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multipotent stem cells hold great promise for regenerative medicine, although small molecules that

control their actions are difficult to identify. Screens are usually performed in vitro using cultured cells1, and positive hits have generally not translated well in vivo. Recently in Cell, Pieper et al. describe a novel in vivo screen in mammals to identify compounds that selectively enhance production of neurons from neural stem cells (NSCs) in situ2. One compound was found to prevent the death of newborn neurons by preserving mitochondrial integrity. This, in turn, enabled their functional incorporation into the brain circuitry and reversed both severe developmental defects and age-related loss of cognition in multiple rodent models in vivo.

Small-molecule probes of NSC function are highly desirable but have proven challenging to discover because of the massive complexity of the mammalian brain. In the rodent brain (and the brains of organisms from finches to humans), new neurons are continually generated in two regions, the subventricular zone lining the lateral ventricles and the subgranular zone (SGZ) of the dentate gyrus in the hippocampus (Fig. 1a)3. The hippocampus functions in memory formation and retrieval, and mounting evidence suggests that the continual birth of new neurons is critical for these functions4. Neuronal production is a stem cell–driven process, and understanding how NSCs are regulated is the subject of intense research. NSC abnormalities are present in diverse pathologies including epilepsy and neurodegeneration3. Screens for small molecules that can manipulate the proliferation and differentiation of NSCs and their offspring have been limited to cell lines5 and ex vivo assays6, and hits generally have unfavorable pharmacokinetics for in vivo use. The report by Pieper et al. elegantly sidesteps these roadblocks to reveal a previously unknown compound with direct translational and clinical potential.

Pieper et al. first performed an in silico screen to identify 1,000 compounds with favorable pharmacokinetic properties.

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your brain on drugsManipulation of stem cells is an important therapeutic goal that has proven difficult to achieve. A recent report describes a novel in vivo small-molecule screen and identifies a modulator of mammalian neurogenesis that partially reverses age-related declines in cognition.

Jonathan P Saxe

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Figure 1 | An in vivo screen for proneurogenic small molecules. (a) Neurogenesis in the dentate gyrus of the hippocampus. Stem cells proliferate and give rise to neuroblasts (red) within the subgranular zone (SGZ) of the dentate gyrus in the hippocampus. Many neuroblasts are pruned by apoptosis. Within a few days, surviving neuroblasts (orange) migrate into the granular zone and begin to mature. Over a few weeks, they terminally differentiate into mature granular neurons (yellow), sending dendrites into the molecular layer of the dentate gyrus and extending projections toward other regions of the hippocampus. (b) An in vivo screen for proneurogenic compounds. Mice were infused with pools of ten compounds over the course of one week, with daily BrdU injections to monitor proliferation and survival of stem cells and their progeny. P7C3 was found to greatly increase the number of BrdU-labeled cells within the SGZ. (c) Effects of systemic treatment with P7C3. Treatment with P7C3 was found to promote mitochondrial integrity under normally apoptotic conditions (top). In a rat model of aging, P7C3 promoted survival of newborn neurons (middle) and was associated with enhanced function in the Morris water maze test of learning and memory (bottom).

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640 nature chemical biology | VOL 6 | SEPTEMBER 2010 | www.nature.com/naturechemicalbiology

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Compounds were pooled into groups of 10 and infused into brains of mice for one week (Fig. 1b). Effects on cell proliferation and survival within the SGZ were monitored, and after successful deconvolution of the pools by retesting individual compounds, eight distinct proneurogenic molecules were identified. One compound, P7C3, showed a highly favorable pharmacological profile and could be formulated for multiple routes of administration.

P7C3 and derivatives designed through structure-activity relationships promoted survival of newborn neurons, rather than NSC proliferation and differentiation per se. To examine this further, the authors used a mouse model in which SGZ neurogenesis is disrupted7 because of elevated cell death (apoptosis) of newborn neurons2. P7C3 rescued SGZ neurogenesis in this model (as well as eliminating electrophysiological and structural defects present in the remaining neurons in the dentate gyrus) by inhibiting apoptosis, which normally prunes many newborn neurons in both wild-type and mutant animals. Apoptosis is generally regulated in part by mitochondrial integrity; P7C3 promotes this integrity in the face of normally proapoptotic calcium flux (Fig. 1c). The molecular targets of P7C3 are unknown but could include calcium-responsive proteins such as Bcl-2 or Apaf-1 or inhibition of mitochondrial caspases. Several chemically distinct compounds having known antiapoptotic activity were

also found to be proneurogenic; notably, one of these compounds (Dimebon) is being evaluated in clinical trials for use in neurodegenerative diseases. Consistently, P7C3 partially reversed age-related cognitive decline in a rat model of aging. Aged rats undergo marked loss of cognitive performance because of decreased neurogenesis, which is associated with increased apoptosis of newborn neurons. Treatment of 18-month-old rats (roughly equivalent in age to a 45-year-old human) with P7C3 for two months reduced the number of apoptotic newborn neurons. This was directly correlated with enhanced performance in a learning and memory assay. Whether factors other than increased survival of neuroblasts play a role in P7C3’s effects remains to be determined.

This paper provides several important advances, including a powerful new screening approach and the biology revealed through the screen. First, it provides benchmarks and methodologies for future in vivo small-molecule screening in mammals, comparable to previous whole-organism screening in invertebrates and nonmammalian vertebrates such as Caenorhabditis elegans8 and zebrafish9. In this screen, hits are biased toward favorable pharmacologic profiles, and the screen allows for easy elimination of compounds with unfavorable toxicity. Moreover, this in vivo setting allows for the manipulation of stem cells in their natural milieu, ruling out possible false positives

that can arise from abnormal behaviors and responses of stem cells cultured ex vivo or as stable cell lines. Second, P7C3 has favorable pharmacokinetics, low toxicity and no obvious teratogenicity. Thus it is an excellent lead compound for future development. Dimebon is currently in phase 3 clinical trials in Alzheimer’s and Huntington’s diseases because of reported memory-enhancing traits. Unfortunately, trials have produced equivocal results10. As P7C3 is ~10–100-fold more potent at promoting survival of newborn neurons and maintaining mitochondrial integrity than Dimebon, P7C3 derivatives could be highly efficacious in the setting of compromised NSC function and neurodegenerative disease. ■

Jonathan P. Saxe is in the Yale Stem Cell Center and Department of Cell Biology, New Haven, Connecticut, USA. e-mail: jonathan.saxe@yale.edu

references1. Ding, S. & Schultz, P.G. Nat. Biotechnol. 22, 833–840 (2004).2. Pieper, A.A. et al. Cell 142, 39–51 (2010).3. Zhao, C., Deng, W. & Gage, F.H. Cell 132, 645–660 (2008).4. Deng, W., Aimone, J.B. & Gage, F.H. Nat. Rev. Neurosci.

11, 339–350 (2010).5. Ding, S. et al. Proc. Natl. Acad. Sci. USA 100, 7632–7637

(2003).6. Saxe, J.P. et al. Chem. Biol. 14, 1019–1030 (2007).7. Pieper, A.A. et al. Proc. Natl. Acad. Sci. USA 102, 14052–14057

(2005).8. Min, J. et al. Nat. Chem. Biol. 3, 55–59 (2007).9. North, T.E. et al. Nature 447, 1007–1011 (2007).10. Miller, G. Science 327, 1309 (2010).

competing financial interestsThe author declares no competing financial interests.

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