VEGF-R2
Farnesoid X receptor (FXR) antagonist[2]
(Z)-Guggulsterone (CAS: 39025-23-5) is a broad-spectrum steroid receptor ligand that functions primarily as an antagonist of the farnesoid X receptor (FXR). Regarding steroid hormone receptors, it acts as an antagonist of the mineralocorticoid receptor (MR, Ki = 37 nM), progesterone receptor (PR, Ki = 224 nM), and glucocorticoid receptor (GR, Ki = 252 nM), while serving as a weak agonist of the androgen receptor (AR, Ki = 315 nM). Furthermore, this compound inhibits angiogenesis by suppressing the VEGF-VEGFR2-Akt signaling axis.

In HUVEC, (Z)-GugguLsterone (10, 20 μM; 24 or 48 hours) lowers the levels of VEGF-R2 protein [1]. Through FXR-mediated ACE2 modulation, (Z)-Guggulsterone (10 μM; 24) decreases primary airways, disturbs ACE2 and SHP levels in organoids, and lessens SARS-CoV-2 infection in many cell types [2].

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The z-guggulsterone treatment inhibited capillary-like tube formation (in vitro neovascularization) by human umbilical vein endothelial cells (HUVEC) and migration by HUVEC and DU145 human prostate cancer cells in a concentration- and time-dependent manner. The z- and E-isomers of guggulsterone seemed equipotent as inhibitors of HUVEC tube formation[1].

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Our previous studies have shown that z-guggulsterone, a constituent of Indian Ayurvedic medicinal plant Commiphora mukul, inhibits the growth of human prostate cancer cells by causing apoptosis. We now report a novel response to z-guggulsterone involving the inhibition of angiogenesis in vitro and in vivo. The z-guggulsterone treatment inhibited capillary-like tube formation (in vitro neovascularization) by human umbilical vein endothelial cells (HUVEC) and migration by HUVEC and DU145 human prostate cancer cells in a concentration- and time-dependent manner. The z- and E-isomers of guggulsterone seemed equipotent as inhibitors of HUVEC tube formation. The z-guggulsterone-mediated inhibition of angiogenesis in vitro correlated with the suppression of secretion of proangiogenic growth factors [e.g., vascular endothelial growth factor (VEGF) and granulocyte colony-stimulating factor], down-regulation of VEGF receptor 2 (VEGF-R2) protein level, and inactivation of Akt. The z-guggulsterone-mediated suppression of DU145 cell migration was increased by knockdown of VEGF-R2 protein level. Ectopic expression of constitutively active Akt in DU145 cells conferred protection against z-guggulsterone-mediated inhibition of cell migration [1].

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Treatment with 10 µM (Z)-Guggulsterone (ZGG), an FXR antagonist, reduced FXR signaling, decreased the presence of FXR on the ACE2 promoter, and downregulated ACE2 expression at both transcript and protein levels in primary human cholangiocyte, airway, and intestinal organoids. This reduction in ACE2 expression subsequently decreased susceptibility to SARS-CoV-2 infection in these organoid models when infected with the virus.[2]
In gall bladder cholangiocyte organoids (GCOs) treated with physiological levels of the bile acid CDCA (an FXR agonist) to simulate baseline FXR activation, co-treatment with 10 µM (Z)-Guggulsterone reduced SARS-CoV-2 infection, as measured by viral RNA quantification and immunofluorescence for the viral spike protein 24 hours post-infection.[2]
Knockdown of FXR using shRNAs in cholangiocyte organoids prevented the upregulation of ACE2 upon CDCA treatment and inhibited viral infection independently of CDCA, UDCA, or ZGG treatment. After FXR knockdown, treatment with (Z)-Guggulsterone had no additional effect on viral infection, confirming that its antiviral effect is mediated through FXR.[2]
In HEK293T cells genetically engineered to stably overexpress ACE2 independent of FXR regulation, treatment with (Z)-Guggulsterone did not affect SARS-CoV-2 replication, confirming that its antiviral effect is specifically dependent on its ability to modulate ACE2 expression via FXR.[2]
A luciferase reporter assay containing the FXR response element (IR-1) from the ACE2 promoter showed that treatment with 50 µM (Z)-Guggulsterone reduced the transcriptional activity associated with this element, and site-directed mutagenesis of the IR-1 site abolished this effect, confirming the specificity of FXR binding.[2]

(Z)-Guggulsterone (silica; 1 mg; 5 x weekly) dramatically lowers wet weight and tumor volume [1].
Oral gavage of 1 mg z-guggulsterone/d (five times/wk) to male nude mice inhibited in vivo angiogenesis in DU145-Matrigel plug assay as evidenced by a statistically significant decrease in tumor burden, microvessel area (staining for angiogenic markers factor VIII and CD31), and VEGF-R2 protein expression. In conclusion, the present study reveals that z-guggulsterone inhibits angiogenesis by suppressing the VEGF-VEGF-R2-Akt signaling axis. Together, our results provide compelling rationale for further preclinical and clinical investigation of z-guggulsterone for its efficacy against prostate cancer[1].

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Oral administration of (Z)-Guggulsterone (1 mg per mouse, approximately 40 mg/kg, five times per week) to male nude mice, starting 2 weeks prior to and continuing for 2 weeks after subcutaneous implantation of DU145 cell-containing Matrigel plugs, significantly inhibited in vivo angiogenesis. This was evidenced by a statistically significant decrease in tumor volume and wet tumor weight compared to vehicle-treated controls.[1]
Immunohistochemical analysis of the excised Matrigel plugs showed that (Z)-Guggulsterone administration significantly reduced the microvessel area (based on staining for angiogenic markers factor VIII and CD31) and decreased VEGF-R2 protein expression compared to controls.[1]

The z-guggulsterone-mediated inhibition of angiogenesis in vitro correlated with the suppression of secretion of proangiogenic growth factors [e.g., vascular endothelial growth factor (VEGF) and granulocyte colony-stimulating factor], down-regulation of VEGF receptor 2 (VEGF-R2) protein level, and inactivation of Akt. The z-guggulsterone-mediated suppression of DU145 cell migration was increased by knockdown of VEGF-R2 protein level. Ectopic expression of constitutively active Akt in DU145 cells conferred protection against z-guggulsterone-mediated inhibition of cell migration[3].

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Chromatin immunoprecipitation followed by quantitative PCR (ChIP-qPCR) was performed on cholangiocyte organoids. Cells were incubated with fresh medium containing 100 µM CDCA, UDCA, or (Z)-Guggulsterone for 2 hours before collection. The lysate was incubated overnight with an FXR antibody or non-immune IgG. Immunoprecipitated DNA was purified and analyzed by qPCR using primers flanking the FXR binding site on the ACE2 promoter. The FXR agonist CDCA promoted FXR binding to the ACE2 promoter, and this binding was reduced by the FXR inhibitors UDCA and (Z)-Guggulsterone.[2]
A luciferase reporter assay was used to assess FXR transcriptional activity. Fragments containing the FXR IR-1 response element from the ACE2 or SHP gene were cloned into a pGL3-promoter luciferase vector. Mutants of the IR-1 sites were generated via site-directed mutagenesis. These reporter constructs were co-transfected with an FXR expression plasmid into HEK293 cells. Twenty-four hours post-transfection, cells were treated with 50 µM CDCA, UDCA, or (Z)-Guggulsterone in fresh medium for 8 hours. Luciferase activity was measured and normalized to an empty pGL3 vector. The assay demonstrated that CDCA increased transcriptional activity via the ACE2 IR-1 element, which was reduced by UDCA and (Z)-Guggulsterone. Mutation of the IR-1 site abolished this activity.[2]

Western Blot Analysis [1]
Cell Types: Vascular Endothelial Growth Factor (VEGF)
Tested Concentrations: 10, 20 μM
Incubation Duration: 24 or 48 hrs (hours)
Experimental Results: Caused a decrease in VEGF-R2 protein levels in HUVEC.\n

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\n\nCell Culture and Cell Viability Assay [1]
\nHUVEC were purchased from Clonetics and maintained in endothelial cell growth medium-2 (EGM2 MV SingleQuots) supplemented with 5% fetal bovine serum. Monolayer cultures of DU145 cells were maintained as we have previously described. Stock solutions of each isomer of guggulsterone were prepared in DMSO and diluted with complete medium. An equal volume of DMSO (final concentration <0.2%) was added to the controls. The effects of z- and E-guggulsterone treatments on HUVEC viability was determined by sulforhodamine B assay as we have previously described.\n

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\n\nIn vitro Capillary-Like Tube Structure Formation and Migration Assays [1]
\nThe effects of z- and E-guggulsterone treatments on in vitro angiogenesis were determined by tube formation assay as we have previously reported. The HUVEC seeded on Matrigel differentiate and form capillary-like tube structures. In some tube formation experiments, the HUVEC were exposed to 20 μmol/L of z-guggulsterone for 24 h in the absence or presence of 1 μmol/L of the Akt-1/2 inhibitor. The effect of z-guggulsterone treatment on in vitro migration by HUVEC or DU145 cells was determined using a Transwell Boyden Chamber containing a polycarbonate filter (pore size 8 μm) as we have previously described. In some migration assays, HUVEC or DU145 cells were treated with 20 μmol/L of z-guggulsterone for 24 h in the absence or presence of 1 μmol/L of Akt-1/2 inhibitor.\n

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\n\nImmunoblotting [1]
\nThe immunoblotting of total Akt, phosphorylated Akt, and VEGF-R2 was done as we have previously described. Briefly, HUVEC or DU145 cells were treated with desired concentrations of z-guggulsterone for specified time periods, and both floating and attached cells were collected. The cell lysates were prepared as we have previously described. The lysate proteins were resolved by 6% to 10% SDS-PAGE and transferred onto polyvinylidene fluoride membrane. After treatment with the desired primary and secondary antibodies, the immunoreactive bands were visualized using an enhanced chemiluminescence method. The blots were stripped and reprobed with antiactin antibody to correct for differences in protein loading. Changes in protein levels were determined by densitometric scanning of the immunoreactive bands. The immunoblotting for each protein was done at least twice using independently prepared lysates.\n

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\n\nAnalyses of Growth Factors, Interleukins, and MMPs [1]
\nHUVEC or DU145 cells (2 × 105) were seeded in 24-well plates and allowed to attach by overnight incubation. Cells were treated with the desired concentrations of z-guggulsterone or DMSO (control) for 24 and 48 h. Subsequently, the culture medium was collected and used to determine the secretion of VEGF, EGF, G-CSF, FGF, IL-12, IL-17, MMP-2, and MMP-9 using commercially available ELISA kits as we have previously described.\n

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\n\nRNA Interference of VEGF-R2 [1]
\nRNA interference of VEGF-R2 was done using a VEGF-R2–targeted short interfering RNA (siRNA). A nonspecific control siRNA was purchased from Qiagen. For transfection, DU145 cells (5 × 104) were seeded in six-well plates and allowed to attach overnight. Cells were transfected with 200 nmol/L of control nonspecific siRNA or VEGF-R2–targeted siRNA using OligofectAMINE according to the manufacturer's recommendations. Twenty-four hours after transfection, the cells were treated with DMSO (control) or 20 μmol/L of z-guggulsterone for 24 h. The cells were collected and processed for analysis of migration and immunoblotting as described above.\n

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\n\nEctopic Expression of Constitutively Active Akt [1]
\nDU145 cells were transiently transfected with pCMV6 vector encoding constitutively active Akt-1 (Myr-Akt1-HA) or empty vector using Fugene 6 transfection regent. Briefly, DU145 cells were seeded in six-well plates at a density of 2 × 105 cells/mL and allowed to attach by overnight incubation. Cells were transfected with the expression vector encoding constitutively active Akt or empty vector. Twenty-four hours after transfection, the cells were treated with 20 μmol/L of z-guggulsterone or DMSO (control) for 24 h and processed for immunoblotting of total or phosphorylated Akt levels and migration assay.\n\n

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\n\nChIP [2]
\nApproximately 6 × 106 cells were used for each ChIP, and cells were incubated with fresh medium with 100 μM of CDCA, UDCA/Ursodeoxycholic acid or z-guggulsterone/ZGG 2 h before collection. ChIP was performed using the True Micro ChiP kit according to the manufacturer’s instructions. In brief, following pre-clearing, the lysate was incubated overnight with the FXR antibody (Supplementary Table 1) or non-immune IgG. ChIP was completed and immunoprecipitated DNA was purified using MicroChip DiaPure columns. Samples were analysed by qPCR using the ΔΔCt approach as previously described51 (see Supplementary Table 3 for primer sequences). Primers flanking the FXRE on the well-known FXR target gene OSTα (also known as SLC51A; ref. 54) were used as a positive control, whereas primers flanking a site distant from the FXRE on the ACE2 promoter were used as a negative control. The results were normalized to the enrichment observed with non-immune IgG ChIP controls.\n

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\n\nLuciferase reporter [2]
\nTwo different fragments containing the FXRE IR-1 in the ACE2 gene and in the SHP gene (also known as NR0B2) were amplified using human genomic DNA as a template and inserted onto a pGL3-promoter luciferase vector. The ACE2 and SHP IR-1 mutants were generated using a site-directed mutagenesis approach. Sequences of primers used are reported in Supplementary Table 4. These gene reporter constructs were co-transfected with a commercially available FXR expression plasmid into HEK293 cells using TransIT-293 Transfection Reagent. Twenty-four hours after transfection, cells were treated with 50 μM of CDCA, UDCA/Ursodeoxycholic acid and z-guggulsterone/ZGG in fresh medium for 8 h. Luciferase activity was determined with the GLO-Luciferase Reporter Assay System and values were normalized to the empty pGL3 vector.\n

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\n\nCytotoxicity and viability [2]
\nPrimary organoids were treated with 0.1 μM–100 μM of CDCA, UDCA/Ursodeoxycholic acid or z-guggulsterone/ZGG and the percentage of viable cells was counted using trypan blue and a Countess II cell counter. Cellular viability in primary organoids treated with 10 μM of CDCA, UDCA or ZGG was measured using the resazurin-based assay PrestoBlue using SoftMax Pro 5.4.4 on a SpectraMax M2.\n

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\n\nLuciferase reporter for SARS-CoV-2 replication [2]
\nA luciferase reporter for SARS-CoV-2 protease activity during viral replication was generated as previously described28 In brief, HEK293T reporter cells stably expressing ACE2, renilla luciferase (Rluc) and SARS-CoV-2 papain-like protease-activatable circularly permuted firefly luciferase (FFluc) were seeded in flat-bottomed 96-well plates. The following morning, cells were treated with the indicated doses of CDCA, UDCA/Ursodeoxycholic acid and z-guggulsterone/ZGG, and infected with SARS-CoV-2 at a MOI of 0.01. The SARS-CoV-2 RdRp inhibitor remdesivir and a neutralizing antibody cocktail blocking the interaction between SARS-CoV-2 spike and ACE2 (REGN-COV2) were included as positive controls. After 24 h, cells were lysed in Dual-Glo Luciferase Buffer diluted 1:1 with PBS and 1% NP-40. Lysates were then transferred to opaque 96-well plates, and viral replication quantified as the ratio of FFluc/Rluc activity measured using the Dual-Glo kit according to the manufacturer’s instructions. FFluc/Rluc ratios were expressed as a fraction of the maximum, then analysed using the Sigmoidal, 4PL, X is log(concentration) function in GraphPad Prism.

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Primary human cholangiocyte organoids (GCOs), airway organoids, and intestinal organoids were used. To modulate FXR activity, organoids were incubated with a final concentration of 10 µM CDCA or 10 µM CDCA in combination with 10 µM UDCA or (Z)-Guggulsterone.[2]
For ACE2 expression analysis, organoids were treated as above. RNA was extracted and ACE2 mRNA levels were measured by quantitative PCR (qPCR) using housekeeping genes HMBS or GAPDH for normalization. Protein expression was assessed by immunofluorescence using specific antibodies against ACE2.[2]
For SARS-CoV-2 infection assays, organoids were pretreated with physiological levels of CDCA to simulate baseline FXR activation, in the presence or absence of FXR inhibitors. Organoids were then infected with SARS-CoV-2 at a multiplicity of infection (MOI) of 1 for 2 hours. After washing, infected organoids were cultured and collected at 24 hours post-infection. Viral infection was quantified by qPCR for SARS-CoV-2 RNA (targeting the RdRp gene) normalized to GAPDH, and by immunofluorescence staining for the SARS-CoV-2 spike protein.[2]
Cytotoxicity and viability assays were performed on primary organoids treated with a concentration range of CDCA, UDCA, or (Z)-Guggulsterone (0.1 µM – 100 µM). Viability was assessed using trypan blue exclusion counting and a resazurin-based metabolic activity assay. No cytotoxic effects were observed at the concentrations used in the experiments (e.g., 10 µM).[2]
FXR knockdown was performed in cholangiocyte organoids using lentiviral particles carrying shRNA against human FXR. Control organoids received lentiviral particles with scrambled shRNA. Successfully transduced organoids were selected with puromycin. Experiments assessing ACE2 expression and SARS-CoV-2 infection were performed 10 days after knockdown.[2]

Animal/Disease Models: Male nude mice (5-6 weeks old) were subcutaneously (sc) (sc) implanted with Matrigel plugs containing DU145 cells.
Doses: 1 mg.
Route of Administration: po (po (oral gavage)) 5 times a week.
Experimental Results: Resulting in statistically significant tumor volume and wet tumor weight. reduce.

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In vivo Matrigel Plug Assay [1]
The effect of z-guggulsterone administration on in vivo angiogenesis was determined by DU145-Matrigel plug assay. Male nude mice (5–6 weeks old) were purchased from Taconic and randomized into two groups of five mice per group. The mice were orally gavaged with 0.1 mL of vehicle (PBS) or 1 mg of z-guggulsterone/mouse in 0.1 mL of PBS (corresponding to ∼40 mg z-guggulsterone/kg body weight) five times per week for 2 weeks prior to Matrigel plug implantation. The Matrigel plugs containing 3 × 106 DU145 cells were implanted s.c. into the flank of each mouse. The z-guggulsterone and vehicle administration was continued for two more weeks. Tumor volume was determined by using a caliper as we have previously described. Body weights of the vehicle-treated control and z-guggulsterone–treated mice were recorded weekly. Mice from each group were also monitored for other symptoms of side effects, including food and water withdrawal and impaired posture or movement. Animals were sacrificed 14 days after Matrigel plug implantation. At the termination of the experiment, the Matrigel plugs from control and z-guggulsterone–treated mice were removed and fixed in 10% neutral-buffered formalin. The fixed Matrigel plugs from control and z-guggulsterone administered mice were dehydrated, embedded in paraffin, and sectioned at 4 μm of thickness. Sections from control and z-guggulsterone administered mice were used for immunohistochemical analysis of CD31, factor VIII, and VEGF-R2. Quantitative image analysis of the microvessel area based on CD31 and factor VIII immunostaining was done using Image Analysis software.

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In vivo Matrigel Plug Angiogenesis Assay: Male nude mice (5-6 weeks old) were randomized into control and treatment groups. Mice in the treatment group received oral gavage of (Z)-Guggulsterone at a dose of 1 mg per mouse (approximately 40 mg/kg body weight), dissolved in 0.1 mL phosphate-buffered saline (PBS), five times per week. Control mice received 0.1 mL of PBS vehicle. This pretreatment regimen continued for two weeks. Subsequently, Matrigel plugs containing 3 x 10^6 DU145 cells were implanted subcutaneously into the flank of each mouse. The oral administration of (Z)-Guggulsterone or vehicle continued for an additional two weeks post-implantation. Tumor volume was measured periodically using calipers. Fourteen days after plug implantation, mice were euthanized. The Matrigel plugs were excised, fixed in formalin, processed for paraffin embedding, and sectioned for immunohistochemical analysis.[1]

cited rat pharmacokinetic study (reference 41 in the paper) reported that the maximum plasma concentration of gucoustin was approximately 3.3 µM after a single oral dose of 50 mg/kg body weight. [1]
The same cited rat study reported that the oral bioavailability of (Z)-gucoustin was approximately 43%. [1]

In in vivo studies, oral administration of (Z)-glucostone (1 mg/mouse, approximately 40 mg/kg, 5 times a week for 4 weeks) to male nude mice did not cause significant changes in body weight compared to the carrier control group. The treated mice appeared healthy and showed no signs of discomfort, limited activity, indigestion, or local reactions such as redness or swelling. [1]
In vitro studies showed that concentrations of (Z)-glucostone up to 20 µM did not significantly affect the viability of human umbilical vein endothelial cells (HUVECs) within 24 hours. [1]

[1]. z-Guggulsterone, a constituent of Ayurvedic medicinal plant Commiphora mukul, inhibits angiogenesis in vitro and in vivo. Mol Cancer Ther. 2008 Jan;7(1):171-80.

[2]. FXR inhibition may protect from SARS-CoV-2 infection by reducing ACE2. Nature. 2023 Mar;615(7950):134-142.

Guggulsterone is a 3-hydroxysteroid with androgenic activity. It has been reported that Guggulsterone exists in Commiphora mukul and Commiphora wightii, with corresponding data. Our previous research showed that Guggulsterone, a component of the Indian Ayurvedic medicinal plant Commiphora mukul, can inhibit the growth of human prostate cancer cells by inducing apoptosis. We now report a novel effect of Guggulsterone: inhibition of angiogenesis in vitro and in vivo. Guggulsterone treatment inhibited capillary-like tubular formation in human umbilical vein endothelial cells (HUVECs) (in vitro angiogenesis) and the migration of HUVECs and DU145 human prostate cancer cells in a concentration- and time-dependent manner. The Z- and E-isomers of Guggulsterone appear to be equally potent inhibitors of HUVEC tubular formation. In vitro experiments showed that Z-glucosterone-mediated angiogenesis inhibition was associated with the inhibition of angiogenic growth factor secretion (e.g., vascular endothelial growth factor (VEGF) and granulocyte colony-stimulating factor), downregulation of VEGF receptor 2 (VEGF-R2) protein levels, and inactivation of Akt. Knockdown of VEGF-R2 protein levels enhanced Z-glucosterone-mediated inhibition of DU145 cell migration. Ectopic expression of constitutively active Akt in DU145 cells protected cells from Z-glucosterone-mediated cell migration inhibition. Gavage administration of 1 mg/day (five weeks) of Z-glucosterone to male nude mice inhibited in vivo angiogenesis in the DU145-Matrigel plug assay, as evidenced by statistically significant reductions in tumor burden, microvessel area (angiogenesis markers factor VIII and CD31 staining), and VEGF-R2 protein expression. In summary, this study demonstrates that z-guggostone inhibits angiogenesis by suppressing the VEGF-VEGF-R2-Akt signaling pathway. Our findings provide a strong theoretical basis for further preclinical and clinical studies of z-guggostone in the treatment of prostate cancer. [1]

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Prophylaxis against SARS-CoV-2 infection by modulating viral host receptors (e.g., angiotensin-converting enzyme 2 (ACE2)) may represent a novel approach to COVID-19 chemoprevention as a complement to vaccination [2,3]. However, the regulatory mechanisms of ACE2 expression remain unclear. This study demonstrates that the farnesoid X receptor (FXR) is a direct regulator of ACE2 transcription in various COVID-19-affected tissues, including the gastrointestinal and respiratory systems. We subsequently used the over-the-counter z-guggostone and the off-patent ursodeoxycholic acid (UDCA) to reduce FXR signaling, thereby downregulating ACE2 expression in human lung, bile duct cells, and intestinal organoids, as well as in corresponding tissues in mice and hamsters. The results showed that UDCA-mediated downregulation of ACE2 reduced susceptibility to SARS-CoV-2 infection in vitro, in vivo, and in vitro perfused human lungs and livers. In addition, we found that UDCA reduced ACE2 expression in human nasal epithelial cells. Finally, using retrospective registry data, we found a correlation between UDCA treatment and positive clinical outcomes after SARS-CoV-2 infection, and confirmed these findings in an independent liver transplant recipient validation cohort. In summary, we demonstrated that FXR plays a role in controlling ACE2 expression and provided evidence that modulating this pathway may help reduce SARS-CoV-2 infection, paving the way for future clinical trials. [2]

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Tumor angiogenesis (neovascularization) is a highly complex process regulated by a variety of pro-angiogenic growth factors and their corresponding receptors. Based on the results of this study, it is reasonable to conclude that inhibition of the VEGF-VEGF-R2-Akt signaling pathway may be an important mechanism by which Z-glucosterone exerts its anti-angiogenic effect. The following observations support this conclusion: (a) Guggulsterone-mediated inhibition of tubular formation and migration is associated with inhibition of VEGF secretion, which provides pro-survival signals to normal and tumor-derived endothelial cells via the receptor tyrosine kinase VEGF-R2; (b) Guggulsterone treatment downregulates VEGF-R2 protein levels; (c) knockdown of VEGF-R2 protein levels enhances Guggulsterone-mediated inhibition of DU145 cell migration; (d) Guggulsterone inhibits Akt in HUVEC and DU145 cells, and the inhibitory effect of the drug on HUVEC tubular formation is enhanced by pharmacological inhibition of Akt. However, the specific mechanisms by which Guggulsterone reduces VEGF secretion or downregulates VEGF-R2 protein levels remain unclear and require further investigation. In summary, this study demonstrates that Guggulsterone can inhibit angiogenesis both in vitro and in vivo. Z-Gigursteine-mediated angiogenesis inhibition is associated with Akt inactivation, inhibition of growth factor (VEGF and G-CSF), IL-17 and MMP-2 secretion, and downregulation of VEGF-R2 protein expression. [1] (Z)-Gigursteine is an over-the-counter phytosterol and an FXR antagonist. [2] This study found that FXR is a direct regulator of ACE2 transcription. (Z)-Gigursteine reduces ACE2 expression in various COVID-19-associated human cell types (respiratory, biliary, and intestinal) by inhibiting the FXR signaling pathway, thereby reducing the susceptibility of cells to SARS-CoV-2 infection in vitro. [2] The mechanism involves (Z)-Gigursteine binding to FXR, preventing FXR from binding to the FXR response element (IR-1) on the ACE2 promoter, leading to a reduction in ACE2 transcription. [2]
This study suggests that modulating host receptors such as ACE2 by inhibiting FXR is a potential host-targeted COVID-19 prevention strategy that may be less susceptible to viral escape mutations compared to direct antiviral strategies. [2]

C21H28O2

312.45

312.208

C, 80.73; H, 9.03; O, 10.24

39025-23-5

39025-23-5 (Z-Guggulsterone); 95975-55-6; 39025-24-6 (E-Guggulsterone)

6450278

White to off-white solid powder

1.1±0.1 g/cm3

463.3±45.0 °C at 760 mmHg

188-190°

172.3±25.7 °C

0.0±1.1 mmHg at 25°C

1.557

3.65

0

2

0

23

640

5

C/C=C/1\C(=O)C[C@H]2[C@@H]3CCC4=CC(=O)CC[C@]4(C)[C@H]3CC[C@]12C

WDXRGPWQVHZTQJ-OSJVMJFVSA-N

InChI=1S/C21H28O2/c1-4-16-19(23)12-18-15-6-5-13-11-14(22)7-9-20(13,2)17(15)8-10-21(16,18)3/h4,11,15,17-18H,5-10,12H2,1-3H3/b16-4+/t15-,17+,18+,20+,21-/m1/s1

(8R,9S,10R,13S,14S,17Z)-17-ethylidene-10,13-dimethyl-1,2,6,7,8,9,11,12,14,15-decahydrocyclopenta[a]phenanthrene-3,16-dione

Z-Guggulsterone; (Z)-Guggulsterone; Z-Guggulsterone; Guggulsterone; 39025-23-5; 95975-55-6; Guggulsterones Z; Cis-Guggulsterone; Guggulsterone E&Z; (Z)-Guggulsterone

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO: 5~10 mg/mL (16.0~32.0 mM)
Ethanol: ~2 mg/mL (~6.4 mM)

Solubility in Formulation 1: ≥ 1 mg/mL (3.20 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 10.0 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.

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Solubility in Formulation 2: 10 mg/mL (32.01 mM) in 50% PEG300 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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