "toxic astrocytes," morphology, functionality) further classified those hits, and a number of compounds affecting known “survival promoting pathways” were identified. The authors then focused on one of these, kenpaullone, and carefully dissected its mechanism of action as an inhibitor of a kinase cascade culminating in c-Jun-activated apoptosis and fingered a particular kinase as a potential drug target for ALS. Interestingly, kenpaullone also suppressed expression of mutant SOD1 with longer treatment. Further analysis of the role of this kinase cascade and apoptosis pathway in normal and pathological processes, as well as other off-target effects of this class of compounds, will be needed to evaluate the risk/benefit of targeting this pathway and to base a chemical optimization strategy on this compound family. Nevertheless, despite questions about the target itself, the use of human- and especially patient-derived neural cells is an important addition to CNS drug discovery to help bridge the preclinical-clinical translational gap.

    Regarding the stem cell motor neuron screening models, one could envisage testing combinations of compounds with different mechanisms of action to see if their effects are additive, or even synergistic, and potentially define a mixture that would provide maximal activity. Most diseases are best treated with combination therapies. Finding a regulatory pathway to test two or more investigational drugs could be the way forward to find effective treatments for neurodegenerative diseases.

    It is intriguing to think that olesoxime might provide additional benefit to compounds acting on the kenpaullone-activated pathway, as its mechanism of action is probably different. We have recently compared olesoxime's mechanism of activity with BDNF, a trophic factor (see Gouarné et al., 2013). Using a cortical neuron neurotoxicity model, we showed that olesoxime did not activate the PI3K/Akt pathways but appears to have a direct effect to stabilize mitochondria and prevent release of apoptotic factors—olesoxime binds to two outer mitochondrial membrane proteins, VDAC and TSPO. Taking all the data on olesoxime accumulated by ourselves and others, it appears to have clear neuroprotective effects. Trophos believes it has a place for the treatment of neurodegenerative diseases, especially where treatment could be started earlier, before neurodegeneration is advanced and accelerating, as in genetic or neuroinflammatory conditions . For this reason, olesoxime is currently being tested in a Phase 2 study in type 2 and type 3 SMA patients and a Phase 1b study in relapsing remitting multiple sclerosis patients as an add-on to β interferon (both trials are registered on ClinicalTrials.gov).


    . Olesoxime protects embryonic cortical neurons from camptothecin intoxication by a mechanism distinct from BDNF. Br J Pharmacol. 2013 Apr;168(8):1975-88. PubMed.

    View all comments by Rebecca M. Pruss

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  1. Stem Cell Screen Points to ALS Disease Target