In murine precision-cut lung slices, 10 µM MG149 produced a small significant reduction in acetylation of histone H4 residues 4–17 after 20 h; 5 µM did not. A separate SAHA comparison also showed reduced acetylation with 10 µM MG149 versus SAHA alone. The assay measured a peptide containing several lysines and did not isolate H4K16 acetylation. [2]
In stimulated murine lung slices, MG149 reduced TNFα, iNOS, IL1β and IL12b mRNA, with no significant reduction in IL6 or KC. Effects varied by gene and concentration. The DMF solvent increased IL12b expression, underscoring the importance of the vehicle comparison. These findings are specific to the LPS/IFNγ-stimulated ex vivo model. [2]
In Parkin-overexpressing SH-SY5Y cells, acute MG149 pretreatment reduced oligomycin/antimycin-induced PINK1/Parkin initiation markers; suppression of ubiquitin phosphorylation was not sustained after 24–48 h pretreatment. In a separate mitoSRAI reporter experiment, MG149 increased mitochondrial delivery to lysosomes without the stimulus and did not prevent stimulus-induced delivery. Reduced initiation markers therefore did not imply complete mitophagy blockade. [3]
Recombinant acetyltransferase inhibition panel
Biochemical assay · Human recombinant Tip60, MOF, p300 and PCAF; radiolabeled acetyl transfer to histone peptides. [1]
Compound 20 inhibited Tip60 and MOF in the micromolar range. Table 2 reported IC50 >200 µM for p300 and PCAF; insolubility prevented determination of exact higher IC50 values.
Literature-Reported Treatment Conditions
- Treatment concentration
- Concentration–response series; at least seven concentrations spanning at least 20-fold around the IC50.
- Exposure duration
- Tip60 7 min; MOF 5 min; p300 5 min; PCAF 3.5 min.
- Method
- Radiometric HAT assay at 30°C; 1 µM acetyl-CoA and 100 µM H4-20 for Tip60/MOF/p300 or H3-20 for PCAF. Enzyme concentrations: 10 nM Tip60, 1 nM MOF, 5 nM p300 and 1 nM PCAF. Enzyme-free background was subtracted.
- Vehicle
- 2% DMSO in assay buffer.
- Reported limits
- Table 2 does not define the meaning of its ± values. Methods state assays were performed at least twice; do not import Figure 1 triplicate SD semantics into Table 2.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: 20
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Table 1 compound 20; Results 3.1; Table 2; Figure 1; Methods 5.3
Tip60 acetyltransferase inhibition: Radiometric assay; reported ± definition unspecified. — Table 2; Methods 5.3
MOF acetyltransferase inhibition: Radiometric assay; reported ± definition unspecified. — Table 2; Methods 5.3
PCAF acetyltransferase inhibition: Table 2 also lists 3% inhibition. — Table 2; Methods 5.3
p300 acetyltransferase inhibition: Table 2 also lists 30% inhibition; do not describe p300 as universally unaffected. — Table 2; Methods 5.3
Tip60 inhibition kinetics relative to two substrates
Biochemical assay · Recombinant human Tip60 with varied acetyl-CoA or H4-20 peptide. [1]
Steady-state fits supported predominantly competitive inhibition relative to acetyl-CoA and a noncompetitive pattern relative to H4-20. A measurable intercept constant indicated possible additional weak interaction.
Literature-Reported Treatment Conditions
- Treatment concentration
- Compound 20: 0, 80 and 120 µM.
- Method
- Initial-rate radiometric assays. For varied acetyl-CoA: 50 nM Tip60 and 100 µM H4-20. For varied H4-20: 25 nM Tip60 and 2 µM acetyl-CoA. Mixed-inhibition equation fitted to initial velocities.
- Vehicle
- 2% DMSO in common radiometric assay buffer.
- Reported limits
- Kinetic reaction duration and fit uncertainty are not specified in the read kinetic report; potency-assay reaction times are not assigned to this design.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: 20
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Results 3.2; Figure 2a–d and caption; Methods 5.3
Inhibition pattern relative to acetyl-CoA: Nonlinear mixed-inhibition fit; authors interpret as essentially competitive. — Results 3.2; Figure 2a–b and caption
Inhibition pattern relative to histone H4-20: Authors describe noncompetitive inhibition against peptide substrate. — Results 3.2; Figure 2c–d and caption
Histone acetylation in murine lung slices
Cell-based study · Murine precision-cut lung slices (ex vivo), stimulated with LPS and IFNγ. [2]
MG149 at 10 µM, but not 5 µM, produced a small significant shift toward unmodified histone H4 residues 4–17. With SAHA, 10 µM MG149 also reduced acetylation relative to SAHA alone. This peptide-level assay does not uniquely quantify H4K16 acetylation.
Literature-Reported Treatment Conditions
- Treatment concentration
- MG149 5 or 10 µM.
- Exposure duration
- 20 h total: 16 h pretreatment followed by 4 h stimulation in continued inhibitor presence.
- Method
- Histone extraction, acetic anhydride-d6 derivatization and LC-MS/MS quantification of acetylation states of H4 residues 4–17; n = 3 independent experiments; mean ± SD; one-way ANOVA with Bonferroni comparisons.
- Vehicle
- 0.2% DMF.
- Co-treatment
- LPS and IFNγ, 10 ng/mL each, during the final 4 h. Separate comparison with SAHA 2 µM, including SAHA-only control.
- Reported limits
- Animal donor characteristics are referred to a previous preparation protocol. Supporting LDH and RAW264.7 datasets were not retrieved; no noncytotoxic-dose or detailed macrophage claim is adopted.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: MG149
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Figure 1; Methods 2.1–2.2, 2.5–2.8; Results 3.2; Figure 3A–B and caption
Acetylation-state distribution of histone H4 residues 4–17: LC-MS/MS; mean ± SD of 3 independent experiments. No precise percentage inferred from bars. — Results 3.2; Figure 3A–B and caption; Methods 2.5–2.6
Inflammatory transcript response in lung slices
Cell-based study · LPS/IFNγ-stimulated murine precision-cut lung slices (ex vivo). [2]
MG149 reduced TNFα, iNOS, IL1β and IL12b mRNA under the tested conditions, while IL6 and KC did not change significantly. The solvent itself increased IL12b expression, making the matched solvent control essential.
Literature-Reported Treatment Conditions
- Treatment concentration
- MG149 5 or 10 µM.
- Exposure duration
- 16 h pretreatment plus 4 h stimulation with MG149 maintained.
- Method
- RT-qPCR, normalized to GAPDH. Mean ± SD from 3–4 independent experiments; one-way ANOVA with Bonferroni post hoc tests.
- Vehicle
- 0.2% DMF; untreated, stimulus-only and stimulus-plus-DMF controls shown.
- Co-treatment
- Final 4 h: LPS and IFNγ at 10 ng/mL each.
- Reported limits
- No exact transcript fold changes reconstructed from plotted bars; supplier identity does not establish Adooq material use.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: MG149
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Methods 2.1–2.3, 2.8; Results 3.3; Figure 5A–F and caption
Inflammatory mRNA response: RT-qPCR; Figure 5 labels verified visually: A TNFα, B iNOS, C IL1β, D IL12b. — Figure 5A–D and caption; Results 3.3; Methods 2.3, 2.8
IL6 and KC mRNA: RT-qPCR; KC was not increased by the inflammatory stimulus; DMF increased IL12b in panel D. — Figure 5E–F; Results 3.3
Acute effects on PINK1/Parkin mitophagy initiation
Cell-based study · Flag-Parkin-overexpressing human SH-SY5Y neuroblastoma cells. [3]
MG149 reduced oligomycin/antimycin-induced ubiquitin Ser65 phosphorylation after 1–3 h pretreatment, but the same suppression was not sustained at 24–48 h. Acute immunoblots also showed reduced Parkin Ser65 phosphorylation and reduced MFN2 degradation. NU9056 did not reproduce these effects.
Literature-Reported Treatment Conditions
- Treatment concentration
- MG149 100 µM; comparator NU9056 10 µM.
- Exposure duration
- MG149/NU9056 pretreatment for 1, 3, 24 or 48 h, then 3 h stimulus for imaging; 3 h pretreatment plus 3 h stimulus for immunoblotting.
- Method
- Immunofluorescence of pUb(Ser65) and TOM20 (n = 3); immunoblots for pUb(Ser65), pParkin(Ser65), MFN2 and GAPDH (n = 3–5). Mean ± SD; two-way ANOVA with Dunnett correction.
- Vehicle
- Equal-volume DMSO controls.
- Co-treatment
- Oligomycin 1 µM plus antimycin A 1 µM (O/A), or DMSO control.
- Reported limits
- Read main text names MG149 without specifying supplier, salt or analytical lot verification. Exact fractional reductions are not read from curves.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: MG149
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Results Sec3; Figure 1A–D; Methods Sec12, Sec15–Sec16, Sec19–Sec20, Sec24; Discussion
PINK1/Parkin initiation markers: Viewed Figure 1; imaging and immunoblots. MG149 alone produced a small nonsignificant pUb increase at acute timepoints. — Results Sec3; Figure 1A–D and caption
Mitochondrial delivery to lysosomes despite reduced initiation markers
Cell-based study · mitoSRAI reporter-expressing, Flag-Parkin-overexpressing SH-SY5Y cells. [3]
Under a 3 h pretreatment/3 h stimulation design, MG149 increased reporter-defined mitochondrial delivery to lysosomes without O/A. It did not prevent O/A-induced delivery, limiting the interpretation of reduced initiation markers as global mitophagy blockade.
Literature-Reported Treatment Conditions
- Treatment concentration
- MG149 100 µM; NU9056 comparator 10 µM.
- Exposure duration
- 3 h pretreatment followed by 3 h with DMSO or O/A.
- Method
- Fixed-cell mitoSRAI reporter: lysosomal mitochondrial area (TOLLES signal without YPet) divided by total TOLLES area; n = 6 independent biological experiments; mean ± SD; two-way ANOVA with Dunnett correction.
- Vehicle
- Equal-volume DMSO.
- Co-treatment
- DMSO or oligomycin 1 µM plus antimycin A 1 µM.
- Reported limits
- This adopted design is Figure 2C–D only. Separate live mitoKeima and PINK1-knockout experiments are not merged into this study or used to assign additional conditions.
Methods and source locations
Compound reported in the paper: MG149; Compound label in the paper: MG149
Research findings for the reported MG149 material and model; no salt conversion, product-lot equivalence, clinical efficacy or target-exclusive cellular action established.
Results Sec4; Figure 2C–D; Methods Sec12, Sec16, Sec23–Sec24; Discussion
Mitochondrial delivery to lysosomes: MitoSRAI index normalized to control O/A. No exact index values inferred from bars. — Results Sec4; Figure 2C–D and caption; Methods Sec23–Sec24
