Search

Disease-causing mutations in many neurodegenerative disorders lead to proteinopathies that trigger endoplasmic reticulum (ER) stress. However, few therapeutic options exist for patients with these diseases. Using an in vitro screening platform to identify compounds that protect human motor neurons from ER stress-mediated degeneration, we discovered that compounds targeting the mitogen-activated protein kinase kinase kinase kinase (MAP4K) family are neuroprotective. The kinase inhibitor URMC-099 (compound 1) stood out as a promising lead compound for further optimization. We coupled structure-based compound design with functional activity testing in neurons subjected to ER stress to develop a series of analogs with improved MAP4K inhibition and concomitant increases in potency and efficacy. Further structural modifications were performed to enhance the pharmacokinetic profiles of the compound 1 derivatives. Prostetin/12k emerged as an exceptionally potent, metabolically stable, and blood-brain barrier-penetrant compound that is well suited for future testing in animal models of neurodegeneration. • MAP4K inhibitors protect motor neurons from ER stress-induced apoptosis • We synthesized URMC-099 analogs to optimize selectivity and pharmacokinetics • Prostetin/12k is an analog with enhanced MAP4K inhibition and neuroprotection • Prostetin/12k shows improved stability, CNS penetrance, and oral bioavailability Using an in vitro screening platform to identify compounds that protect neurons from ER stress, a common feature of many neurodegenerative disorders, Bos et al. identified MAP4Ks as important shared targets of neuroprotective compounds. They then developed a series of potent, stable, and brain-penetrant MAP4K inhibitors, including Prostetin/12k. [ABSTRACT FROM AUTHOR]

Progressive degeneration of motor neurons (MNs) is the hallmark of amyotrophic lateral sclerosis (ALS). Limb-innervating lateral motor column MNs (LMC-MNs) seem to be particularly vulnerable and are among the first MNs affected in ALS. Here, we report association of this differential susceptibility with reduced expression of the mir-17∼92 cluster in LMC-MNs prior to disease onset. Reduced mir-17∼92 is accompanied by elevated nuclear PTEN in spinal MNs of presymptomatic SOD1 G93A mice. Selective dysregulation of the mir-17∼92/nuclear PTEN axis in degenerating SOD1 G93A LMC-MNs was confirmed in a double-transgenic embryonic stem cell system and recapitulated in human SOD1 +/L144F -induced pluripotent stem cell (iPSC)-derived MNs. We further show that overexpression of mir-17∼92 significantly rescues human SOD1 +/L144F MNs, and intrathecal delivery of adeno-associated virus (AAV)9-mir-17∼92 improves motor deficits and survival in SOD1 G93A mice. Thus, mir-17∼92 may have value as a prognostic marker of MN degeneration and is a candidate therapeutic target in SOD1-linked ALS. VIDEO ABSTRACT., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease selectively targeting motor neurons in the brain and spinal cord. The reasons for differential motor neuron susceptibility remain elusive. We developed a stem cell-based motor neuron assay to study cell-autonomous mechanisms causing motor neuron degeneration, with implications for ALS. A small-molecule screen identified cyclopiazonic acid (CPA) as a stressor to which stem cell-derived motor neurons were more sensitive than interneurons. CPA induced endoplasmic reticulum stress and the unfolded protein response. Furthermore, CPA resulted in an accelerated degeneration of motor neurons expressing human superoxide dismutase 1 (hSOD1) carrying the ALS-causing G93A mutation, compared to motor neurons expressing wild-type hSOD1. A secondary screen identified compounds that alleviated CPA-mediated motor neuron degeneration: three kinase inhibitors and tauroursodeoxycholic acid (TUDCA), a bile acid derivative. The neuroprotective effects of these compounds were validated in human stem cell-derived motor neurons carrying a mutated SOD1 allele (hSOD1 A4V ). Moreover, we found that the administration of TUDCA in an hSOD1 G93A mouse model of ALS reduced muscle denervation. Jointly, these results provide insights into the mechanisms contributing to the preferential susceptibility of ALS motor neurons, and they demonstrate the utility of stem cell-derived motor neurons for the discovery of new neuroprotective compounds., (Copyright © 2018 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

The generation of human pluripotent stem cell (hPSC)-derived ventricular progenitors and their assembly into a 3-dimensional in vivo functional ventricular heart patch has remained an elusive goal. Herein, we report the generation of an enriched pool of hPSC-derived ventricular progenitors (HVPs), which can expand, differentiate, self-assemble, and mature into a functional ventricular patch in vivo without the aid of any gel or matrix. We documented a specific temporal window, in which the HVPs will engraft in vivo. On day 6 of differentiation, HVPs were enriched by depleting cells positive for pluripotency marker TRA-1-60 with magnetic-activated cell sorting (MACS), and 3 million sorted cells were sub-capsularly transplanted onto kidneys of NSG mice where, after 2 months, they formed a 7 mm × 3 mm × 4 mm myocardial patch resembling the ventricular wall. The graft acquired several features of maturation: expression of ventricular marker (MLC2v), desmosomes, appearance of T-tubule-like structures, and electrophysiological action potential signature consistent with maturation, all this in a non-cardiac environment. We further demonstrated that HVPs transplanted into un-injured hearts of NSG mice remain viable for up to 8 months. Moreover, transplantation of 2 million HVPs largely preserved myocardial contractile function following myocardial infarction. Taken together, our study reaffirms the promising idea of using progenitor cells for regenerative therapy., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

Motor neurons (MNs) are unique because they project their axons outside of the CNS to innervate the peripheral muscles. Limb-innervating lateral motor column MNs (LMC-MNs) travel substantially to innervate distal limb mesenchyme. How LMC-MNs fine-tune the balance between survival and apoptosis while wiring the sensorimotor circuit en route remains unclear. Here, we show that the mir-17∼92 cluster is enriched in embryonic stem cell (ESC)-derived LMC-MNs and that conditional mir-17∼92 deletion in MNs results in the death of LMC-MNs in vitro and in vivo. mir-17∼92 overexpression rescues MNs from apoptosis, which occurs spontaneously during embryonic development. PTEN is a primary target of mir-17∼92 responsible for LMC-MN degeneration. Additionally, mir-17∼92 directly targets components of E3 ubiquitin ligases, affecting PTEN subcellular localization through monoubiquitination. This miRNA-mediated regulation modulates both target expression and target subcellular localization, providing LMC-MNs with an intricate defensive mechanism that controls their survival.

Although many distinct mutations in a variety of genes are known to cause Amyotrophic Lateral Sclerosis (ALS), it remains poorly understood how they selectively impact motor neuron biology and whether they converge on common pathways to cause neuronal degeneration. Here, we have combined reprogramming and stem cell differentiation approaches with genome engineering and RNA sequencing to define the transcriptional and functional changes that are induced in human motor neurons by mutant SOD1. Mutant SOD1 protein induced a transcriptional signature indicative of increased oxidative stress, reduced mitochondrial function, altered subcellular transport, and activation of the ER stress and unfolded protein response pathways. Functional studies demonstrated that these pathways were perturbed in a manner dependent on the SOD1 mutation. Finally, interrogation of stem-cell-derived motor neurons produced from ALS patients harboring a repeat expansion in C9orf72 indicates that at least a subset of these changes are more broadly conserved in ALS., (Copyright © 2014 Elsevier Inc. All rights reserved.)