KLF4 suppresses anticancer effects of brusatol via transcriptional upregulating NCK2 expression in melanoma.

Brusatol (Bru), a main extract from traditional Chinese medicine Brucea javanica, has been reported to exist antitumor effect in many tumors including melanoma. However, the underlying mechanism in its anti-melanoma effect still need further exploration. Here, we reported that the protein expression of KLF4 in melanoma cells were significantly downregulated in response to brusatol treatment. Overexpression of KLF4 suppressed brusatol-induced melanoma cell apoptosis; while knockdown of KLF4 enhanced antitumor effects of brusatol on melanoma cells not only in vitro but also in vivo. Further studies on the mechanism revealed that KLF4 bound to the promoter of NCK2 directly and facilitated NCK2 transcription, which suppressed the antitumor effect of brusatol on melanoma. Furthermore, our findings showed that miR-150-3p was dramatically upregulated under brusatol treatment which resulted in the downregulation of KLF4. Our results suggested that the miR-150-3p/KLF4/NCK2 axis might play an important role in the antitumour effects of brusatol in melanoma.

The Experimental and In Silico-Based Evaluation of NRF2 Modulators, Sulforaphane and Brusatol, on the Transcriptome of Immortalized Bovine Mammary Alveolar Cells.

Changes during the production cycle of dairy cattle can leave these animals susceptible to oxidative stress and reduced antioxidant health. In particular, the periparturient period, when dairy cows must rapidly adapt to the sudden metabolic demands of lactation, is a period when the production of damaging free radicals can overwhelm the natural antioxidant systems, potentially leading to tissue damage and reduced milk production. Central to the protection against free radical damage and antioxidant defense is the transcription factor NRF2, which activates an array of genes associated with antioxidant functions and cell survival. The objective of this study was to evaluate the effect that two natural NRF2 modulators, the NRF2 agonist sulforaphane (SFN) and the antagonist brusatol (BRU), have on the transcriptome of immortalized bovine mammary alveolar cells (MACT) using both the RT-qPCR of putative NRF2 target genes, as well as RNA sequencing approaches. The treatment of cells with SFN resulted in the activation of many putative NRF2 target genes and the upregulation of genes associated with pathways involved in cell survival, metabolism, and antioxidant function while suppressing the expression of genes related to cellular senescence and DNA repair. In contrast, the treatment of cells with BRU resulted in the upregulation of genes associated with inflammation, cellular stress, and apoptosis while suppressing the transcription of genes involved in various metabolic processes. The analysis also revealed several novel putative NRF2 target genes in bovine. In conclusion, these data indicate that the treatment of cells with SFN and BRU may be effective at modulating the NRF2 transcriptional network, but additional effects associated with cellular stress and metabolism may complicate the effectiveness of these compounds to improve antioxidant health in dairy cattle via nutrigenomic approaches.

Brusatol hinders the progression of bladder cancer by Chac1/Nrf2/SLC7A11 pathway.

Bladder cancer is a common tumor that impacts the urinary system and marked by a significant fatality rate and an unfavorable prognosis. Promising antineoplastic properties are exhibited by brusatol, which is obtained from the dried ripe fruit of Brucea javanica. The present study aimed to evaluate the influence of brusatol on the progression of bladder cancer and uncover the molecular mechanism involved. We used Cell Counting Kit-8, colony formation and EdU assays to detect cell numbers, viability and proliferation. We used transwell migration assay to detect cell migration ability. The mechanism of brusatol inhibition of bladder cancer proliferation was studied by flow cytometry and western blotting. It was revealed that brusatol could reduce the viability and proliferation of T24 and 5637 cells. The transwell migration assay revealed that brusatol was able to attenuate the migration of T24 and 5637 cells. We found that treatment with brusatol increased the levels of reactive oxygen species, malondialdehyde and Fe2+, thereby further promoting ferroptosis in T24 and 5637 cells. In addition, treatment with RSL3 (an agonistor of ferroptosis) ferrostatin-1 (a selective inhibitor of ferroptosis) enhanced or reversed the brusatol-induced inhibition. In vivo, treatment with brusatol significantly suppressed the tumor growth in nude mice. Mechanistically, brusatol induced ferroptosis by upregulating the expression of ChaC glutathione-specific gamma-glutamylcyclotransferase (Chac1) and decreasing the expression of SLC7A11 and Nrf2 in T24 and 5637 cells. To summarize, the findings of this research demonstrated that brusatol hindered the growth of bladder cancer and triggered ferroptosis via the Chac1/Nrf2/SLC7A11 pathway.

Ailanthone inhibits non-small cell lung cancer growth and metastasis through targeting UPF1/GAS5/ULK1 signaling pathway.

BACKGROUND: Targeting long non-coding RNAs (LncRNAs) is a novel and promising approach in cancer therapy. In our previous study, we investigated the effects of ailanthone (aila), the main active compound derived from the stem barks of Ailanthus altissima (Mill.) Swingle, on the growth of non-small cell lung cancer (NSCLC) cells. Although we observed significant inhibition of NSCLC cell growth of aila, the underlying mechanisms involving LncRNAs, specifically LncRNA growth arrest specific 5 (GAS5), remain largely unknown. METHODS: To further explore the impact of aila on NSCLC, we performed a series of experiments. Firstly, we confirmed the inhibitory effect of aila on NSCLC cell growth using multiple assays, including MTT, wound healing, transwell assay, as well as subcutaneous and metastasis tumor mice models in vivo. Next, we utilized cDNA microarray and RT-QPCR to identify GAS5 as the primary target of aila. To verify the importance of GAS5 in aila-induced tumor inhibition, we manipulated GAS5 expression levels by constructing GAS5 over-expression and knockdown NSCLC cell lines. Furthermore, we investigated the upstream and downstream signaling pathways of GAS5 through western blot and RT-QPCR analysis. RESULTS: Our results showed that aila effectively increased GAS5 expression, as determined by microarray analysis. We also observed that aila significantly enhanced GAS5 expression in a dose- and time-dependent manner across various NSCLC cell lines. Notably, over-expression of GAS5 led to a significant suppression of NSCLC cell tumor growth; while aila had minimal inhibitory effect on GAS5-knockdown NSCLC cells. Additionally, we discovered that aila inhibited ULK1 and autophagy, and this inhibition was reversed by GAS5 knockdown. Moreover, we found that aila up-regulated GAS5 expression by suppressing UPF1-mediated nonsense-mediated mRNA decay (NMD). CONCLUSION: In summary, our findings suggest that aila promotes GAS5 expression by inhibiting UPF1-mediated NMD, leading to the repression of ULK1-mediated autophagy and subsequent inhibitory effects on NSCLC cells. These results indicate that aila is a potent enhancer of GAS5 and holds promising potential for application in NSCLC therapy. However, our research is currently focused only on NSCLC. It remains to be determined whether aila can also inhibit the growth of other types of tumors through the UPF1/GAS5/ULK1 signaling pathway. In future studies, we can further investigate the mechanisms by which aila suppresses other types of tumors and potentially broaden the scope of its application in cancer therapy.

Bruceantinol targeting STAT3 exerts promising antitumor effects in in vitro and in vivo osteosarcoma models.

Bruceantinol (BOL) is a quassinoid compound found in the fruits of Brucea javanica. Previous research has highlighted the manifold physiological and pharmacological activities of BOL. Notably, BOL has demonstrated antitumor cytotoxic and antibacterial effects, lending support to its potential as a promising therapeutic agent for various diseases. Despite being recognized as a potent antitumor inhibitor in multiple cancer types, its efficacy against osteosarcoma (OS) has not been elucidated. In this work, we investigated the antitumor properties of BOL against OS. Our findings showed that BOL significantly decreased the proliferation and migration of OS cells, induced apoptosis, and caused cell death without affecting the cell cycle. We further confirmed that BOL potently suppressed tumor growth in vivo. Mechanismly, we discovered that BOL directly bound to STAT3, and prevent the activation of STAT3 signaling at low nanomolar concentrations. Overall, our study demonstrated that BOL potently inhibited the growth and metastasis of OS, and efficiently suppressed STAT3 signaling pathway. These results suggest that BOL could be a promising therapeutic candidate for OS.

Bruceine A: Suppressing metastasis via MEK/ERK pathway and invoking mitochondrial apoptosis in triple-negative breast cancer.

Triple-negative breast cancer (TNBC), as the most aggressive subtype of breast cancer, presents a scarcity of miraculous drugs in suppressing its proliferation and metastasis. Bruceine A (BA) is a functional group-rich quassin compound with extensive and distinctive pharmacological activities. Within the present study, we investigated the capabilities of BA in suppressing TNBC proliferation and metastasis as well as its potential mechanisms. The results displayed that BA dramatically repressed the proliferation of MDA-MB-231 and 4T1 cells with corresponding IC50 values of 78.4 nM and 524.6 nM, respectively. Concurrently, BA arrested cells in G1 phase by downregulating cycle-related proteins Cyclin D1 and CDK4. Furthermore, BA distinctly induced mitochondrial dysfunction as manifested by diminished mitochondrial membrane potential, elevated reactive oxygen species generation, minimized ATP production, and Caspase-dependent activation of the mitochondrial apoptosis pathway. Additionally, BA restrained the invasion and metastasis of TNBC cells by repressing MMP9 and MMP2 expression. Intriguingly, after pretreatment with MEK activator C16-PAF, the inhibitory effect of BA on MEK/ERK pathway was notably diminished, while the proliferation suppression and metastasis repression exerted by BA were all strikingly curtailed. Molecular docking illustrated that BA potently combined with residues on the MEK1 protein with the presence of diverse intermolecular interactions. Ultimately, BA effectively suppressed tumor growth in the 4T1 xenograft tumor model with no detectable visceral toxicity in the high-dose group and, astonishingly, repressed tumor metastasis in the 4T1-luc lung metastasis model. Collectively, our study demonstrates that BA is a promising chemotherapeutic agent for treating TNBC and suppressing lung metastasis.

Quassinoid analogs exert potent antitumor activity via reversible protein biosynthesis inhibition in human colorectal cancer.

Cellular protein synthesis is accelerated in human colorectal cancer (CRC), and high expression of protein synthesis regulators in CRC patients is associated with poor prognosis. Thus, inhibition of protein synthesis may be an effective therapeutic strategy for CRC. We previously demonstrated that the quassinoid bruceantinol (BOL) had antitumor activity against CRC. Herein, potent tumor growth suppression (>80%) and STAT3 inhibition was observed in two different mouse models following BOL administration. Loss of body and spleen weight was observed but was eliminated upon nanoparticle encapsulation while maintaining strong antitumor activity. STAT3 siRNA knockdown exhibited modest suppression of cell proliferation. Surprisingly, STAT3 inhibition using a PROTAC degrader (SD-36) had little effect on cancer cell proliferation suggesting the possibility of additional mechanism(s) of action for quassinoids. BOL-resistant (BR) cell lines, HCT116BR and HCA7BR, were equally sensitive to standard CRC therapeutic agents and known STAT3 inhibitors but resistant to homoharringtonine (HHT), a known protein synthesis inhibitor. The ability of quassinoids to inhibit protein synthesis was dependent on the structure of the C15 sidechain. Of note, BOL did not inhibit protein synthesis in normal human colon epithelial cells whereas HHT and napabucasin remained effective in these normal cells. Novel quassinoids were designed, synthesized, and evaluated in pre-clinical CRC models. Treatment with the most potent analog, 5c, resulted in significant inhibition of cell proliferation and protein synthesis at nanomolar concentrations. These quassinoid analogs may represent a novel class of protein synthesis inhibitors for the treatment of human CRC.

Quassinoids from Eurycoma longifolia Jack roots and their potential inhibitory activity against human benign prostatic hyperplasia cells (BPH-1) and testosterone-induced BPH rat model.

Benign prostate hyperplasia (BPH) is an enlargement of the prostate gland, because of hormonal changes in aging males which contribute significantly to excessive proliferation over apoptosis of prostatic cells. The anti-proliferative and induced apoptotic activities of Eurycoma longifolia quassinoids on cancer cell lines could be promising therapeutic targets on BPH. Hitherto, no report of the quassinoids against BPH problem was available. In this study, a systematic phytochemical fractionation of the root extract, TAF2 was performed, which led to the discovery of nine previously described C20 quassinoids (1-9). Two undescribed C20 (10 and 12) and one undescribed (11) C19 quassinoids were identified by detailed NMR and HR-ESI-MS data analysis. Their absolute configurations were assigned by ECD spectral analysis. The quassinoids (1-12) were tested for inhibitory activity against the proliferation of human BPH-1 and human skin Hs27 fibroblast cells cultured in vitro. 1, 2 and 3 at 10 µM significantly reduced BPH-1 cell viability and were cytotoxic to Hs27 fibroblast cells. 2 was selected for further study of anti-BPH activity against testosterone induced BPH rats. At 5 mg/kg, 2 reduced the rat prostatic weight and prostatic index, consistent with the decrease in papillary acini number and epithelial thickness of the prostate tissues. These quassinoids may be potential anti-BPH compounds that require further studies.

ASFV transcription reporter screening system identifies ailanthone as a broad antiviral compound.

African swine fever (ASF) is an acute, highly contagious and deadly viral disease in swine that jeopardizes the worldwide pig industry. Unfortunately, there are no authoritative vaccine and antiviral drug available for ASF control. African swine fever virus (ASFV) is the etiological agent of ASF. Among the ASFV proteins, p72 is the most abundant component in the virions and thus a potential target for anti-ASFV drug design. Here, we constructed a luciferase reporter system driven by the promoter of p72, which is transcribed by the co-transfected ASFV RNA polymerase complex. Using this system, we screened over 3200 natural product compounds and obtained three potent candidates against ASFV. We further evaluated the anti-ASFV effects and proved that among the three candidates, ailanthone (AIL) inhibits the replication of ASFV at the nanomolar concentration (IC50 â€‹= â€‹15 â€‹nmol/L). Our in vitro experiments indicated that the antiviral effect of AIL is associated with its inhibition of the HSP90-p23 cochaperone. Finally, we showed the antiviral activity of AIL on Zika virus and hepatitis B virus (HBV), which supports that AIL is a potential broad-spectrum antiviral agent.

Quassinoids from Eurycoma longifolia with antiviral activities by inhibiting dengue virus replication.

BACKGROUND: Dengue caused by dengue virus (DENV) spreads rapidly around the world. However, there are no worldwide licensed vaccines or specific antivirals to combat DENV infection. Quassinoids are the most characteristic components of Eurycoma longifolia, which have been reported to display a variety of biological activities. However, whether quassinoids exert anti-DENV activities remains unknown. PURPOSE: To test the quassinoids of E. longifolia for their activity against DENV and to clarify the potential mechanisms. METHODS: The quassinoids from E. longifolia were isolated by chromatography techniques, and their chemical structures were elucidated by spectroscopic analysis. The anti-DENV activities of quassinoids on baby hamster kidney cells BHK-21 were determined by lactate dehydrogenase (LDH) assay. The synthesis of progeny virus was measured by plaque assay. The expression levels of envelope protein (E) and non-structural protein 1 (NS1) were evaluated by qRT-PCR, Western blot and immunofluorescence assays. Molecular docking was used to screen the potential targets of the most active quassinoid against DENV-2, and surface plasmon resonance analysis was employed to confirm the direct binding between the most active quassinoid and potential target. RESULTS: Twenty-four quassinoids, including three new quassinoids (1 - 3), were isolated from the ethanol extract of E. longifolia. Quassinoids 4, 5, 9, 11, 12, 15, 16, 17, 19 and 20 significantly reduced the LDH release at the stages of viral binding and entry or intracellular replication. Among them, 19 (6α-hydroxyeurycomalactone, 6α-HEL) exhibited the best anti-DENV-2 activities with an EC50 value of 0.39 ± 0.02 µM. Further experiments suggested that 6α-HEL remarkably inhibited progeny virus synthesis and mRNA and protein expression levels of E and NS1 of DENV-2. Time-of-drug-addition assay suggested that 6α-HEL inhibited intracellular replication of DENV-2 at an early stage. Moreover, 6α-HEL was shown to interact with NS5-RdRp domain at a binding affinity of -8.15 kcal/mol. SPR assay further verified 6α-HEL bound to RdRp protein with an equilibrium dissociation constant of 1.49 × 10-7 M. CONCLUSION: Ten quassinoids from E. longifolia showed anti-DENV activities at processes of virus binding and entry or intracellular replication. The most active quassinoid 6α-HEL exerts the anti-DENV-2 activities at intracellular replication stage by directly targeting the NS5-RdRp protein. These results suggest that 6α-HEL could be a promising candidate for the treatment of DENV-2 infection.

Brusatol: A potential sensitizing agent for cancer therapy from Brucea javanica.

Cancer is currently the most important problem endangering human health. As antitumor drugs have always been the most common methods for treating cancers, searching for new antitumor agents is of great significance. Brusatol, a quassinoid from the seeds of Brucea javanica, exhibits a potent tumor-suppressing effect with improved disease outcome. Studies have shown that brusatol not only shows potential tumor inhibition through multiple pharmacological effects, such as promoting apoptosis and inhibiting metastasis but also exhibits significant synergistic antitumor effects in combination with chemotherapeutic agents and overcoming chemical resistance in a wide range of cancer types. In this paper, the antitumor effects and mechanisms of brusatol were reviewed to provide evidence that brusatol has the exact antitumor efficacy of chemotherapeutic agents and show the potential of brusatol to be developed as a promising antitumor drug.

Quassinoids from Picrasma chinensis with Insecticidal Activity against Adults and Larvae of Diaphorina citri Kuwayama and Neuroprotective Effect.

Eleven new tetracyclic quassinoids, picrachinensin A-K (1-11), along with six known congeners, were isolated from the stems and leaves of Picrasma chinensis. Their structures were elucidated by integrated multiple spectroscopic techniques, single-crystal X-ray diffraction analysis, and electronic circular dichroism. Notably, compounds 3 and 4 are a pair of undescribed epimers, and 8 and 9 are unusual quassinoids with a hydroxymethyl group at C-13. Biologically, compound 7 exhibited insecticidal activity on both adults and larvae of Diaphorina citri Kuwayama even more effectively than the positive control (abamectin), with an LD50 of 55.69 mg/L for adults and a corrected mortality rate of 30.42 ± 2.78% for larvae (100 mg/L). According to preliminary structure-activity relationship investigations, the hydroxymethyl at the C-13 position of quassinoids was beneficial for their insecticidal activity. In addition, compounds 1, 4, and 12 exhibited excellent neuroprotective effect against H2O2-induced oxidative injury on SH-SY5Y cells, with more potent activity than the positive control (Trolox), and all the compounds exhibited no cytotoxicity to SH-SY5Y and BV-2 cells at the indicated concentrations.

Quassinoids from Eurycoma longifolia and their bone formation evaluation in zebrafish, C3H10 cells and silico.

Phytochemicals with bone formation potential in traditional medicines captured more and more attentions due to their advantages to bone loss and fewer side effects. As a famous aphrodisiac phytomedicine, Eurycoma longifolia (EL) has acquired general recognition in improving male sexual health, and thus been considered as traditional medicine for the treatment of androgen-deficient osteoporosis. Although the aqueous extract of EL had been proved to be beneficial to bone loss, the active constituents and the mechanisms underlying the effects are still obscure. The current study performed a chemical investigation on the roots of EL, which resulted in the isolation and identification of ten quassinoids (EL-1-EL-10), and then conducted their osteogenic activity evaluations in vivo zebrafish model with or without dexamethasone (Dex) and in vitro C3H10 cell model. The result displayed that most tested concentrations of EL-1-EL-5 could significantly increase the mineralization areas and integrated optical densities (IODs) of skull in both zebrafish model. The majority tested concentrations of EL-1-EL-5 could also improve the mRNA expression of early osteogenic associated genes ALPL, Runx2a, Sp7 in zebrafish model without Dex, but only a few could accelerate the mRNA expression of late osteogenic associated genes OCN. These results suggested the ability of EL-1-EL-5 to increase bone formation mainly by accelerating osteogenic differentiation at the early stage. The structure-based virtual screening based on the pharmacophores in ePharmaLib, as well as the molecular docking study, implied that the effects of the quassinoids (EL-1-EL-5) on the enhancement of bone formation might be related with improving the content and the activity of androgen through binding with CYP19A, SHBG and AKR1C2, and activating bone metabolism-related ANDR target genes and signal pathways by combining with ANDR directly. Although the assumptions are in silico model-based and further in vitro and in vivo validations are still necessary, we provided a new perspective to explore the potential of EL to be used as an alternative treatment for not only androgen-deficient osteoporosis, but also estrogen-deficient bone loss, by combining with SHBG.

Bruceine D may affect the phenylpropanoid biosynthesis by acting on ADTs thus inhibiting Bidens pilosa L. seed germination.

Bruceine D is a natural quassinoid, which was successfully isolated in our research group from the residue of Brucea javanica (L.) seeds. Our previous research showed that Bruceine D prevented Bidens pilosa L. seed germination by suppressing the activity of key enzymes and the expression levels of key genes involved in the phenylpropanoid biosynthesis pathway. In this study, integrated analyses of non-targeted metabolomic and transcriptomic were performed. A total of 356 different accumulated metabolites (DAMs) were identified, and KEGG pathway analyses revealed that most of these DAMs were involved in phenylpropanoid biosynthesis. The decreased expression of ADTs and content of L-phenylalanine implicates that Bruceine D may suppress the downstream phenylpropanoid biosynthesis pathway by disrupting primary metabolism, that is, the phenylalanine biosynthesis pathway, thus inhibiting the final products, resulting in the interruption of B. pilosa seed germination. These results suggest that Bruceine D may inhibit the B. pilosa seed germination by suppressing phenylpropanoid biosynthesis through acting on ADTs.

A Quassinoid Diterpenoid Eurycomanone from Eurycoma longifolia Jack Exerts Anti-Cancer Effect through Autophagy Inhibition.

Eurycomanone (EN) is one of the representative quassinoid diterpenoids from roots of Eurycoma longifolia Jack, a natural medicine that is widely distributed in Southeast Asia. Previous studies showed that EN induces cancer cell apoptosis and exhibits anti-cancer activity, but the molecular mechanism of EN against cancer has still not been elucidated. In this study, we examined the regulatory effect of EN on autophagy to reveal the mechanism of EN-mediated colon cancer growth inhibition. First, we found that EN is able to inhibit colon cancer cell proliferation and colony formation. The angiogenesis level in cancer cells was inhibited as well. Next, the treatment of EN led to the suppression of autophagy, which was characterized by the downregulation of the LC3-II level and the formation of GFP-LC3 puncta under EN treatment in colon cancer. Moreover, we revealed that the mTOR signaling pathway was activated by EN in a time- and concentration-dependent manner. Finally, autophagy induction protected colon cancer cells from EN treatment, suggesting that autophagy improves cell survival. Taken together, our findings revealed the mechanism of EN against colon cancer through inhibiting autophagy and angiogenesis in colon cancer, supporting that the autophagy inhibitor EN could be developed to be a novel anti-cancer agent.

Brusatol suppresses the growth of intrahepatic cholangiocarcinoma by PI3K/Akt pathway.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is a malignancy with a hidden onset, high metastasis recurrence rate, and poor prognosis. Research on effective drugs for ICC is important for improving the prognosis of patients in the clinic. Brusatol is a quassinoid extracted from the seeds of Brucea sumatrana and has been shown to have the potential to inhibit tumor metastasis and proliferation. There has been no scientific research on the therapeutic effect of brusatol on ICC. Our study offers a novel strategy for the therapy of ICC. PURPOSE: Explore effects of brusatol treatment on ICC and clarify the possible mechanism. STUDY DESIGN: Various cell functional experiments and basic experimental techniques were applied to ICC cell lines to explore the influences of brusatol on ICC cells; this conclusion was further verified in animal models. METHODS: The anti-cancer effects of the drug on the cell, protein, and RNA level were verified by cell functional experiments, WB blotting and transcriptome sequencing experiments, respectively. Finally, the experimental results were verified using subcutaneous tumor experiments in nude mice. RESULTS: The consequences exhibited that the levels of epithelial markers of ICC cells increased after brusatol treatment, and the levels of interstitial indicators decreased, suppressing the epithelial-mesenchymal transition (EMT) process. Brusatol inhibited proliferation, induced apoptosis, and suppressed the migration and invasion abilities of Hucc-T1 and RBE oncocytes via activating PI3K/Akt pathway. It also suppressed the growth of Hucc-T1 xenografts in nude mice. CONCLUSION: Brusatol inhibits the proliferation and EMT process in ICC oncocytes by the PI3K/Akt pathway and promotes apoptosis in oncocytes.

Brusatol Inhibits Proliferation and Metastasis of Colorectal Cancer by Targeting and Reversing the RhoA/ROCK1 Pathway.

Brusatol (BRU) is an important compound extracted from Brucea javanica oil, whose pharmacological effects are able to induce a series of biological effects, including inhibition of tumor cell growth, anti-inflammatory, antiviral, and antitumor. Currently, there are so few studies about the brusatol effects on colorectal cancer that its anticancer mechanism has not been clearly defined. In this study, we made an in-depth investigation into the brusatol effect towards the proliferation and metastasis of colon cancer and the possible mechanism. The inhibitory effect of BRU on the proliferation of colorectal cancer cells was unveiled via CCK-8 method and colony formation assay, while the inhibitory effect of BRU on migration and invasion of colorectal cancer cells was revealed by scratch assay and transwell assay. In addition, Western blot results also revealed that BRU inhibited not only the expressions of RhoA and ROCK1 but also the protein expressions of EMT-related markers e-cadherin, N-cadherin, Vimentin, MMP2, and MMP9 in colon cancer cells. Through the xenotransplantation model, our in vivo experiment further verified the antitumor effect of BRU on colon cancer cells in vitro, and the results were consistent with the protein expression trend. In conclusion, BRU may inhibit the proliferation and metastasis of colorectal cancer by influencing EMT through RhoA/ROCK1 pathway.

A New C22-Quassinoid with Anti-Pancreatic Adenocarcinoma Activity from Seeds of Brucea javanica.

An undescribed C22-quassinoid named sergeolide A (1) and fifteen known quassinoids (2-16) were obtained from the seeds of Brucea javanica (Simaroubaceae). All chemical structures were established based on spectroscopic data and X-ray diffraction analysis. Sergeolide A (1) is the first example of a naturally occurring C22-quassinoid bearing a butenolide group fused the A ring of the bruceolide skeleton from Brucea genus. And this is the first report of the NMR data for desmethyl-bruceines B (2) and C (3) and the crystal structure for bruceolide (11). In addition, all isolates were evaluated for their anti-pancreatic adenocarcinoma activity by measuring the growth inhibitory of the MIA PaCa-2 cell lines. Consequently, compounds 1, 7-10, and 12-16 exhibited potent anti-pancreatic cancer activity in vitro (IC50 =0.054∼0.357 µM).

Bruceine A protects against diabetic kidney disease via inhibiting galectin-1.

Bruceine A is a natural quassinoid compound extracted from the fruit of the Traditional Chinese Medicine Brucea javanica (L.) Merr. that has various types of various biological activities. However, whether the compound has a protective effect on diabetic kidney disease remains unknown. Galectin-1 is actively involved in a variety of chronic inflammation-relevant human diseases including diabetic kidney disease. Here, we identified Bruceine A as a kidney protective molecule against a model of diabetic kidney disease in db/db mice with potent anti-inflammatory activity both in vitro and in vivo. Mechanistically, by selectively binding to the conserved carbohydrate-recognition domain of galectin-1 and disrupting the interaction between galectin-1 and the receptor for activated protein C kinase 1, Bruceine A was found to inhibit galectin-1-mediated inflammatory signal transduction under high glucose stress in rat mesangial HBZY-1 cells. Thus, our findings reveal Bruceine A as an unidentified galectin-1 inhibitor affording significant protection against diabetic kidney disease and may provide novel pharmacological therapeutics for the disease.

Brucein D augments the chemosensitivity of gemcitabine in pancreatic cancer via inhibiting the Nrf2 pathway.

BACKGROUND: Gemcitabine (GEM) is the first-line chemotherapeutic drug used to treat pancreatic ductal adenocarcinoma carcinoma (PDAC), but chemoresistance is often encountered clinically. Nrf2, an oxidative stress responsive transcription factor, is an important contributor to chemoresistance and poor prognosis of PDAC. Brucein D (BD), a naturally occurring quassinoid, has been reported to exert anti-tumor effect in several cancers including PDAC. In this study, we aimed to investigate the efficacy of BD and the role of Nrf2 axes on the chemosensitivity of GEM and elucidate the underlying molecular mechanisms. METHODS: Analyses of clinical samples of PDAC and GEPIA database were first conducted to identify the expression of Nrf2 in PDAC. We then established cell lines with stable deletion of Nrf2 through transfecting lentivirus into PDAC cells. Quantitative real-time PCR (qRT-PCR) and Western blotting were performed to determine the expression of Nrf2 in these cell lines. The effects of BD and Nrf2 axes on PDAC cell proliferation, colony-formation, tumor growth and chemosensitivity were determined both in vitro and in vivo. Orthotopic xenograft and genetically engineered KPC mouse models of PDAC were used to evaluate the anti-pancreatic cancer effects of BD and GEM. RESULTS: Nrf2 was highly expressed in PDAC in the clinical samples and GEPIA analysis. Gain- and lost-function study demonstrated that Nrf2 affected the chemosensitivity of GEM on PDAC cells both in vitro and in vivo. We further found that BD effectively inhibited PDAC cell proliferation and enhanced the chemosensitivity of GEM. Mechanistic studies revealed that BD sensitized GEM in PDAC cells through the ubiquitin-proteasome-dependent degradation of Nrf2, and downregulating the Nrf2 pathway. Silencing of Nrf2 plus BD treatment resulted in more potent inhibitory effects of GEM. In contrast, Nrf2 activation attenuated the chemosensitivity of GEM, indicating that the action of BD was Nrf2 dependent. Finally, the efficacy of BD alone and in combination with GEM on PDAC was validated on both orthotopic xenograft and genetically engineered KPC mouse models. CONCLUSIONS: BD was able to enhance the chemosensitivity of GEM in PDAC through inhibition of the Nrf2 pathway. Our experimental findings indicate that BD, a potent Nrf2 inhibitor, holds promise for further development into a novel adjuvant therapy for PDAC.