Novel potent liposome agonists of triggering receptor expressed on myeloid cells 2 phenocopy antibody treatment in cells.

The receptor Triggering Receptor Expressed on Myeloid cells 2 (TREM2) is associated with several neurodegenerative diseases including Alzheimer's Disease and TREM2 stimulation represents a novel therapeutic opportunity. TREM2 can be activated by antibodies targeting the stalk region, most likely through receptor dimerization. Endogenous ligands of TREM2 are suggested to be negatively charged apoptotic bodies, mimicked by phosphatidylserine incorporated in liposomes and other polyanionic molecules likely binding to TREM2 IgV fold. However, there has been much discrepancy in the literature on the nature of phospholipids (PLs) that can activate TREM2 and on the stability of the corresponding liposomes over time. We describe optimized liposomes as robust agonists selective for TREM2 over TREM1 in cellular system. The detailed structure/activity relationship studies of lipid polar heads indicate that negatively charged lipid heads are required for activity and we identified the shortest maximally active PL sidechain. Optimized liposomes are active on both TREM2 common variant and TREM2 R47H mutant. Activity and selectivity were further confirmed in different native TREM2 expressing cell types including on integrated cellular responses such as stimulation of phagocytic activity. Such tool agonists will be useful in further studies of TREM2 biology in cellular systems alongside antibodies, and in the design of small molecule synthetic TREM2 agonists.

Elongator subunit 3 (ELP3) modifies ALS through tRNA modification.

Amyotrophic lateral sclerosis (ALS) is a fatal degenerative motor neuron disorder of which the progression is influenced by several disease-modifying factors. Here, we investigated ELP3, a subunit of the elongator complex that modifies tRNA wobble uridines, as one of such ALS disease modifiers. ELP3 attenuated the axonopathy of a mutant SOD1, as well as of a mutant C9orf72 ALS zebrafish model. Furthermore, the expression of ELP3 in the SOD1G93A mouse extended the survival and attenuated the denervation in this model. Depletion of ELP3 in vitro reduced the modified tRNA wobble uridine mcm5s2U and increased abundance of insoluble mutant SOD1, which was reverted by exogenous ELP3 expression. Interestingly, the expression of ELP3 in the motor cortex of ALS patients was reduced and correlated with mcm5s2U levels. Our results demonstrate that ELP3 is a modifier of ALS and suggest a link between tRNA modification and neurodegeneration.

Astrocyte-derived Jagged-1 mitigates deleterious Notch signaling in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a late-onset devastating degenerative disease mainly affecting motor neurons. Motor neuron degeneration is accompanied and aggravated by oligodendroglial pathology and the presence of reactive astrocytes and microglia. We studied the role of the Notch signaling pathway in ALS, as it is implicated in several processes that may contribute to this disease, including axonal retraction, microgliosis, astrocytosis, oligodendrocyte precursor cell proliferation and differentiation, and cell death. We observed abnormal activation of the Notch signaling pathway in the spinal cord of SOD1G93A mice, a well-established model for ALS, as well as in the spinal cord of patients with sporadic ALS (sALS). This increased activation was particularly evident in reactive GFAP-positive astrocytes. In addition, one of the main Notch ligands, Jagged-1, was ectopically expressed in reactive astrocytes in spinal cord from ALS mice and patients, but absent in resting astrocytes. Astrocyte-specific inactivation of Jagged-1 in presymptomatic SOD1G93A mice further exacerbated the activation of the Notch signaling pathway and aggravated the course of the disease in these animals without affecting disease onset. These data suggest that aberrant Notch signaling activation contributes to the pathogenesis of ALS, both in sALS patients and SOD1G93A mice, and that it is mitigated in part by the upregulation of astrocytic Jagged-1.

Oligodendrocyte dysfunction in the pathogenesis of amyotrophic lateral sclerosis.

Oligodendrocytes are well known targets for immune-mediated and infectious diseases, and have been suggested to play a role in neurodegeneration. Here, we report the involvement of oligodendrocytes and their progenitor cells in the ventral grey matter of the spinal cord in amyotrophic lateral sclerosis, a neurodegenerative disease of motor neurons. Degenerative changes in oligodendrocytes were abundantly present in human patients with amyotrophic lateral sclerosis and in an amyotrophic lateral sclerosis mouse model. In the mouse model, morphological changes in grey matter oligodendrocytes became apparent before disease onset, increasingly so during disease progression, and oligodendrocytes ultimately died. This loss was compensated by increased proliferation and differentiation of oligodendrocyte precursor cells. However, these newly differentiated oligodendrocytes were dysfunctional as suggested by their reduced myelin basic protein and monocarboxylate transporter 1 expression. Mutant superoxide dismutase 1 was found to directly affect monocarboxylate transporter 1 protein expression. Our data suggest that oligodendroglial dysfunction may be a contributor to motor neuron degeneration in amyotrophic lateral sclerosis.

Regulation of human microglial gene expression and function via RNAase-H active antisense oligonucleotides in vivo in Alzheimer's disease.

BACKGROUND: Microglia play important roles in maintaining brain homeostasis and neurodegeneration. The discovery of genetic variants in genes predominately or exclusively expressed in myeloid cells, such as Apolipoprotein E (APOE) and triggering receptor expressed on myeloid cells 2 (TREM2), as the strongest risk factors for Alzheimer's disease (AD) highlights the importance of microglial biology in the brain. The sequence, structure and function of several microglial proteins are poorly conserved across species, which has hampered the development of strategies aiming to modulate the expression of specific microglial genes. One way to target APOE and TREM2 is to modulate their expression using antisense oligonucleotides (ASOs). METHODS: In this study, we identified, produced, and tested novel, selective and potent ASOs for human APOE and TREM2. We used a combination of in vitro iPSC-microglia models, as well as microglial xenotransplanted mice to provide proof of activity in human microglial in vivo. RESULTS: We proved their efficacy in human iPSC microglia in vitro, as well as their pharmacological activity in vivo in a xenografted microglia model. We demonstrate ASOs targeting human microglia can modify their transcriptional profile and their response to amyloid-ß plaques in vivo in a model of AD. CONCLUSIONS: This study is the first proof-of-concept that human microglial can be modulated using ASOs in a dose-dependent manner to manipulate microglia phenotypes and response to neurodegeneration in vivo.

Effects of microglial depletion and TREM2 deficiency on Aß plaque burden and neuritic plaque tau pathology in 5XFAD mice.

Dystrophic neuronal processes harboring neuritic plaque (NP) tau pathology are found in association with Aß plaques in Alzheimer's disease (AD) brain. Microglia are also in proximity to these plaques and microglial gene variants are known risk factors in AD, including loss-of-function variants of TREM2. We have further investigated the role of Aß plaque-associated microglia in 5XFAD mice in which NP tau pathology forms after intracerebral injection of AD brain-derived pathologic tau (AD-tau), focusing on the consequences of reduced TREM2 expression and microglial depletion after treatment with the colony-stimulating factor 1 (CSFR1) inhibitor, PLX3397. Young 5XFAD mice treated with PLX3397 had a large reduction of brain microglia, including cortical plaque-associated microglia, with a significant reduction of Aß plaque burden in the cortex. A corresponding decrease in cortical APP-positive dystrophic processes and NP tau pathology were observed after intracerebral AD-tau injection in the PLX3397-treated 5XFAD mice. Consistent with prior reports, 5XFAD × TREM2-/- mice showed a significant reduction of plaque-associated microglial, whereas 5XFAD × TREM2+/- mice had significantly more plaque-associated microglia than 5XFAD × TREM2-/- mice. Nonetheless, AD-tau injected 5XFAD × TREM2+/- mice showed greatly increased AT8-positive NP tau relative to 5XFAD × TREM2+/+ mice. Expression profiling revealed that 5XFAD × TREM2+/- mice had a disease-associated microglial (DAM) gene expression profile in the brain that was generally intermediate between 5XFAD × TREM2+/+ and 5XFAD × TREM2-/- mice. Microarray analysis revealed significant differences in cortical and hippocampal gene expression between AD-tau injected 5XFAD × TREM2+/- and 5XFAD × TREM2-/- mice, including pathways linked to microglial function. These data suggest there is not a simple correlation between the extent of microglia plaque interaction and plaque-associated neuritic damage. Moreover, the differences in gene expression and microglial phenotype between TREM2+/- and TREM2-/- mice suggest that the former may better model the single copy TREM2 variants associated with AD risk.

Conditional deletion of Id2 or Notch1 in oligodendrocyte progenitor cells does not ameliorate disease outcome in SOD1G93A mice.

Oligodendrocytes are essential for structural and trophic support of motor axons. Their impairment has been implicated in amyotrophic lateral sclerosis (ALS), a neurodegenerative disorder of motor neurons. Oligodendrocyte progenitor cells fail to differentiate into mature oligodendrocytes and thereby jeopardize the health of motor neurons. Here, we report that oligodendrocytic ablation of inhibitor of DNA binding 2 (Id2) or Notch receptor 1 (Notch1), 2 negative master modulators of oligodendrocyte differentiation, fails to alleviate oligodendrocyte dysfunction or alter disease outcome in a murine model of ALS. Our data suggest that these inhibitors are not suitable targets for intervention in ALS.

The role of oligodendroglial dysfunction in amyotrophic lateral sclerosis.

Healthy oligodendrocytes are crucial for neurons and abnormal functioning of these cells is involved in several neurodegenerative diseases. We will focus on oligodendroglial pathology in amyotrophic lateral sclerosis (ALS), an adult-onset progressive neurodegenerative disease characterized by selective motor neuron loss. Recent discoveries shed new light on the crucial role of oligodendrocytes in this fatal disease. We will first give an overview of the importance of good-functioning oligodendrocytes for neuronal health, in particular for motor neurons. Subsequently, we will discuss the recent data on oligodendroglial abnormalities in ALS. We conclude that oligodendrocytes should be considered as important contributors to motor neuron degeneration. As a consequence, oligodendrocytes are a promising new therapeutic target for ALS and other neurodegenerative diseases.