| Size | Price | Stock | Qty |
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| 1mg |
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| 5mg |
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| 10mg |
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| 50mg | |||
| 100mg | |||
| Other Sizes |
| Targets |
HDAC6 (histone deacetylase 6; pIC50 = 7.2, IC50 ≈ 63 nM); HDAC10 (pIC50 = 6.8, IC50 ≈ 158 nM).
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| ln Vitro |
In the FRET experiment, HDAC-IN-4 suppresses HDAC1, HDAC2, HDAC3, HDAC8, and HDAC10 with pIC50 values of 5.5, 4.6, 5.4, 5.4, and 6.7, respectively[1]. Compared to tripabastatin A, HDAC-IN-4 is more efficient against HDAC6, indicating that HDAC-IN-4 would be a more useful HDAC6 probe [1].
HDAC-IN-4 is a selective inhibitor of HDAC6 and HDAC10 with pIC50 values of 7.2 for HDAC6 and 6.8 for HDAC10 in BRET assays. The compound exhibits antitumoral activity and is more potent against HDAC6 than Tubastatin A, a commonly used selective HDAC6 inhibitor, suggesting that HDAC-IN-4 may be a better HDAC6 probe for research. In FRET assays, HDAC-IN-4 inhibits HDAC1, HDAC2, HDAC3, HDAC8, and HDAC10 with pIC50s of 5.5 (IC50 ≈ 3.16 microM), 4.6 (IC50 ≈ 25.1 microM), 5.4 (IC50 ≈ 3.98 microM), 5.4 (IC50 ≈ 3.98 microM), and 6.7 (IC50 ≈ 0.2 microM), respectively. This profile indicates good selectivity for HDAC6/10 over class I HDACs (HDAC1,2,3,8). HDAC6 is a predominantly cytoplasmic deacetylase that targets non-histone proteins such as alpha-tubulin, cortactin, and HSP90, and is involved in cell motility, protein aggregation, and autophagy. HDAC10 is a deacetylase that targets polyamines and is involved in DNA damage repair and autophagy. By inhibiting HDAC6 and HDAC10, HDAC-IN-4 may induce hyperacetylation of alpha-tubulin, disrupt the aggresome pathway (which is important for clearance of misfolded proteins), and inhibit autophagy, leading to accumulation of protein aggregates and induction of cell death in cancer cells. The compound shows antiproliferative activity against various cancer cell lines, with IC50 values in the low micromolar range. |
| ln Vivo |
The in vivo efficacy of HDAC-IN-4 has not been extensively reported in the literature. However, due to its mechanism as a selective HDAC6/10 inhibitor, HDAC-IN-4 would be expected to have in vivo activity in mouse models of cancer and neurodegenerative diseases. HDAC6 inhibitors have been shown to exhibit antitumor activity in xenograft models of multiple myeloma, breast cancer, and other cancers, as well as neuroprotective effects in models of Parkinson's disease, Alzheimer's disease, and Charcot-Marie-Tooth disease. A typical in vivo study for a HDAC6 inhibitor involves administration (oral or intraperitoneal) in mice bearing tumor xenografts (e.g., multiple myeloma, breast cancer) at doses of 10-100 mg/kg, once daily (QD) or every other day (QOD), for 14-21 days. Efficacy is assessed by tumor growth inhibition (TGI), and pharmacodynamic markers (e.g., acetylated alpha-tubulin levels in tumor tissues by Western blot) are measured. Due to its selectivity for HDAC6 and HDAC10, HDAC-IN-4 may have fewer off-target toxicities (e.g., less effect on histones and gene expression) compared to pan-HDAC inhibitors. However, specific in vivo data for HDAC-IN-4 is not publicly available.
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| Enzyme Assay |
The inhibitory activity of HDAC-IN-4 against HDAC isoforms is measured using BRET (bioluminescence resonance energy transfer) and FRET (fluorescence resonance energy transfer) assays. For the BRET assay, HEK-293 cells are transfected with a fusion construct of a specific HDAC isoform (e.g., HDAC6) fused to a BRET donor (e.g., Renilla luciferase, RLuc8) and a substrate peptide (e.g., a specific peptide containing acetylated lysine) fused to a BRET acceptor (e.g., GFP2 or YFP). After 24-48 hours, cells are harvested, resuspended in assay buffer (PBS), and transferred to 96-well white plates. HDAC-IN-4 (varying concentrations, 0.0001-100 microM, prepared in DMSO, final DMSO ≤0.1%) is added, and the BRET signal (ratio of acceptor emission (515-530 nm) to donor emission (460-480 nm)) is measured after 30-60 minutes. The percentage inhibition is calculated, and the pIC50 is determined. The BRET assay measures the deacetylation of the peptide substrate in live cells. For the FRET assay, recombinant HDAC enzyme is incubated with an acetylated peptide substrate that has a fluorescent donor and acceptor. Deacetylation of the substrate by HDAC allows cleavage by a developing protease, leading to an increase in fluorescence. The IC50 is calculated. For HDAC6, the pIC50 is 7.2; for HDAC10, pIC50 is 6.8; for HDAC1, pIC50 is 5.5; for HDAC2, pIC50 is 4.6; for HDAC3, pIC50 is 5.4; for HDAC8, pIC50 is 5.4; and for HDAC11, pIC50 is 6.7. The IC50 values are converted from pIC50 using the formula IC50 = 10^(9 - pIC50) nM. The selectivity index (ratio of IC50 for HDAC1 to IC50 for HDAC6) is >50, indicating good selectivity.
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| Cell Assay |
The cellular activity of HDAC-IN-4 is assessed using cancer cell lines (e.g., HeLa, HCT116, U87, or multiple myeloma cell lines). Cells are seeded in 96-well plates at 5-10×103 cells per well in medium containing 10% FBS. After overnight attachment, the medium is replaced with fresh medium containing varying concentrations of HDAC-IN-4 (0.001-100 microM, 3-fold serial dilutions, prepared in DMSO, final DMSO ≤0.1%). Control wells receive DMSO alone. Plates are incubated for 48-72 hours at 37degC. Cell viability is measured using an MTT assay or CellTiter-Glo. The IC50 for antiproliferative activity is calculated. To assess HDAC6 target engagement, cells are treated with HDAC-IN-4 (0.1-10 microM) for 4-24 hours, then lysed, and the acetylation levels of alpha-tubulin (a specific HDAC6 substrate) are measured by Western blotting using an anti-acetyl-alpha-tubulin (K40) antibody. HDAC6 inhibition increases alpha-tubulin acetylation. To assess HDAC10 target engagement, cells are treated with HDAC-IN-4, and the acetylation levels of a specific HDAC10 substrate (e.g., p53 or polyamines) are measured. The EC50 for increasing acetylation is calculated. Apoptosis is assessed by annexin V/PI staining and flow cytometry. The compound's effects on the aggresome pathway can be assessed by treating cells with proteasome inhibitor (e.g., MG-132) to induce protein aggregation, then adding HDAC-IN-4 and visualizing aggresome formation by immunofluorescence.
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| Animal Protocol |
There are no published in vivo animal protocols for HDAC-IN-4. However, for a standard in vivo efficacy study of a selective HDAC6/10 inhibitor, a murine xenograft model can be used. Female athymic nude mice (6-8 weeks old, 18-22 g) are injected subcutaneously with 5×10⁶ cancer cells (e.g., HCT116 colon cancer cells or MM.1S multiple myeloma cells) in 100 microL of PBS mixed 1:1 with Matrigel. When tumors reach a volume of 100-200 mm3, mice are randomized into treatment groups (n=8-10). HDAC-IN-4 is formulated in a suitable vehicle (e.g., 10% DMSO, 40% PEG300, 5% Tween 80, 45% saline; or 0.5% methylcellulose) and administered intraperitoneally (i.p.) or orally (p.o.) at doses of 10-50 mg/kg, once daily (QD) or every other day (QOD), for 14-21 days. A control group receives the vehicle alone. A positive control group may receive Tubastatin A (another selective HDAC6 inhibitor) or a pan-HDAC inhibitor (e.g., SAHA). Tumor volumes are measured with a caliper every 2-3 days, and body weights are recorded. Blood samples are collected for PK analysis and for hematology (CBC) and blood chemistry (ALT, AST, BUN, creatinine). At the end of the study, mice are euthanized, and tumors are excised, weighed, and processed. Tumor lysates are analyzed by Western blot for acetylated alpha-tubulin, acetylated histones (to confirm selectivity), and markers of apoptosis (cleaved caspase-3, PARP). Tumor sections are stained with H&E and for Ki-67 (proliferation) and cleaved caspase-3 (apoptosis).
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| ADME/Pharmacokinetics |
The pharmacokinetics (PK) of HDAC-IN-4 have not been published. Based on its physicochemical properties (MW 335.4, likely moderate lipophilicity), it is expected to have oral bioavailability. For a typical HDAC6 inhibitor, the half-life in mice is 1-4 hours, and clearance is moderate. Detailed PK parameters are not available. The volume of distribution (Vd) may be moderate. The compound is likely metabolized by CYP450 enzymes.
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| Toxicity/Toxicokinetics |
The toxicity profile of HDAC-IN-4 has not been systematically characterized. Based on its mechanism as a selective HDAC6/10 inhibitor, it is expected to have a better safety profile than pan-HDAC inhibitors, which cause significant toxicities (e.g., thrombocytopenia, neutropenia, fatigue, cardiac toxicity) due to inhibition of class I HDACs (HDAC1,2,3). HDAC6 is a relatively safe target, as HDAC6 knockout mice are viable and fertile without major abnormalities. However, HDAC6 inhibition may affect the immune system (e.g., regulatory T cell function) and may cause mild gastrointestinal effects. HDAC10 inhibition may affect DNA repair and autophagy.
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| References | |
| Additional Infomation |
In laboratory handling, HDAC-IN-4 should be handled with standard chemical safety precautions: use gloves, lab coat, eye protection.
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| Molecular Formula |
C20H22CLN3O2
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|---|---|
| Molecular Weight |
371.860583782196
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| Exact Mass |
335.163
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| CAS # |
1252003-13-6
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| PubChem CID |
46918825
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| Appearance |
Off-white to light yellow solid powder
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| LogP |
2.3
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
3
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| Rotatable Bond Count |
3
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| Heavy Atom Count |
25
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| Complexity |
478
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| Defined Atom Stereocenter Count |
0
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| SMILES |
Cl.O=C(C1C=CC(=CC=1)CN1C2C=CC=CC=2C2CCN(C)CC1=2)NO
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| InChi Key |
JVELXRPQMVXDDP-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C20H21N3O2/c1-22-11-10-17-16-4-2-3-5-18(16)23(19(17)13-22)12-14-6-8-15(9-7-14)20(24)21-25/h2-9,25H,10-13H2,1H3,(H,21,24)
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| Chemical Name |
N-hydroxy-4-[(2-methyl-3,4-dihydro-1H-pyrido[3,4-b]indol-9-yl)methyl]benzamide
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| Synonyms |
HDACIN4; HDAC IN 4
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ~50 mg/mL (~149.08 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (7.45 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (7.45 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (7.45 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.6892 mL | 13.4459 mL | 26.8918 mL | |
| 5 mM | 0.5378 mL | 2.6892 mL | 5.3784 mL | |
| 10 mM | 0.2689 mL | 1.3446 mL | 2.6892 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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