Cat. No.: HY-184378

MDB5

MDB5 is a Hedgehog pathway and Smoothened inhibitor that binds to the 7-TM domain of Smo. MDB5 reduces hepatic stellate cell activation, extracellular matrix-related gene expression, collagen deposition, hydroxyproline content and epithelial-mesenchymal transition, and prevents sinusoidal endothelial cell capillarization. MDB5 induces G1 and S phase cell cycle arrest, triggers apoptosis by upregulating Bax and downregulating Bcl-2, and also decreases oxygen consumption rate, glucose uptake, transglutaminase activity and fibronectin matrix assembly. MDB5 reduces the levels of liver injury markers, hepatic triglyceride deposition and hepatic steatosis, restores the tissue structure of the kidney and spleen, and inhibits pancreatic tumor growth without causing body weight loss. MDB5 can be loaded into PEG-PCC-g-DC micelles, achieving high drug loading, sustained release, improved water solubility, enhanced cellular uptake and optimized hepatic distribution, with good tolerance in mice. MDB5 is applicable to research related to alcohol-associated liver disease, liver fibrosis and pancreatic cancer.

For research use only. We do not sell to patients.

CAS No.: 2922221-22-3
Formula: C₃₁H₂₃Cl₂N₅O₂
Molecular Weight:568.45

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Please store the product under the recommended conditions in the Certificate of Analysis.

MDB5 Chemical Structure

CAS No.: 2922221-22-3

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Strong Fluorescence Signal / Multi-Disciplinary Applications / High Fluorescence Specificity

Biological Activity
- Description
- In Vitro
- In Vivo
Chemical Information
Purity & Documentation
References
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Biological Activity

Description

MDB5 is a Hedgehog pathway and Smoothened inhibitor that binds to the 7-TM domain of Smo^[1]^[2]^[3]. MDB5 reduces hepatic stellate cell activation, extracellular matrix-related gene expression, collagen deposition, hydroxyproline content and epithelial-mesenchymal transition, and prevents sinusoidal endothelial cell capillarization^[1]^[2]. MDB5 induces G1 and S phase cell cycle arrest, triggers apoptosis by upregulating Bax and downregulating Bcl-2, and also decreases oxygen consumption rate, glucose uptake, transglutaminase activity and fibronectin matrix assembly^[2]^[3]. MDB5 reduces the levels of liver injury markers, hepatic triglyceride deposition and hepatic steatosis, restores the tissue structure of the kidney and spleen, and inhibits pancreatic tumor growth without causing body weight loss^[1]^[2]^[3]. MDB5 can be loaded into PEG-PCC-g-DC micelles, achieving high drug loading, sustained release, improved water solubility, enhanced cellular uptake and optimized hepatic distribution, with good tolerance in mice^[2]. MDB5 is applicable to research related to alcohol-associated liver disease, liver fibrosis and pancreatic cancer^[1]^[2]^[3].

All Bcl-2 Family Isoforms :

In Vitro

OthersAnti-cancerCell ProliferationCell Cycle RegulationMitochondrial FunctionProtein/Amino Acid MetabolismApoptosis

Others

MDB5 (50 μM) inhibits ethanol-induced upregulation of Gli1 and Gli2 expression in AML12 mouse hepatocytes^[1].

Others

MDB5 (50 μM; 48 h) regulates the expression of fibrosis-related genes (Smad7, Tgf-β1, Gli1) in ethanol-exposed HSC-T6 rat hepatic stellate cells, and acts synergistically with anti-miR-96 to correct dysregulated gene expression^[1].

Anti-cancer

Cell Proliferation

MDB5 (1-100 μM; 48 h) potently inhibits the viability of HSC-T6 cells, with an IC₅₀ of 25 μM after 48 h of incubation^[2].

Anti-cancer

Cell Cycle Regulation

MDB5 (50 μM; 48 h) disrupts the cell cycle progression of HSC-T6 cells by increasing the proportions of cells in the G₀/G₁ and G₂/M phases and decreasing the proportion of cells in the S phase^[2].

Others

MDB5 (50 μM; 48 h) potently downregulates components of the Hh pathway (GLI1, PTCH1), the metabolic enzyme GLS1, and the fibrosis marker α-SMA at both mRNA and protein levels in HSC-T6 cells^[2].

Anti-cancer

Mitochondrial Function

MDB5 (50 μM; 24 h) significantly reduces the basal oxygen consumption rate of HSC-T6 cells, and this effect appears after 24 h of cell treatment with 50 μM, indicating that it inhibits mitochondrial metabolic activity. Meanwhile, it significantly suppresses glucose uptake by HSC-T6 cells^[2].

Anti-cancer

Protein/Amino Acid Metabolism

Treatment with MDB5 (25 μM; 24 h) at the IC₅₀ concentration of 25 μM for 24 h significantly reduces transglutaminase activity in HSC-T6 cells^[2].

Others

MDB5 (50 μM; 24 h) significantly inhibits fibronectin matrix assembly in HSC-T6 cells, and after treatment with 50 μM for 24 h, the matrix formation decreases to approximately 80% of that in the control group^[2].

Anti-cancer

Cell Proliferation

Micelle-encapsulated MDB5 (1-100 μM; 48 h) potently inhibits the viability of HSC-T6 cells in a dose-dependent manner^[2].

Others

MDB5-loaded PEG-PCC-g-DC micelles (up to 9.8% w/w; 60 h) exhibit a uniform spherical morphology with a particle size of 40-50 nm, a maximum drug loading capacity of 9.8% w/w, enhanced water solubility, and sustained drug release properties, with approximately 60% of the drug released within 24 h^[2].

Anti-cancer

MDB5 exhibits the strongest binding affinity to wild-type human Smo protein, with a docking score of −12.84 kcal/mol, and forms unique additional interactions within the 7-TM domain^[3].

Anti-cancer

MDB5 (0-100 μM; 72 h) potently reduces the viability of human pancreatic cancer cell line MIA PaCa-2, with an IC₅₀ of 55.57 μM; it decreases the viability of human pancreatic cancer cell line PANC-1 in a dose-dependent manner^[3].

Anti-cancer

MDB5 (incubated for 24 h after siRNA transfection; 50 μM) exerts cytotoxic effects on human pancreatic cancer cells MIA PaCa-2 via the Smo-dependent Hedgehog pathway, as its activity is completely abolished in Smo-knockdown cells^[3].

Anti-cancer

MDB5 (50 μM; 72 h) effectively and significantly downregulates the expression of Hedgehog pathway transcripts (Gli-1, Gli-2, Ptch-1 and Shh) in human pancreatic cancer cell line MIA PaCa-2^[3].

Anti-cancer

Apoptosis

MDB5 (50 μM; 72 h) downregulates Hedgehog pathway proteins (Gli-1, Shh) and the anti-apoptotic protein Bcl-2, while upregulating the pro-apoptotic protein Bax in human pancreatic cancer cells MIA PaCa-2^[3].

Anti-cancer

Apoptosis

MDB5 (50 μM; 72 h) downregulates pancreatic cancer stem cell (CSC) markers (ALDH1/2, CD44, Oct-3/4) in the human pancreatic cancer cell line MIA PaCa-2^[3].

Anti-cancer

Apoptosis

MDB5 (50 μM; 72 h) downregulates Hedgehog pathway proteins (Gli-1, Shh) and the anti-apoptotic protein Bcl-2, while upregulating the pro-apoptotic protein Bax^[3] in human pancreatic cancer cell line PANC-1.

Anti-cancer

Apoptosis

MDB5 (50 μM; 72 h) induces S-phase cell cycle arrest in human pancreatic cancer cell line MIA PaCa-2, and 45.1% of MIA PaCa-2 human pancreatic cancer cells undergo apoptosis^[3].

Anti-cancer

MDB5 (50 μM; 72 h post-seeding) significantly reduces the migratory potential of human pancreatic cancer cell line MIA PaCa-2^[3].

Anti-cancer

MDB5 (50 μM; 7 days) significantly reduces the tumorigenic (colony-forming) potential of human pancreatic cancer cells MIA PaCa-2^[3].

Anti-cancer

MDB5 (50 μM; applied on day 4 of spheroid culture) effectively reduces the volume of MIA PaCa-2 human pancreatic cancer 3D spheroids^[3].

Anti-cancer

Nanoparticle-encapsulated MDB5 (10-100 μM; 72 h) reduces the viability of human pancreatic cancer cell line MIA PaCa-2 in a dose-dependent manner^[3].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

Real Time qPCR^[1]

Cell Line:

HSC-T6 rat hepatic stellate cells (ethanol-exposed)
Concentration:

50 μM
Incubation Time:

48 h
Result:

Increased mRNA expression of Smad7 and decreased mRNA expression of Tgf-β1 and Gli1 compared to untreated or scrambled miRNA-transfected cells.
Showed significantly greater efficiency in correcting dysregulated gene expression when combined with anti-miR-96 than when used alone.

Cell Viability Assay^[2]

Cell Line:

rat hepatic stellate cell line (HSC-T6)
Concentration:

1-100 μM; 25 μM
Incubation Time:

48 h
Result:

Caused a significant dose-dependent decrease in HSC-T6 cell viability at 48 h.
Exhibited a 50% inhibitory concentration (IC₅₀) of 25 μM.
Reduced cell viability at concentrations of 50 μM and 100 μM.

Apoptosis Analysis^[2]

Cell Line:

rat hepatic stellate cell line (HSC-T6)
Concentration:

50 μM
Incubation Time:

48 h
Result:

Resulted in a significantly higher percentage of apoptotic cells (16.3%) untreated control cells.

Cell Cycle Analysis^[2]

Cell Line:

rat hepatic stellate cell line (HSC-T6)
Concentration:

50 μM
Incubation Time:

48 h
Result:

Increased the percentage of cells in the G₀/G₁ phase to 57.25%, the percentage of cells in the G₂/M phase to 12.9%, and decreased the percentage of cells in the S phase to 30.65%, compared to control cells (46.7% G₀/G₁, 7.86% G₂/M, 45.62% S).

Real Time qPCR^[2]

Cell Line:

rat hepatic stellate cell line (HSC-T6)
Concentration:

50 μM
Incubation Time:

48 h
Result:

Caused significant downregulation of GLI1, PTCH1, GLS1, and α-SMA mRNA expression relative to untreated control cells.
Significantly reduced protein levels of GLI1, PTCH1, GLS1, α-SMA, and YAP1.

Cell Viability Assay^[2]

Cell Line:

rat hepatic stellate cell line (HSC-T6)
Concentration:

1-100 μM
Incubation Time:

48 h
Result:

Caused a dose-dependent decrease in HSC-T6 cell viability at 48 h.
Showed enhanced efficacy compared to free MDB5 due to micellar formulation.

Cell Cytotoxicity Assay^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

0-100 μM
Incubation Time:

72 h
Result:

Reduced MIA PaCa-2 cell viability in a dose-dependent manner.
Achieved an IC₅₀ value of 55.57 μM.

Cell Cytotoxicity Assay^[3]

Cell Line:

PANC-1 human pancreatic cancer cells
Concentration:

0-100 μM
Incubation Time:

72 h
Result:

Reduced PANC-1 cell viability in a dose-dependent manner.

Cell Cytotoxicity Assay^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

post 24 h siRNA transfection incubation
Result:

Showed no cytotoxic effect in MIA PaCa-2 cells transfected with Smo-specific siRNA.
Exhibited significant cytotoxicity in cells transfected with non-targeting control siRNA, matching results seen in untransfected cells.

Real Time qPCR^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h
Result:

Significantly reduced the relative expression of Gli-1, Gli-2, Ptch-1, and Shh transcripts.

Western Blot Analysis^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h
Result:

Significantly reduced the protein levels of Gli-1, Shh, and the anti-apoptotic protein Bcl-2.
Increased the protein level of the pro-apoptotic protein Bax.\n
Significantly reduced the protein levels of pancreatic CSC markers ALDH1/2, CD44, and Oct-3/4.

Western Blot Analysis^[3]

Cell Line:

PANC-1 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h
Result:

Significantly reduced the protein levels of Gli-1, Shh, and Bcl-2.
Increased the protein level of Bax.

Apoptosis Analysis^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h
Result:

Induced total apoptosis in 45.1% of MIA PaCa-2 cells.
Showed a significantly greater apoptotic effect than untreated control cells (5.7%).

Cell Cycle Analysis^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h
Result:

Significantly increased the percentage of cells in S-phase to 29.27%.
Significantly reduced the percentage of cells in G1-phase.

Cell Migration Assay^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

50 μM
Incubation Time:

72 h post-seeding
Result:

Significantly reduced the number of migratory MIA PaCa-2 cells.

Cell Cytotoxicity Assay^[3]

Cell Line:

MIA PaCa-2 human pancreatic cancer cells
Concentration:

10-100 μM
Incubation Time:

72 h
Result:

Reduced MIA PaCa-2 cell viability in a dose-dependent manner.
Showed efficacy comparable to free MDB5.

In Vivo

Lipid MetabolismCytokines & ChemokinesProtein/Amino Acid MetabolismAnti-cancerApoptosisCell Cycle RegulationOthers

Lipid Metabolism

Cytokines & Chemokines

MDB5 loaded in MDC2 LNPs (20 mg/kg; i.v.; three times per week; 2 weeks) significantly ameliorates alcohol-associated liver injury in mice, reducing plasma ALT by ~32.8% and AST by ~38.8% relative to untreated ethanol-fed mice, while also reducing hepatic triglyceride levels and normalizing key fibrosis- and inflammation-related protein expression^[1].

Protein/Amino Acid Metabolism

MDB5 loaded micelles (10 mg/kg; i.v.; 3 times weekly; 2 weeks) exerts superior antifibrotic, hepatoprotective, and Hh pathway inhibitory effects compared to GDC-0449 in CBDL-induced liver fibrosis in C57BL/6 mice, including significant reductions in serum liver enzymes, collagen deposition, and Hh target gene expression, and restoration of liver sinusoidal structure^[2].

Anti-cancer

Apoptosis

Cell Cycle Regulation

MDB5 (20 mg/kg; i.v.; thrice weekly; 3 weeks) delivered via mPEG-b-P(CB-co-LA) nanoparticles significantly inhibits subcutaneous pancreatic tumor growth in NSG mice, reducing mean tumor volume to 103.56 mm³ with no systemic toxicity^[3].

Others

MDB5 (20-40 mg/kg; i.p.; thrice weekly; 4 weeks) administered via intraperitoneal emulsion at doses up to 40 mg/kg does not cause significant hepatic injury in NSG mice with subcutaneous pancreatic tumors^[3].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

Animal Model:

C57BL/6 (male)^[1]
Dosage:

20 mg/kg (free MDB5); 20 mg/kg (MDB5 loaded in MDC2 LNPs)
Administration:

i.v.; three times per week; 2 weeks
Result:

Decreased plasma ALT levels to 58.76 U/L and AST levels to 111.5 U/L (MDB5 LNPs group).
Decreased plasma ALT to 70.7 U/L and AST to 137.5 U/L (free MDB5 group).
Reduced hepatic triglyceride levels to 28.3 mg/g (MDB5 LNPs group).
Reduced hepatic protein levels of GLI2 and SHH, restored hepatic FOXO3A staining, and decreased TGF-β1 staining (MDB5 LNPs group).
Reduced liver injury, hepatic steatosis, collagen deposition, and lipid accumulation (MDB5 LNPs group).

Animal Model:

C57BL/6 wild-type (male, 8-10 weeks old, CBDL-induced liver fibrosis)^[2]
Dosage:

10 mg/kg
Administration:

i.v.; 3 times a week; 2 weeks
Result:

Significantly reduced serum levels of ALT, AST, TBIL, and ALP compared to CBDL control and GDC-0449 loaded micelles.
Restored normal liver morphology, reduced hepatic steatosis and lesions, and decreased collagen deposition more effectively than GDC-0449.
Significantly downregulated hepatic expression of Hh pathway components (GLI1, PTCH1), glutaminolysis enzyme GLS1, proliferation marker Ki-67, profibrogenic factor OPN, and mesenchymal marker α-SMA, while restoring epithelial marker E-cadherin expression.
Prevented liver sinusoidal endothelial capillarization, maintaining sinusoidal porosity comparable to sham mice.
Enhanced intrahepatic distribution of fluorescently labeled micelles compared to GDC-0449 loaded micelles.
Fully restored kidney histology and showed superior protective effects on spleen structural restoration relative to GDC-0449.

Animal Model:

NOD.Cg-Prkdcscid Il2rgtm1wjl/SzJ (NSG) (male, 8-10 weeks old, subcutaneous injection of 3×10⁶ MIA PaCa-2 cells to establish flank tumors)^[3]
Dosage:

20 mg/kg
Administration:

i.v.; thrice weekly; 3 weeks
Result:

Reduced final mean tumor volume to 103.56 mm³.
Decreased final mean tumor weight significantly compared to control and GDC-0449 groups.
Reduced Ki-67 staining in treated tumors relative to controls and GDC-0449-treated tumors.
Increased cleaved caspase-3 staining in treated tumors relative to controls and GDC-0449-treated tumors.
Increased E-cadherin staining in treated tumors relative to controls and GDC-0449-treated tumors.
Caused no significant changes in body weight during treatment.
Showed no pathological changes in major organs (kidney, spleen, liver, lung, heart) via H&E staining.

Animal Model:

NOD.Cg-Prkdcscid Il2rgtm1wjl/SzJ (NSG) (male, subcutaneous flank tumors established with MIA PaCa-2 cells grown to 200 mm³)^[3]
Dosage:

20 mg/kg; 40 mg/kg
Administration:

i.p.; thrice weekly; 4 weeks
Result:

Caused no significant changes in hepatic biochemical parameters (alanine aminotransferase, total bilirubin) compared to control mice at 20 mg/kg dose.
Caused no significant changes in hepatic biochemical parameters (alanine aminotransferase, total bilirubin) compared to control mice at 40 mg/kg dose.

Related Protocols :

Chemical Information

CAS No. 2922221-22-3
Molecular Weight 568.45
Formula C₃₁H₂₃Cl₂N₅O₂
SMILES

O=C(C1=CC=C(C(N(CC2=NC=CC=C2)CC3=NC=CC=C3)=O)C=C1Cl)NC4=CC(C5=NC=CC=C5)=C(Cl)C=C4
Shipping

Room temperature in continental US; may vary elsewhere.
Storage

Please store the product under the recommended conditions in the Certificate of Analysis.

Purity & Documentation

Handling Instructions (2659 KB)

References

[1]. Kumar V, et al. Anti-miR-96 and Hh pathway inhibitor MDB5 synergistically ameliorate alcohol-associated liver injury in mice. Biomaterials. 2023 Apr;295:122049. [Content Brief]

[2]. Kumar V, et al. The use of micelles to deliver potential hedgehog pathway inhibitor for the treatment of liver fibrosis. Theranostics. 2019;9(25):7537-7555. [Content Brief]

[3]. Kumar V, et al. Design, Synthesis and Biological Evaluation of novel Hedgehog Inhibitors for treating Pancreatic Cancer. Scientific reports. 2017 May 10;7(1):1665. [Content Brief]

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MDB5

All Bcl-2 Family Isoforms