Journal: Advanced drug delivery reviews

Article Title: Progress in tumor-associated macrophage (TAM)-targeted therapeutics

doi: 10.1016/j.addr.2017.04.010

Figure Lengend Snippet: Selected compilation of drugs in clinical trials whose mechanisms of action may involve modulation of macrophages/TAMs

Article Snippet: Pexidartinib/PLX3397 (Oral c-fms/KIT/FLT3 tyrosine kinase inhibitor) , Centre Leon Berard and Plexxikon, AstraZeneca , 1 Ongoing , Advanced/metastatic colorectal or pancreatic cancer , With Durvalumab (Anti-PD-L1 antibody) , NCT02777710 , May 12, 2016.

Techniques: Clinical Proteomics, Mutagenesis, Inhibition, Variant Assay

Journal: The Journal of Cell Biology

Article Title: Autophagy gene FIP200 in neural progenitors non–cell autonomously controls differentiation by regulating microglia

doi: 10.1083/jcb.201609093

Figure Lengend Snippet: Suppression of microglia activation and infiltration rescued the defective neurogenesis in 2cKO mice. (A) The percentage of ramified, round, and amoeboid microglia per SVZ section of Ctrl, FIP cKO, 2cKO, and p53 cKO mice treated with vehicle (Veh) or minocycline (Mino; three mice each). (B–D) Number of iNOS + microglia (B), IL-6 + microglia (C), or TNF + microglia (D) per SVZ section in Ctrl, FIP cKO, 2cKO, and p53 cKO mice treated with vehicle (Veh) or minocycline (Mino; mean ± SEM; three mice each). (E–G) Number of DCX + neuroblasts (E) and GFAP + astrocytes (F) per SVZ section or the density of NeuN + neurons per olfactory bulb section (G) in Ctrl, FIP cKO, 2cKO, and p53 cKO mice treated with vehicle or minocycline are shown (mean ± SEM; three mice each). (H) Number of Iba1 + microglia per SVZ section of 2cKO mice treated with vehicle (Veh) or TAK-779 (TAK; mean ± SEM; three mice each). (I–K) Number of DCX + neuroblasts (I) and GFAP + astrocytes (K) per SVZ section or the density of NeuN + neurons per olfactory bulb section (J) in 2cKO mice treated with vehicle (Veh) or TAK-779 (TAK; mean ± SEM; three mice each). (L) Number of Iba1 + microglia per SVZ section in Ctrl, FIP cKO, 2cKO, and p53 cKO mice treated with vehicle (Veh) or PLX3397 (PLX; mean ± SEM; four mice each). (M–O) Number of DCX + neuroblasts (M) and GFAP + astrocytes (O) per SVZ section or the density of NeuN + neurons per olfactory bulb section (N) in Ctrl, FIP cKO, 2cKO, and p53 cKO mice treated with vehicle (Veh) or PLX3397 (PLX) are shown (mean ± SEM; four mice each). **, P < 0.01; ***, P < 0.001.

Article Snippet: PLX3397 (ApexBio) was dissolved in DMSO and then added immediately before using to a solution of 0.5% methyl cellulose (M0512; Sigma-Aldrich) and 1.0% Tween 80 (P1754; Sigma-Aldrich).

Techniques: Activation Assay

Journal: Cell reports

Article Title: Developmental Apoptosis Promotes a Disease-Related Gene Signature and Independence from CSF1R Signaling in Retinal Microglia

doi: 10.1016/j.celrep.2019.04.062

Figure Lengend Snippet: (A) Tamoxifen dosing regimen. (B) Confocal images of retinal wholemounts of control at P9 (tamoxifen at P4, P6, and P8) (left) and homozygous floxed at P8 (tamoxifen at P4 and P6) (right). Scale bars, 100 µm. Experiment repeated twice, n = 4 animals each. (C) PLX3397 (PLX) dosing regimen. (D) Confocal images of retinal wholemounts at P7 from vehicle- (left) and PLX-treated (right) animals. Scale bar, 100 µm. (E) Representative flow cytometry plot and gate for collecting GFP + CD45 + microglia in vehicle- (left) and PLX-treated (right) animals. (F) Percent microglia (CD45 + GFP + ) of total single cells by flow cytometry. (±SEM; n = 8 each). Two-tailed unpaired t test t(14) = 4.438, ***p = 0.0006. (G) Percent CD11c HI of total CD45 + GFP + microglia by flow cytometry. (±SEM; n = 8 each) Two-tailed unpaired t test t(14) = 4.662, ***p = 0.0004. (H) Log-transformed relative gene expression by qPCR (±SEM) in sorted microglia from PLX-treated retinas relative to vehicle controls. WT (n≥8 except n = 5 Cd68, Lamp1, Lyz2, P2ry12); PLX (n = 9 except n = 8 for Itgax, n = 5 Cd68, Lamp1, Lyz2, P2ry12). Two-tailed unpaired t test *p ≤ 0.05, **p ɛ≤ 0.01, ***p ≤ 0.001. #, Welch’s correction. (I) HCR of vehicle- and PLX-treated retinas. Scale bars, 50µm. (J) Ratio (±SEM) of PLX-treated over genotyped-matched control density (CD45 + Cd11b + /singlets). WT (n = 17); Trem2 KO (n = 6); Bax KO (n = 5). One-way BrownForsythe ANOVA F(2,24.29) = 11.76, p = 0.0003 and Games-Howell’s multiple comparisons test. *p ≤ 0.05, ***p ≤ 0.001. (K) Percent (±SEM) of CD11c HI microglia in total CD45 + CD11b=population after PLX by flow cytometry. WT littermates (n = 5), Trem2 KO (n = 7); Bax KO (n = 5). One-way ANOVA F(2,14) = 24.55, p < 0.0001 and Tukey’s multiple comparisons test ***p = 0.0005, ****p < 0.0001. See also and .

Article Snippet: PLX3397 (AdooQ BioScience, A15520) was dissolved in corn oil and 10% DMSO and administered to postnatal pups by intraperitoneal injection on P4, P5, and P6 at 0.25mg/g body weight.

Techniques: Control, Flow Cytometry, Two Tailed Test, Transformation Assay, Gene Expression

Journal: Cell reports

Article Title: Developmental Apoptosis Promotes a Disease-Related Gene Signature and Independence from CSF1R Signaling in Retinal Microglia

doi: 10.1016/j.celrep.2019.04.062

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: PLX3397 (AdooQ BioScience, A15520) was dissolved in corn oil and 10% DMSO and administered to postnatal pups by intraperitoneal injection on P4, P5, and P6 at 0.25mg/g body weight.

Techniques: Recombinant, Red Blood Cell Lysis, Blocking Assay, In Situ Hybridization, cDNA Synthesis, Software

Journal: bioRxiv

Article Title: Retinal ganglion cell survival after severe optic nerve injury is modulated by crosstalk between JAK/STAT signaling and innate immune responses in the zebrafish retina

doi: 10.1101/2021.04.08.439090

Figure Lengend Snippet: Immunostaining of 4C4 (magenta) on isl2b :GFP retinal cryosections (A) and retinal flat mounts at 1dpi with Imaris surface renderings of 4C4 + macrophages/microglia (B). (C) Images of 1dpi retinal flat-mounts from isl2b :GFP; mpeg1 :mCherry animals; macrophages/microglia (magenta). (D) Quantification of the GCL surface area occupied by mCherry + macrophages/microglia at 1dpi (n=4/condition). Shown are mean±SD; ***p<0.001; Mann-Whitney test. (E) Violin plot showing a significant increase in sphericity of mCherry+ macrophages/microglia in ONT+ retinae compared to ONT- controls (n=140 in ONT- and n=272 in ONT+). ****p<0.0001; unpaired t-test with Welch’s correction. (F) Flat-mount images of isl2b :GFP; mpeg1 :mCherry retinae after intravitreal injection of dexamethasone (Dex) or DMSO (Ctr) at 7dpi. (G) RGC survival in dexamethasone-treated retinae increased significantly at 7dpi when compared to control (n=4/condition). Shown are mean±SD; *p<0.05; Kruskal Wallis ANOVA test with Dunn’s multiple comparisons. (H) Quantification of mCherry + macrophage/microglia coverage of the GCL after ONT and dexamethasone or DMSO injection (n=3/condition). Shown are mean±SD; Kruskal Wallis ANOVA test with Dunn’s multiple comparisons. No significant differences were detected. (I) Flat-mount images of isl2b :GFP; mpeg1 :mCherry retinae after PLX3397 or control treatment (Ctr) at 7dpi. (J) Quantification of mCherry + macrophage/microglia coverage of the GCL after ONT and PLX3397 treatment (n=4/condition). Shown are mean±SD; *p<0.05; Kruskal Wallis ANOVA test with Dunn’s multiple comparisons. (K) RGC survival in PLX3397-treated retinae increased significantly at 7dpi when compared to control. Similarly, RGC survival in PLX3397-treated retinae increased significantly after P6 addition over DMSO controls (n=4/condition). Shown are mean±SD; *P<0.05; Mann-Whitney test. Scale bars = 50μm.

Article Snippet: To deplete macrophages/microglia, fish were immersed in 500nM PLX3397 (Fisher Scientific) in system water, as previously described ( ).

Techniques: Immunostaining, MANN-WHITNEY, Injection

Journal: Expert opinion on emerging drugs

Article Title: An update: emerging drugs to treat squamous cell carcinomas of the head and neck

doi: 10.1080/14728214.2018.1543400

Figure Lengend Snippet: Emerging immunotherapeutic agents currently active, under investigation, or recruiting in head and neck squamous cell carcinoma

Article Snippet: Pexidartinib (PLX3397) , Plexxikon, Daiichi Sankyo, Merck , CSF1R , Inhibition of TAM by inhibiting binding of CSF-1 to CSF1R , I , Combined with pembrolizumab.

Techniques: Inhibition, Activation Assay, Binding Assay, Injection, Immunopeptidomics, Lysis, Expressing

Journal: Immunity

Article Title: Stellate Cells, Hepatocytes, and Endothelial Cells Imprint the Kupffer Cell Identity on Monocytes Colonizing the Liver Macrophage Niche

doi: 10.1016/j.immuni.2019.08.017

Figure Lengend Snippet: Replenishment of KC Pool by Ly6C hi Monocytes (A and B) Expression of GFP, Ly6C, and F4/80 of monocytes (green gate), em-KCs (black gate), and mo-KCs (red gate) after DT injection in (A) Clec4f DTR/+ - Ccr2 GFP/+ mice or (B) Clec4f DTR/+ - Ccr2 GFP/GFP mice. Flow-cytometry plots are pre-gated on live CD45 + CD11b + Lyve-1 − SiglecF − Ly6G − single cells. Data are representative of 2–3 experiments. (C) Proportion of Ly6C hi monocytes (green lines), em-KCs (black lines), and mo-KCs (red lines) in the liver of Clec4f DTR/+ - Ccr2 GFP/+ mice (solid lines) or Clec4f DTR/+ - Ccr2 GFP/GFP mice (dashed lines) as a percentage of live CD45 + cells after DT injection. Pooled data are from 2–3 experiments; n = 5 (0,5d), 6 (PBS, 1d; 1,5d; 2d; 5d; 6d), and 8 mice (3d; 4d). (D) Heatmap showing the top 30 of upregulated genes in mo-KCs 3 days compared with 1 and 7 days after DT injection. (E) Expression of Ki-67 and EdU incorporation by em-KCs and mo-KCs in (top) Clec4f DTR/+ - Ccr2 GFP/+ mice or (bottom) in Clec4f DTR/+ - Ccr2 GFP/GFP mice after DT injection. Flow-cytometry plots are pre-gated as in (A). Data are representative of 2–3 experiments. (F) Percentage of EdU + cells in indicated populations during the differentiation kinetic of mo-KCs in Clec4f DTR/+ - Ccr2 GFP/+ mice (solid line) or Clec4f DTR/+ - Ccr2 GFP/GFP (dash line). Pooled data are from 2–3 experiments; n = 5 (0,5d), 6 (PBS, 1d, 1,5d, 2d, 5d, 6d), and 8 mice (3d, 4d). Two-way ANOVA with Tukey post-test. ∗ p < 0.05, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. (G) Percentage of EdU + mo-KCs 3 days after DT injection in liver of mice treated with PLX3397 (α-CSF1R) or vehicle. Pooled data are from 2 experiments, n = 7. Mann-Whitney t test. ∗∗∗ p < 0.001. Related to Figure S1 .

Article Snippet: PLX3397 , Achemblock , Cat#H-8970.

Techniques: Expressing, Injection, Flow Cytometry, MANN-WHITNEY

Journal: Immunity

Article Title: Stellate Cells, Hepatocytes, and Endothelial Cells Imprint the Kupffer Cell Identity on Monocytes Colonizing the Liver Macrophage Niche

doi: 10.1016/j.immuni.2019.08.017

Figure Lengend Snippet:

Article Snippet: PLX3397 , Achemblock , Cat#H-8970.

Techniques: Control, Recombinant, Blocking Assay, Irradiation, Enzyme-linked Immunosorbent Assay, cDNA Synthesis, Microarray, Software, Microscopy

Journal: iScience

Article Title: Microglia depletion prevents lactation by inhibition of prolactin secretion

doi: 10.1016/j.isci.2023.106264

Figure Lengend Snippet: The effect of systemic PLX3394 treatment on the body weight of pups and prolactin signaling and behavior in mice whose treatment started during pregnancy (Experiment 1 - Exp. 1) (A) The timeline of the protocols. The mating of animals (n = 6 animals per group) was controlled for accuracy of the treatment. (B) Disappearance of Iba1-positive microglial cells in response to PLX3397-treatment. b1, Iba1-labeled coronal section of the mediobasal hypothalamus in control animals. Labeled cells are shown in the arcuate nucleus (Arc) and the ventromedial subdivision of the ventrolateral hypothalamic nucleus (VMHvl). b2, The same field of a coronal section as in b1 is shown in a PLX3397 treated animal demonstrating a reduced number of microglia cells. The scale bar is 300 μm. b3, The number of Iba1 labeled cells per 1 mm 2 are shown in the Arc. The column representing the density of Iba1 labeled cells is yellow for the control animals and green for the treated group (n = 6 mothers per group). Statistical analysis was performed using two-way ANOVA test with the type of treatment (PLX3397 versus control) being one parameter and the brain regions being the repeated parameter followed by Sidek’s multiple comparison test. (C) The effect of PLX3397 treatment on the body weight of pups (n = 18 pups) in Exp. 1. The statistical analysis of weight gain was performed with two-way ANOVA with the treatment being the independent parameter and the body weights on consecutive days the repeated parameter. C1, The change of the body weight of pups in Exp. 1 in percentage of body weight at postpartum day 3 (test day 2, treated day 23). A large difference was detected the following day when the measurement ended for the treated mothers to save the life of pups. C2, The apparent body weight change of pups during a 1-h suckling bout after 4 h of maternal separation in Exp. 1. D1, The percentage of the time mothers (n = 6 mothers per group) spent with parental behavior in Exp. 1 when treated mothers received oral PLX3397 treatment during the pregnancy. There was no difference between the two groups (control group - yellow column, treated group – green column) in nursing, and other parental or non-parental behaviors. D2, The latency of retrieval of the third pup into the nest. There was a significant difference between control and microglia depleted groups in Exp. 1 as the treatment increased the latency of pup retrieval (n = 6 mothers per group). The statistical analysis was performed with unpaired t-test using the Welch’s correction because the F test indicated significant differences between the variances of the control and treated groups. E1, The effect of PLX3397 treatment on the behavior of mother (n = 6 per group) in an arena (open field test).e1, There was no difference between the groups in terms of the part of the arena they stayed. E2, Duration of behavioral elements exhibited by the two groups of mice did not differ, either. (F) The results of the forced swim test (FST). The animals did not show any difference in the time they spent with swimming, climbing and floating. (G) The results of the elevated plus maze (EPM) test. There was no difference in the time spent in the arms between the treated and control groups. Data are represented as mean ± SEM.

Article Snippet: Control animals received the same food without PLX3397 (SM R/M-H Placebo; Ssniff).

Techniques: Labeling, Control, Comparison

Journal: iScience

Article Title: Microglia depletion prevents lactation by inhibition of prolactin secretion

doi: 10.1016/j.isci.2023.106264

Figure Lengend Snippet: The effect of systemic PLX3394 treatment on the body weight of pups and prolactin signaling and behavior in mice whose treatment started during pregnancy (Experiment 2) (A) The timeline of the protocols. The mating of animals (n = 6 animals per group) was controlled for accuracy of the treatment. (B) The effect of PLX3397 treatment on the body weight of pups (3 pups/mother) in Exp. 2 (n = 6 mothers per group). The statistical analysis of weight gain was performed with two-way ANOVA with the treatment being the independent parameter and the body weights on consecutive days the repeated parameter. b1, The change of the body weight of foster pups given to mothers at their 21st postpartum day in Exp. 2. The body weights of the foster pups are expressed in percentage of their body weight at the 23rd postpartum day (second test day, 23rd treated day) of the mothers. The body weight of the pups was reduced 3 days later for the treated mothers. b2, The effect of the treatment on the body weight gain of pups during a 1 h long suckling bout following 4 h of maternal separation performed at the 25th postpartum day (fourth test day, 25th treated day) of the mothers using foster pups. The apparent weight gain of pups during suckling was highly significantly lower in the treated than the control group. (C) The effect of PLX3397 treatment on the number of prolactin sensitive cells in the mediobasal hypothalamus (n = 6 mothers per group). C1, pSTAT5-positive cells (black dots) are present in the Arc and the VMHvl of a control animal. C2, A brain section showing the same field as in e1 demonstrates that pSTAT5-positive neurons are not visible in the Arc and VMHvl of a PLX3397-treated animal. The scale bar is 300 μm. C3, Quantification of the density of pSTAT5-positive cells in the mediobasal hypothalamus. The columns represent the number of pSTAT5-positive cells in the control animals (yellow) on the left and the treated group (green) on the right. Additional abbreviations: 3V – third ventricle, f – fornix. ∗: p < 0.05; ∗∗∗: p < 0.001. D1, The effect of postpartum oral PLX3397 treatment on the maternal behavior of dams. The columns show the percentage of time what mothers spent with nursing, other maternal behaviors, or non-parental behaviors. D2, The effect of postpartum oral PLX3397 treatment on behavior of mice dams in the pup retrieval test. E1, The effect of PLX3397 treatment on the behavior of mother (n = 6 per group) in an arena (open field test). There was no difference between the groups in terms of the part of the arena they stayed. E2, Duration of behavioral elements exhibited by the 2 groups of mice did not differ, either. (F) The results of the forced swim test (FST). The animals did not show any difference in the time they spent with swimming, climbing and floating. (G) The results of the elevated plus maze (EPM) test. There was no difference in the time spent in the arms between the treated and control groups. Data are represented as mean ± SEM.

Article Snippet: Control animals received the same food without PLX3397 (SM R/M-H Placebo; Ssniff).

Techniques: Control

Journal: iScience

Article Title: Microglia depletion prevents lactation by inhibition of prolactin secretion

doi: 10.1016/j.isci.2023.106264

Figure Lengend Snippet: The effect of intracerebroventricular PLX3394 treatment of postpartum mothers on the body weight of pups and prolactin secretion and behavior in rats (Experiment 3) (A) Illustration of the third experiment (Exp. 3), in which rat mothers (n = 10 treated and 9 control mothers) received PLX3397 treatment only in the postpartum period directly to the lateral ventricle via osmotic minipumps. (B) The effect of PLX3397 treatment on the weight of litters (10 pups per litter; n = 9 litters for control and 10 for treated mothers). The statistical analysis of weight gain was performed with two-way ANOVA with the treatment being the independent parameter and the body weights on consecutive days the repeated parameter. B1, The change of the body weight of foster pups given to mothers at their 23rd postpartum day in Exp. 3. The body weight of the foster pups is expressed in percentage of their body weight at the 25 th postpartum day (first test day, 23 rd treated day) of the mothers. The body weight of the pups was reduced 2 and 3 days later for the treated mothers. B2, The weight gain of pups during a 1-h long suckling bout. Although pups of treated mothers gained some weight during suckling, the weight gain was significantly higher in the control group (p < 0.001). (C) The effect of PLX3397 treatment on suckling-induced serum prolactin levels. The level of prolactin was equally low in the 2 groups after 4 h of pup separation. In turn, significant difference was found 30 and 60 min after the pups were given back to the mothers (p < 0.001, n = 10 treated and 9 control mothers) as the serum prolactin level was dramatically increased in the control group whereas only a small and late increase was detected in the treated mothers. The statistical analysis was carried out with two-way ANOVA followed by Sidak’s multiple comparison test with the type of treatment (PLX3397 versus control) being one parameter and time points of prolactin level measurements being the repeated parameter. ∗: p < 0.05; ∗∗: p < 0.01, ∗∗∗: p < 0.001. d1, pSTAT5-ir cells (black dots) are present in the Arc and VMHvl nuclei in a control rat. D2, The brain section represents the corresponding field as in d1 in a PLX3397-treated mother. It is demonstrated that the treatment causes a reduced number of pSTAT5-positive neurons. The scale bar is 300 μm. D3, Quantification of the number of pSTAT5-ir cells in the mediobasal hypothalamus. The columns represent the density of pSTAT5-ir cells in the control animals (yellow) on the left and the treated group (green) on the right. f – fornix. The statistical analysis to compare the density of cells in the mediobasal hypothalamus between control and PLX3397 treated groups was performed with unpaired t-test. ∗∗∗: p < 0.001. E1, The effect of postpartum oral PLX3397 treatment on the maternal behavior of dams. The columns show the percentage of time what mothers spent with nursing, other maternal behaviors, or non-parental behaviors. E2, The effect of postpartum oral PLX3397 treatment on behavior of mice dams in the pup retrieval test. (F) The results of the elevated plus maze (EPM) test. There was no difference in the time spent in the arms between the treated and control groups. (G) The results of the forced swim test (FST). The animals did not show any difference in the time they spent with swimming, climbing and floating. Data are represented as mean ± SEM.

Article Snippet: Control animals received the same food without PLX3397 (SM R/M-H Placebo; Ssniff).

Techniques: Control, Comparison

Journal: iScience

Article Title: Microglia depletion prevents lactation by inhibition of prolactin secretion

doi: 10.1016/j.isci.2023.106264

Figure Lengend Snippet: The effect of PLX3398 treatment of the pups on the body weight during a suckling bout Vehicle and PLX3397 treated pups gained similar amounts of weight during a 1-h long suckling bout. Data are represented as mean ± SEM.

Article Snippet: Control animals received the same food without PLX3397 (SM R/M-H Placebo; Ssniff).

Techniques:

Journal: iScience

Article Title: Microglia depletion prevents lactation by inhibition of prolactin secretion

doi: 10.1016/j.isci.2023.106264

Figure Lengend Snippet:

Article Snippet: Control animals received the same food without PLX3397 (SM R/M-H Placebo; Ssniff).

Techniques: Software, Recombinant, Plasmid Preparation

Journal: bioRxiv

Article Title: Aggregation of HAPLN2, a component of the perinodal extracellular matrix, is a hallmark of physiological brain aging in mice

doi: 10.1101/2025.01.09.632092

Figure Lengend Snippet: (A) Immunohistochemistry images of the cerebellum from mice fed AIN-76A control chow or AIN-76A containing 290 mg/kg PLX3397 from one month to three months of age, stained with anti-ionized calcium-binding adaptor molecule 1 (Iba1) antibody as a microglia marker. n = 3. (B) Immunohistochemistry images of the cerebellum from the mice in (A), stained with anti-HAPLN2 antibody. n = 3. (C) Quantification of the area of each HAPLN2-positive punctum in (B). (D) Immunoblot analysis of the cerebellum from the mice in (B), fractionated as described in . n = 3. (E) Densitometric quantitation of (D). Protein expression levels were normalized to α-tubulin in the S1 fractions. Error bars represent mean ± S.D. P -values were calculated using Student’s t -test.

Article Snippet: PLX3397 [ ] was synthesized as described previously [ ] and formulated at 290 mg/kg in AIN-76A standard chow by Research Diets Inc.

Techniques: Immunohistochemistry, Control, Staining, Binding Assay, Marker, Western Blot, Quantitation Assay, Expressing