SJA710-6 targets mesenchymal stem cells, promoting their differentiation into hepatocyte-like cells. Its mechanism involves the activation of hepatic differentiation pathways, leading to the expression of liver-specific genes and the acquisition of hepatocyte functions. The compound does not target a specific enzyme or receptor but rather modulates the cellular differentiation program. Its ability to selectively differentiate MSCs toward hepatocyte-like cells makes it a valuable tool for studying liver development and regeneration.

SJA710-6 at 5 μM induced rMSCs to exhibit polygonal hepatocyte-like morphology with granular cytoplasm and refractile borders after 14 days of differentiation [1].
Periodic acid-Schiff (PAS) staining showed that SJA710-6-treated rMSCs accumulated glycogen (magenta staining) comparable to the positive control (growth factors), while DMSO control showed no glycogen storage [1].
Albumin secretion measured by ELISA: SJA710-6-treated cells secreted 36.54 ± 7.73 μg/mL albumin, significantly higher than DMSO control (13.77 ± 3.16 μg/mL, p < 0.01) [1].
Urea production after exposure to 6 mM NH₄Cl for 24 h: SJA710-6-differentiated rMSCs produced 12.63 ± 3.17 μg/mL urea, compared to 7.61 ± 1.04 μg/mL for DMSO control (p < 0.01) [1].
LDL uptake: SJA710-6-treated cells showed significantly increased uptake of Dil-Ac-LDL (1641.39 ± 326.62 ng/well) versus DMSO control (352.48 ± 88.26 ng/well, p < 0.01) [1].
Flow cytometry analysis after 24 days of induction: 47.14% of SJA710-6-treated cells exhibited hepatic differentiation, comparable to the growth factor positive control [1].
RT-PCR analysis after 28 days: SJA710-6 treatment significantly increased mRNA expression of hepatocyte-specific genes albumin, AFP, CK18, c-Met, CYP1A1, CYP2B1, and HNF3β, whereas DMSO control expressed none of these genes [1].
Western blot analysis: SJA710-6 treatment significantly enhanced FoxH1 protein expression compared to DMSO control (p < 0.01), similar to the growth factor positive control [1].
In vitro, SJA710-6 induces the differentiation of rat mesenchymal stem cells (rMSCs) toward hepatocyte-like cells. Within 14 days of treatment with SJA710-6, the differentiated cells exhibit typical morphological characteristics of hepatocytes. These cells are able to store glycogen, secrete albumin, produce urea, and uptake LDL. The compound also increases the mRNA levels of hepatocyte-specific genes such as albumin, AFP, and CK18. These results demonstrate the potent hepatic differentiation-inducing activity of SJA710-6.

SJA710-6 does not have established in vivo pharmacological activity as a therapeutic agent. Its primary application is in regenerative medicine research, where it is used to study the potential of stem cell differentiation for liver disease treatment. The compound's ability to promote the differentiation of MSCs into functional hepatocyte-like cells suggests potential for cell-based therapies for liver diseases, but specific in vivo efficacy data in animal models are not extensively detailed in the available literature.

In vitro differentiation assays for SJA710-6 involve culturing rat mesenchymal stem cells (rMSCs) in appropriate media supplemented with the compound. Cells are treated with SJA710-6 for up to 14 days. The differentiation into hepatocyte-like cells is assessed by morphological examination, as well as by measuring the expression of hepatocyte-specific genes (albumin, AFP, CK18) using RT-PCR. Functional assays include glycogen storage (PAS staining), albumin secretion (ELISA), urea production (colorimetric assay), and LDL uptake (fluorescence microscopy).

Cell isolation: rMSCs were obtained by flushing femurs and tibias of 6-week-old female Sprague-Dawley rats with D-Hanks' solution. Cells were cultured in low-glucose DMEM with 10% FBS, 10 mM HEPES, 2 mM glutamine, 100 U/mL penicillin, 100 μg/mL streptomycin at 37 °C, 5% CO₂ [1].
Hepatic differentiation protocol (4 steps): Step 1 (pre-induction): rMSCs in serum-free DMEM for 24 h. Step 2 (induction): 2% FBS/DMEM with 100 ng/mL activin A and 10 ng/mL FGF4 for 2 days. Step 3 (differentiation): 2% FBS/DMEM with 1× ITS and either DMSO (negative control), growth factors (10 ng/mL FGF4 + 20 ng/mL HGF, positive control), or 5 μM SJA710-6 for 7 days. Step 4 (maturation): 2% FBS/DMEM with 1 μM dexamethasone, 1× ITS and same treatments for 14 days. Media changed every 3 days [1].
PAS staining: Cells fixed with 4% formaldehyde, permeabilized with 0.1% Triton X-100, oxidized with 1% periodic acid for 1 h at 22 °C, rinsed, treated with Schiff's reagent for 1 h, then examined under light microscope [1].
Albumin ELISA: Culture medium collected after 24 h was analyzed using an ELISA quantitation kit according to manufacturer's instructions; absorbance read on a plate reader [1].
Urea production assay: Cells exposed to 6 mM NH₄Cl for 24 h; culture medium urea concentration measured using an automated chemistry analyzer with a urea assay kit [1].
LDL uptake assay: Differentiated cells incubated with 10 μg/mL Dil-Ac-LDL in high-glucose DMEM for 24 h at 37 °C; supernatant collected and assayed per manufacturer's instructions [1].
Flow cytometry: After 24 days of differentiation, cells treated with brefeldin A (3 μg/mL) for 6 h, fixed in 70% cold ethanol overnight, permeabilized with D-Hanks' solution containing 0.3% Triton X-100, 0.1% NaN₃, 0.1% saponin, 10 mM HEPES, 1% normal serum for 15 min. Cells incubated with primary goat anti-albumin antibody, then with FITC-conjugated rabbit anti-goat secondary antibody, and analyzed by flow cytometry [1].
RT-PCR: Total RNA extracted with TRIzol, reverse transcribed to cDNA. PCR amplification at 94 °C for 40 s, 56–62 °C for 50 s, 72 °C for 60 s for 35 cycles after initial denaturation at 94 °C for 5 min. Primers for albumin, AFP, CK18, c-Met, CYP1A1, CYP2B1, PEPCK, FoxH1, HNF4α, HNF3β, and GAPDH were used [1].
Western blot: Cell lysates prepared with protein extraction reagent plus protease inhibitors. Protein samples heated at 100 °C for 5 min, separated on 20% SDS-PAGE, transferred to PVDF membrane. Membrane blocked for ≥1.5 h, then incubated overnight at 4 °C with primary antibodies against FoxH1 or β-actin. After washing, incubated with HRP-conjugated secondary antibodies for ≥1.5 h at room temperature, developed using chemiluminescence, and images captured on X-ray film [1].
In vitro cell-based assays for SJA710-6 are conducted using rat mesenchymal stem cells (rMSCs). Cells are cultured in appropriate media (e.g., DMEM with 10% FBS) and treated with SJA710-6 at various concentrations for up to 14 days. The differentiation into hepatocyte-like cells is assessed by morphological examination, immunocytochemistry for hepatocyte markers (albumin, CK18, AFP), and functional assays (glycogen storage, albumin secretion, urea production, LDL uptake). Gene expression of hepatocyte-specific markers is analyzed by RT-PCR. Cell viability is assessed by MTT assays.

In vivo animal studies for SJA710-6 are not extensively detailed in the available literature. Potential studies could involve transplanting MSCs that have been pre-treated with SJA710-6 into animal models of liver disease to assess their ability to engraft, differentiate, and restore liver function. Alternatively, the compound could be administered directly to promote endogenous hepatic differentiation. However, specific protocols and efficacy data are not widely available in the public domain.

SJA710-6 has a molecular weight of approximately 439.33 g/mol and a molecular formula of C₂₂H₂₀BrFN₄. Specific pharmacokinetic properties, such as half-life and bioavailability, are not extensively characterized in the available literature. As a cell-permeable small molecule, it is expected to be able to enter cells and exert its effects. It is soluble in DMSO and is typically used in in vitro studies at concentrations determined by its activity in differentiation assays.

SJA710-6 is a research compound and is not intended for human therapeutic use. Comprehensive toxicological data are not available in the public literature. As a small molecule used in cell culture, it is generally handled with standard laboratory safety precautions. The compound is supplied with a purity of ≥98% and should be stored as recommended by the manufacturer. It is for research use only.

ChemMedChem.2012 Aug;7(8):1447-52.

SJA710-6 was identified through a phenotypic screen using PAS staining as a readout for glycogen accumulation, a hallmark of hepatocyte differentiation. The compound induces rMSCs to differentiate into hepatocyte-like cells with functional characteristics comparable to those induced by growth factors (FGF4 and HGF). The differentiation efficiency with SJA710-6 (47.14%) was similar to that of growth factors. Mechanistically, SJA710-6 enhances FoxH1 expression, suggesting involvement of the Nodal signaling pathway in hepatic fate specification. No in vivo animal efficacy, pharmacokinetic, or toxicity data are reported in this study [1].
SJA710-6 is a novel hepatic differentiation inducer that promotes the differentiation of mesenchymal stem cells into hepatocyte-like cells. It is a cell-permeable imidazopyridinamine compound with significant potential in regenerative medicine, particularly for liver diseases. The compound induces the expression of hepatocyte-specific genes and functional markers, making it a valuable tool for studying liver development, stem cell biology, and potential cell-based therapies. This product is for research use only.

C22H20BRFN4

439.323407173157

438.09

C, 60.15; H, 4.59; Br, 18.19; F, 4.32; N, 12.75

1397255-09-2

71466685

Solid powder

5.8

1

4

6

28

479

0

CCCN1C2=C(C=CC(=N2)NCC3=CC=C(C=C3)F)N=C1C4=CC=C(C=C4)Br

RPNQWWSSADJSGS-UHFFFAOYSA-N

InChI=1S/C22H20BrFN4/c1-2-13-28-21(16-5-7-17(23)8-6-16)26-19-11-12-20(27-22(19)28)25-14-15-3-9-18(24)10-4-15/h3-12H,2,13-14H2,1H3,(H,25,27)

2-(4-bromophenyl)-N-[(4-fluorophenyl)methyl]-3-propylimidazo[4,5-b]pyridin-5-amine

SJA710 6; SJA710-6; SJA-710-6; SJA 710-6

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~569.06 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)