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Abstract Background The neuronal ceroid lipofuscinoses (CLN diseases) are fatal lysosomal storage diseases causing neurodegeneration in the CNS. We have previously shown that neuroinflammation comprising innate and adaptive immune reactions drives axonal damage and neuron loss in the CNS of palmitoyl protein thioesterase 1-deficient (Ppt1 −/−) mice, a model of the infantile form of the diseases (CLN1). Therefore, we here explore whether pharmacological targeting of innate immune cells modifies disease outcome in CLN1 mice. Methods We applied treatment with PLX3397 (150 ppm in the chow), a potent inhibitor of the colony stimulating factor-1 receptor (CSF-1R) to target innate immune cells in CLN1 mice. Experimental long-term treatment was non-invasively monitored by longitudinal optical coherence tomography and rotarod analysis, as well as analysis of visual acuity, myoclonic jerks, and survival. Treatment effects regarding neuroinflammation, neural damage, and neurodegeneration were subsequently analyzed by histology and immunohistochemistry. Results We show that PLX3397 treatment attenuates neuroinflammation in CLN1 mice by depleting pro-inflammatory microglia/macrophages. This leads to a reduction of T lymphocyte recruitment, an amelioration of axon damage and neuron loss in the retinotectal system, as well as reduced thinning of the inner retina and total brain atrophy. Accordingly, long-term treatment with the inhibitor also ameliorates clinical outcomes in CLN1 mice, such as impaired motor coordination, visual acuity, and myoclonic jerks. However, we detected a sex- and region-biased efficacy of CSF-1R inhibition, with male microglia/macrophages showing higher responsiveness toward depletion, especially in the gray matter of the CNS. This results in a better treatment outcome in male Ppt1 −/− mice regarding some histopathological and clinical readouts and reflects heterogeneity of innate immune reactions in the diseased CNS. Conclusions Our results demonstrate a detrimental impact of innate immune reactions in the CNS of CLN1 mice. These findings provide insights into CLN pathogenesis and may guide in the design of immunomodulatory treatment strategies.

Abstract Background Cosmic radiation exposures have been found to elicit cognitive impairments involving a wide-range of underlying neuropathology including elevated oxidative stress, neural stem cell loss, and compromised neuronal architecture. Cognitive impairments have also been associated with sustained microglia activation following low dose exposure to helium ions. Space-relevant charged particles elicit neuroinflammation that persists long-term post-irradiation. Here, we investigated the potential neurocognitive benefits of microglia depletion following low dose whole body exposure to helium ions. Methods Adult mice were administered a dietary inhibitor (PLX5622) of colony stimulating factor-1 receptor (CSF1R) to deplete microglia 2 weeks after whole body helium irradiation (4He, 30 cGy, 400 MeV/n). Cohorts of mice maintained on a normal and PLX5622 diet were tested for cognitive function using seven independent behavioral tasks, microglial activation, hippocampal neuronal morphology, spine density, and electrophysiology properties 4–6 weeks later. Results PLX5622 treatment caused a rapid and near complete elimination of microglia in the brain within 3 days of treatment. Irradiated animals on normal diet exhibited a range of behavioral deficits involving the medial pre-frontal cortex and hippocampus and increased microglial activation. Animals on PLX5622 diet exhibited no radiation-induced cognitive deficits, and expression of resting and activated microglia were almost completely abolished, without any effects on the oligodendrocyte progenitors, throughout the brain. While PLX5622 treatment was found to attenuate radiation-induced increases in post-synaptic density protein 95 (PSD-95) puncta and to preserve mushroom type spine densities, other morphologic features of neurons and electrophysiologic measures of intrinsic excitability were relatively unaffected. Conclusions Our data suggest that microglia play a critical role in cosmic radiation-induced cognitive deficits in mice and, that approaches targeting microglial function are poised to provide considerable benefit to the brain exposed to charged particles.

Abstract Numerous clinical studies have established the debilitating neurocognitive side effects of chemotherapy in the treatment of breast cancer, often referred as chemobrain. We hypothesize that cognitive impairments are associated with elevated microglial inflammation in the brain. Thus, either elimination of microglia or restoration of microglial function could ameliorate cognitive dysfunction. Using a rodent model of chronic Adriamycin (ADR) treatment, a commonly used breast cancer chemotherapy, we evaluated two strategies to ameliorate chemobrain: 1) microglia depletion using the colony stimulating factor-1 receptor (CSF1R) inhibitor PLX5622 and 2) human induced pluripotent stem cell-derived microglia (iMG)-derived extracellular vesicle (EV) treatment. In strategy 1 mice received ADR once weekly for 4 weeks and were then administered CSF1R inhibitor (PLX5622) starting 72 h post-ADR treatment. ADR-treated animals given a normal diet exhibited significant behavioral deficits and increased microglial activation 4–6 weeks later. PLX5622-treated mice exhibited no ADR-related cognitive deficits and near complete depletion of IBA-1 and CD68+ microglia in the brain. Cytokine and RNA sequencing analysis for inflammation pathways validated these findings. In strategy 2, 1 week after the last ADR treatment, mice received retro-orbital vein injections of iMG-EV (once weekly for 4 weeks) and 1 week later, mice underwent behavior testing. ADR-treated mice receiving EV showed nearly complete restoration of cognitive function and significant reductions in microglial activation as compared to untreated ADR mice. Our data demonstrate that ADR treatment elevates CNS inflammation that is linked to cognitive dysfunction and that attenuation of neuroinflammation reverses the adverse neurocognitive effects of chemotherapy.

Genetics implicate microglia in Alzheimer’s disease pathogenesis, but their roles remain unclear. Here, the authors find that microglial depletion in a mouse model of Alzheimer’s disease impairs plaque formation and that Aβ-induced changes in neuronal gene expression are microglia-mediated.

Kynurenine 3-monooxygenase (KMO), a key player in the kynurenine pathway (KP) of tryptophan degradation, regulates the synthesis of the neuroactive metabolites 3-hydroxykynurenine (3-HK) and kynurenic acid (KYNA). KMO activity has been implicated in several major brain diseases including Huntington’s disease (HD) and schizophrenia. In the brain, KMO is widely believed to be predominantly localized in microglial cells, but verification in vivo has not been provided so far. Here, we examined KP metabolism in the brain after depleting microglial cells pharmacologically with the colony stimulating factor 1 receptor inhibitor PLX5622. Young adult mice were fed PLX5622 for 21 days and were euthanized either on the next day or after receiving normal chow for an additional 21 days. Expression of microglial marker genes was dramatically reduced on day 22 but had fully recovered by day 43. In both groups, PLX5622 treatment failed to affect Kmo expression, KMO activity or tissue levels of 3-HK and KYNA in the brain. In a parallel experiment, PLX5622 treatment also did not reduce KMO activity, 3-HK and KYNA in the brain of R6/2 mice (a model of HD with activated microglia). Finally, using freshly isolated mouse cells ex vivo, we found KMO only in microglia and neurons but not in astrocytes. Taken together, these data unexpectedly revealed that neurons contain a large proportion of functional KMO in the adult mouse brain under both physiological and pathological conditions.

Abstract Background The 18-kDa translocator protein (TSPO) is an important target for assessing neuroimmune function in brain with positron-emission tomography (PET) imaging. The goal of this work was to assess two [11C]PBR28 imaging paradigms for measuring dynamic microglia changes in Macaca mulatta. Methods Dynamic [11C]PBR28 PET imaging data with arterial blood sampling were acquired to quantify TSPO levels as [11C]PBR28 V T. Scans were acquired at three timepoints: baseline, immediately post-drug, and prolonged post-drug. Results In one animal, a colony-stimulating factor 1 receptor kinase inhibitor, previously shown to deplete brain microglia, reduced [11C]PBR28 V T in brain by 46 ± 3% from baseline, which recovered after 12 days to 7 ± 5% from baseline. In a different animal, acute lipopolysaccharide administration, shown to activate brain microglia, increased [11C]PBR28 V T in brain by 39 ± 9% from baseline, which recovered after 14 days to −11 ± 3% from baseline. Conclusions These studies provide preliminary evidence of complementary paradigms to assess microglia dynamics via in vivo TSPO imaging.

Purpose: To evaluate the safety, recommended phase II dose (RP2D) and efficacy of pexidartinib, a colony stimulating factor receptor 1 (CSF-1R) inhibitor, in combination with weekly paclitaxel in patients with advanced solid tumors. Patients and Methods: In part 1 of this phase Ib study, 24 patients with advanced solid tumors received escalating doses of pexidartinib with weekly paclitaxel (80 mg/m 2 ). Pexidartinib was administered at 600 mg/day in cohort 1. For subsequent cohorts, the dose was increased by ⩽50% using a standard 3+3 design. In part 2, 30 patients with metastatic solid tumors were enrolled to examine safety, tolerability and efficacy of the RP2D. Pharmacokinetics and biomarkers were also assessed. Results: A total of 51 patients reported ≥1 adverse event(s) (AEs) that were at least possibly related to either study drug. Grade 3–4 AEs, including anemia (26%), neutropenia (22%), lymphopenia (19%), fatigue (15%), and hypertension (11%), were recorded in 38 patients (70%). In part 1, no maximum tolerated dose was achieved and 1600 mg/day was determined to be the RP2D. Of 38 patients evaluable for efficacy, 1 (3%) had complete response, 5 (13%) partial response, 13 (34%) stable disease, and 17 (45%) progressive disease. No drug–drug interactions were found. Plasma CSF-1 levels increased 1.6- to 53-fold, and CD14dim/CD16+ monocyte levels decreased by 57–100%. Conclusions: The combination of pexidartinib and paclitaxel was generally well tolerated. RP2D for pexidartinib was 1600 mg/day. Pexidartinib blocked CSF-1R signaling, indicating potential for mitigating macrophage tumor infiltration.

How microglia contribute to brain injury or repair is unclear. Here combining microglia manipulations and calcium imaging, the authors show that selective elimination of microglia leads to disrupted neuronal calcium dynamics and markedly increased brain injury after cerebral ischemia.

Microglia are the primary immune cell in the brain and are postulated to play important roles outside of immunity. Administration of the dual colony-stimulating factor 1 receptor (CSF1R)/c-Kit kinase inhibitor, PLX3397, to adult mice results in the elimination of ~99% of microglia, which remain eliminated for as long as treatment continues. Upon removal of the inhibitor, microglia rapidly repopulate the entire adult brain, stemming from a central nervous system (CNS) resident progenitor cell. Using this method of microglial elimination and repopulation, the role of microglia in both healthy and diseased states can be explored. Here, we examine the responsiveness of newly repopulated microglia to an inflammatory stimulus, as well as determine the impact of these cells on behavior, cognition, and neuroinflammation. Two month-old wild-type mice were placed on either control or PLX3397 diet for 21 d to eliminate microglia. PLX3397 diet was then removed in a subset of animals to allow microglia to repopulate and behavioral testing conducted beginning at 14 d repopulation. Finally, inflammatory profiling of the microglia-repopulated brain in response to lipopolysaccharide (LPS; 0.25 mg/kg) or phosphate buffered saline (PBS) was determined 21 d after inhibitor removal using quantitative real time polymerase chain reaction (RT-PCR), as well as detailed analyses of microglial morphologies. We find mice with repopulated microglia to perform similarly to controls by measures of behavior, cognition, and motor function. Compared to control/resident microglia, repopulated microglia had larger cell bodies and less complex branching in their processes, which resolved over time after inhibitor removal. Inflammatory profiling revealed that the mRNA gene expression of repopulated microglia was similar to normal resident microglia and that these new cells appear functional and responsive to LPS. Overall, these data demonstrate that newly repopulated microglia function similarly to the original resident microglia without any apparent adverse effects in healthy adult mice.

Tumor associated macrophages (TAM) can promote angiogenesis, invasiveness and immunosuppression. The cytokine CSF-1 (or M-CSF) is an important factor of TAM recruitment and differentiation and several pharmacological agents targeting the CSF-1 receptor (CSF-1R) have been developed to regulate TAM in solid cancers. We show that the kinase inhibitor PLX3397 strongly dampened the systemic and local accumulation of macrophages driven by B16F10 melanomas, without affecting Gr-1(+) myeloid derived suppressor cells. Removal of intratumoral macrophages was remarkably efficient and a modest, but statistically significant, delay in melanoma outgrowth was observed. Importantly, CSF-1R inhibition strongly enhanced tumor control by immunotherapy using tumor-specific CD8 T cells. Elevated IFNγ production by T cells was observed in mice treated with the combination of PLX3397 and immunotherapy. These results support the combined use of CSF-1R inhibition with CD8 T cell immunotherapy, especially for macrophage-stimulating tumors.